Eubacter

Biofactors, 1993 May, 4(2), 95 - 104
Hypusine: its post-translational formation in eukaryotic initiation factor 5A and its potential role in cellular regulation; Park MH et al.; The amino acid, hypusine {N epsilon-(4-amino-2-hydroxybutyl) lysine}, a unique component of one cellular protein, eukaryotic translation initiation factor 5A (eIF-5A, old terminology eIF-4D), is formed post-translationally in two enzymatic steps: (i) transfer of the 4-aminobutyl moiety of the polyamine spermidine to the epsilon-amino group of a single specific lysine residue in the eIF-5A precursor protein to form an intermediate, deoxyhypusine, and (ii) subsequent hydroxylation in this 4-aminobutyl portion . Hypusine is produced soon after the translation of eIF-5A mRNA; the modification is essentially irreversible . Hypusine is found in all eukaryotes examined as well as in archaebacteria; it does not occur in eubacteria . The protein containing hypusine from each species displays a high degree of amino acid identity; the sequence of amino acids surrounding the hypusine residue is strictly conserved, suggesting the importance of the hypusine modification throughout evolution . Expression of one of the two yeast eIF-5A genes is required for survival and the lysine codon at the site of hypusine synthesis is vital for yeast growth . The precise cellular function of eIF-5A remains to be elucidated; however, eIF-5A stimulates methionyl-puromycin synthesis in a model assay for translation initiation and eIF-5A precursors containing lysine in place of hypusine are inactive in this assay . This provides evidence that the hypusine modification is needed for eIF-5A activity . In view of the important role of hypusine in eIF-5A and because of the narrow specificities of the enzymes involved in formation of this unusual amino acid, the hypusine biosynthetic steps offer promising targets for intervention in cellular proliferation . Spermidine analogs that are inhibitors of deoxyhypusine synthase in vitro also cause inhibition of hypusine formation in cells, together with a reduction in protein synthesis and in cell growth . In addition, certain metal chelating inhibitors of deoxyhypusine hydroxylase exhibit anti-proliferative effects by arresting mammalian cells at the G1/S boundary of the cell cycle . These results lay the foundation for the potential regulation of cellular events through the application of specific and potent inhibitors of hypusine biosynthesis.

Plant Physiol, 1993 May, 102(1), 155 - 63
Treatment of pea (Pisum sativum L.) protoplasts with DNA-damaging agents induces a 39-kilodalton chloroplast protein immunologically related to Escherichia coli RecA; Cerutti H et al.; Organisms must have efficient mechanisms of DNA repair and recombination to prevent alterations in their genetic information due to DNA damage . There is evidence for DNA repair and recombination in plastids of higher plants, although very little is known at the biochemical level . Many chloroplast proteins are of eubacterial ancestry, suggesting that the same could be true for the components of a DNA repair and recombination system . A 39-kD protein, immunologically related to Escherichia coli RecA, is present in chloroplasts of pea (Pisum sativum L.) . Bandshift gel assays suggest that it binds single-stranded DNA . Its steady-state level is increased by several DNA-damaging agents . These results are consistent with it being a plastid homolog of E . coli RecA protein, presumably involved in DNA repair and recombination, and with the existence of an SOS-like response in pea leaf cells . Experiments with protein synthesis inhibitors suggest that the 39-kD chloroplast protein is encoded in the nucleus.

FEMS Microbiol Lett, 1993 Apr 15, 108(3), 255 - 8
Aspartame as a source of essential phenylalanine for the growth of oral anaerobes; Wyss C; Phenylalanine and aspartic acid requirements were determined for 13 species of oral bacteria using the chemically defined medium OMIZ-W1 . None of Actinobacillus actinomycetemcomitans, Bacteroides forsythus, Eikenella corrodens, Selenomonas sputigena, Treponema pectinovorum, T . socranskii, or Wolinella recta required either of these amino acid constituents of aspartame (L-aspartyl-L-phenylalanine methylester) . Phenylalanine was essential for the growth of Capnocytophaga gingivalis, Eubacterium timidum, Fusobacterium nucleatum, Porphyromonas gingivalis, T . denticola, and T . vincentii, while aspartic acid was not required . With the exception of E . timidum, all phenylalanine-dependent strains could grow when the free amino acid was replaced by aspartame at concentrations at least 10-fold lower than those used for aspartame as an artificial sweetener.

Proc Natl Acad Sci U S A, 1993 Apr 15, 90(8), 3511 - 5
Heterogeneity of the principal sigma factor in Escherichia coli: the rpoS gene product, sigma 38, is a second principal sigma factor of RNA polymerase in stationary-phase Escherichia coli; Tanaka K et al.; The rpoS gene of Escherichia coli encodes a putative RNA polymerase sigma factor that is considered to be the central regulator of gene expression in stationary phase . The gene product (sigma 38) was overproduced using the cloned rpoS gene and purified to homogeneity . Reconstituted RNA polymerase holoenzyme (E sigma 38) was found to recognize in vitro a number of typical sigma 70-type promoters, including the lacUV5 and trp promoters . Some, however, were recognized exclusively or preferentially by E sigma 70, whereas at least one, fic, was favored by E sigma 38 . Thus E . coli promoters can be classified into three groups: the first group is recognized by E sigma 70 and E sigma 38, but the second and third groups are recognized substantially by either E sigma 70 or E sigma 38 alone . In contrast to other minor sigma factors, sigma 38 shares a set of amino acid sequences common among the principal sigma factors of eubacteria and is therefore a member of the RpoD-related protein family . The intracellular level of sigma 38 was demonstrated to increase in vivo upon entry into stationary phase . These results together indicate that sigma 38 is a second principal sigma factor in stationary-phase E . coli.

Mol Microbiol, 1993 Apr, 8(1), 1 - 4
Mitochondrial transcription: is a pattern emerging?
Jaehning JA.
Despite the striking similarities of RNA polymerases and transcription signals shared by eubacteria, archaebacteria and eukaryotes, there has been little indication that transcription in mitochondria is related to any previously characterized model . Only in yeast has the subunit structure of the mitochondrial RNA polymerase been determined . The yeast enzyme is composed of a core related to polymerases from bacteriophage T7 and T3, and a promoter recognition factor similar to bacterial sigma factors . Soluble systems for studying mitochondrial transcript initiation in vitro have been described from several organisms, and used to determine consensus sequences at or near transcription start sites . Comparison of these sequences from fungi, plants, and amphibians with the T7/T3 promoter suggests some intriguing similarities . Mammalian mitochondrial promoters do not fit this pattern but instead appear to utilize upstream sites, the target of a transcriptional stimulatory factor, to position the RNA polymerase . The recent identification of a possible homologue of the mammalian upstream factor in yeast mitochondria may indicate that a pattern will eventually be revealed relating the transcriptional machineries of all eukaryotic mitochondria.

Int J Syst Bacteriol, 1993 Apr, 43(2), 302 - 4
Eubacterium saphenum sp . nov., isolated from human periodontal pockets {corrected}; Uematsu H et al.; A new species, Eubacterium saphenum {corrected} sp . nov., established on the basis of the results of DNA-DNA hybridization, was proposed for strains isolated from human periodontal pockets . Differential characteristics are given.

J Clin Microbiol, 1993 Apr, 31(4), 1001 - 2
UV red fluorescence of Eubacterium lentum; Mosca A et al.; Twenty-nine clinical isolates of Eubacterium lentum and two type species were evaluated for the ability to fluoresce under UV light . Twenty-one of the 29 isolates and both of the reference strains showed orange-to-red fluorescence . This fluorescence did not require blood or hemin in the culture media and did not fade upon air exposure . The fluorescent pigment, after extraction by 1 N NaOH, showed peak excitation at a wavelength of around 400 nm . The capacity of E . lentum to produce fluorescence may be a useful and time-sparing laboratory aid for its identification.

J Bacteriol, 1993 Apr, 175(7), 1871 - 8
Eubacterial components similar to small nuclear ribonucleoproteins: identification of immunoprecipitable proteins and capped RNAs in a cyanobacterium and a gram-positive eubacterium; Kovacs SA et al.; Small nuclear ribonucleoprotein (snRNP) particles play an important role in the processing of pre-mRNA . snRNPs have been identified immunologically in a variety of cells, but none have ever been observed in prokaryotic systems . This report provides the first evidence for the presence of snRNP-like components in two types of prokaryotic cells: those of the cyanobacterium Synechococcus leopoliensis and those of the gram-positive eubacterium Bacillus subtilis . These components consist of snRNP-immunoreactive proteins and RNAs, including some with the snRNP-unique 5' m2,2,7G (m3G) cap . Immunoreactivity was determined by immunoprecipitation procedures, with either antinuclear-antibody-positive (RNP- and Sm-monospecific) patient sera or a m3G monoclonal antibody, with radiolabelled cell extracts that were preadsorbed with antinuclear-antibody-negative sera . S . leopoliensis immunoprecipitates showed the presence of high-molecular-mass proteins (14 to 70 kDa) and RNAs (138 to 243 nucleotides) that are analogous in size to proteins and RNAs found in human (HEp-2) cell immunoprecipitates but absent in Escherichia coli immunoprecipitates . Thin-layer chromatography of S . leopoliensis immunoprecipitates confirmed the presence of a capped nucleotide similar to a capped nucleotide in HEp-2 immunoprecipitates; no such nucleotide was observed in E . coli immunoprecipitates . Immunoreactive RNAs (117-170 nucleotides) were identified in a second eubacterium, B . subtilis, as well . This work suggests that snRNPs or their evolutionary predecessors predate the emergence of eukaryotic cells.

Oral Microbiol Immunol, 1993 Apr, 8(2), 105 - 10
Identification of Actinobacillus actinomycetemcomitans: polymerase chain reaction amplification of lktA-specific sequences; Goncharoff P et al.; Actinobacillus actinomycetemcomitans has been strongly implicated in the etiology of localized juvenile periodontitis . Techniques used in the identification of this periodontal pathogen include cultural, biochemical, immunological and DNA hybridization analysis . In this study, we report the use of polymerase chain reaction (PCR) to amplify unique sequences of A . actinomycetemcomitans . Specific oligonucleotide primers LKT2 and LKT3 were designed to hybridize to the A . actinomycetemcomitans lktA gene, which encodes leukotoxin, a putative A . actinomycetemcomitans virulence factor . The LKT2 and LKT3 primers amplified lktA-specific sequences from all 12 A . actinomycetemcomitans strains tested . In another set of experiments, 13 other bacterial species, most of which are normal residents of the oral cavity, were tested with these primers . These PCR amplifications also contained 2 additional primers, RRN4 and RRN5, which served as positive controls; RRN4 and RRN5 were designed to amplify specific sequences of eubacterial 16S ribosomal DNA (rDNA) . PCR amplifications of all bacterial species tested, including A . actinomycetemcomitans, yielded 16S rDNA-specific DNA fragments . Furthermore, each bacterial species tested, with the exception of A . actinomycetemcomitans, failed to amplify lktA sequences . The LKT and RRN primers were used in further PCR experiments to detect A . actinomycetemcomitans directly from gingival fluid samples . The results clearly demonstrate the simplicity, rapidity, specificity and accuracy of the LKT primers in the identification of A . actinomycetemcomitans.

Mol Microbiol, 1993 Apr, 8(2), 243 - 52
Complex transcription of an operon encoding the SalI restriction-modification system of Streptomyces albus G; Alvarez MA et al.; High-resolution S1 nuclease mapping of mRNA synthesised in vivo, in vitro run-off transcription with RNA polymerase from Streptomyces lividans and gene fusions were used to analyse the transcriptional organization of the SalI restriction-modification system of Streptomyces albus G . The salIR and salIM genes that encode the restriction endonuclease and its cognate methyltransferase constitute an operon which is mainly transcribed from sal-pR1, a promoter located immediately upstream of salIR, with two possible minor promoters further upstream . Another promoter, sal-pM, is within the 3' end of the salIR coding region, and allows expression of the modification gene in the absence of sal-pR1 . The sal-pM promoter might be involved in the establishment of modification prior to restriction endonuclease activity . Sequences upstream of the apparent transcriptional start sites for sal-pR1 and sal-pM show similarity with the -10 region of typical vegetatively expressed eubacterial promoters, but appropriately centered -35 regions are absent.

Infect Agents Dis, 1993 Apr, 2(2), 87 - 99
Why is Chlamydia sensitive to penicillin in the absence of peptidoglycan?
Moulder JW.
Most eubacteria are sensitive to penicillin because the antibiotic inhibits synthesis of peptidoglycan, an essential constituent of their cell walls . A few eubacteria have no measurable peptidoglycan, and, with one exception, they are not susceptible to penicillin . The exception is the genus Chlamydia whose members are just as sensitive to penicillin as peptidoglycan-containing bacteria . A numbers of ways to resolve this anomaly, penicillin sensitivity without peptidoglycan, are proposed . It is concluded that there are serious objections to each one and that the chlamydial anomaly remains unresolved . However, examination of the relation between penicillin and chlamydiae is useful because it reveals how little is known of the evolutionary history of penicillin, penicillin-binding proteins, and peptidoglycan.

Curr Microbiol, 1993 Apr, 26(4), 233 - 7
Aspects of energy-yielding metabolism in the aphid, Schizaphis graminum, and its endosymbiont: detection of gene fragments potentially coding for the ATP synthase beta-subunit and glyceraldehyde-3-phosphate dehydrogenase; Clark MA et al.; Specialized cells within the aphid, Schizaphis graminum, contain intracellular, vesicle-enclosed eubacterial endosymbionts (Buchnera aphidicola) . Using oligonucleotide probes derived from conserved sequences of the ATP synthase beta-subunit and glyceraldehyde-3-phosphate dehydrogenase, and the polymerase chain reaction (PCR), we have amplified, cloned, and sequenced three DNA fragments . Amino acid sequence similarity indicated that two of these fragments corresponded to endosymbiont and host genes potentially coding for the beta-subunit of ATP synthase . The host gene fragment contained two putative introns . The third DNA fragment corresponded to a portion of a gene coding for a glyceraldehyde-3-phosphate dehydrogenase that was highly related to one of the enzymes from Escherichia coli (GapA) . These results indicate that B . aphidicola may have an ATP synthase and consequently could synthesize ATP from a proton motive force generated within the intracellular vesicles of host cells containing the endosymbionts . The detection of a gene fragment coding for a protein similar to glyceraldehyde-3-phosphate dehydrogenase suggests the presence of this glycolytic enzyme in the endosymbiont and its involvement in energy-yielding metabolism.

Science, 1993 Mar 26, 259(5103), 1892 - 6
Structural relationship of bacterial RecA proteins to recombination proteins from bacteriophage T4 and yeast; Story RM et al.; RecA protein is essential in eubacteria for homologous recombination and promotes the homologous pairing and strand exchange of DNA molecules in vitro . Recombination proteins with weak sequence similarity to bacterial RecA proteins have been identified in bacteriophage T4, yeast, and other higher organisms . Analysis of the primary sequence relationships of DMC1 from Saccharomyces cerevisiae and UvsX of T4 relative to the three-dimensional structure of RecA from Escherichia coli suggests that both proteins are structural homologs of bacterial RecA proteins . This analysis argues that proteins in this group are members of a single family that diverged from a common ancestor that existed prior to the divergence of prokaryotes and eukaryotes.

Nucleic Acids Res, 1993 Mar 25, 21(6), 1449 - 55
The sequence of 28S ribosomal RNA varies within and between human cell lines; Leffers H et al.; The primary structure of 28S ribosomal RNA constitutes a conserved core which is similar among most 23S-like rRNAs and expansion segments which occur at specific positions in the sequence . The expansion segments account for most of the size difference between prokaryotic (archaeal and eubacterial) and eukaryotic rRNAs and they exhibit a sequence variation which is unique among rRNAs . We have investigated the sequence variation of one of the expansion segments, V8, by sequencing a total of 111 V8 segments from 9 different human cell lines and tissues and have found 35 different variants . The variation occur mainly at two 'hot spots' which are separated by 170 nucleotides in the primary sequence but are neighbours in the secondary structure . The sequence of V8 segments varies both within and between human cell lines and tissues . The implications for the evolution of the eukaryotic 28S rRNA are discussed together with possible functions of the expansion segments . We also present a secondary structure model for the V8 segment based on comparative sequence analysis and chemical and enzymatic foot printing.

Nucleic Acids Res, 1993 Mar 25, 21(6), 1335 - 8
Lack of peptide-release activity responding to codon UGA in Mycoplasma capricolum; Inagaki Y et al.; In Mycoplasma capricolum, a relative of Gram-positive eubacteria with a high genomic AT-content (75%), codon UGA is assigned to tryptophan instead of termination signal . Thus, in this bacterium the release factor 2 (RF-2), that recognizes UAA and UGA termination codons in eubacteria such as Escherichia coli and Bacillus subtilis, would be either specific to UAA or deleted . To test this, we have constructed a cell-free translation system using synthetic mRNA including codon UAA {mRNA(UAA)}, UAG {mRNA(UAG)} and UGA {mRNA(UGA)} in-frame . In the absence of tryptophan, the translation of mRNA(UGA) ceased at UGA sites without appreciable release of the synthesized peptides from the ribosomes, whereas with mRNA(UAA) or mRNA(UAG) the bulk of the peptides was released . Upon addition of the E.coli S-100 fraction or B.subtilis S-100 fraction to the translation system, the synthesized peptides with mRNA(UGA) were almost completely released from the ribosomes, presumably because of the presence of RF-2 active to UGA in the added S-100 fraction . These data suggest that RF-2 is deleted or its activity to UGA is strongly weakened in M.capricolum.

J Biol Chem, 1993 Mar 25, 268(9), 6097 - 106
Old Yellow Enzyme . The discovery of multiple isozymes and a family of related proteins; Stott K et al.; Using fast protein liquid chromatography, we have separated native Old Yellow Enzyme from Brewer's Bottom Yeast into three distinct fractions . Two of these fractions are homodimeric forms of the enzyme while the third is the corresponding heterodimeric form . One of these homodimeric fractions is identical in every respect to OYE1, originally cloned from Brewer's Bottom Yeast (Saito, K., Thiele, D . J., Davio, M., Lockridge, O., and Massey, V . (1991) J . Biol . Chem . 266, 20720-20724) . We have cloned, sequenced, and expressed a second Old Yellow Enzyme gene from Saccharomyces cerevisiae, showing close similarity, but not identity, with OYE1 . Native Old Yellow Enzyme samples were also affinity-purified from a strain of S . cerevisiae and an OYE deletion mutant constructed from it . A total of at least seven isozymes of Old Yellow Enzyme have been discovered, each having slightly different characteristics ranging from surface charge to NADPH dehydrogenase activities with different electron acceptors, as well as N-terminal amino acid sequence . In addition, both recombinant enzymes showed considerable similarity to two proteins in the GenBank/EMBL data bank, a 60,000-dalton bile acid-inducible polypeptide in Eubacterium sp . (Mallonee, D . H., White, W . B., and Hylemon, P . B . (1990) J . Lipid Res . 172, 7011-7019) and a 72,000-dalton NADH oxidase in Thermoanaerobium brockii.

Proc Natl Acad Sci U S A, 1993 Mar 15, 90(6), 2305 - 9
From elongator tRNA to initiator tRNA; Varshney U et al.; We show that the two most important properties needed for a tRNA to function in initiation in Escherichia coli are its ability to be formylated and its ability to bind to the ribosomal P site . This conclusion is based on conversion of two different elongator tRNAs to ones that can act as initiators in E . coli . We transplanted the features unique to E . coli and eubacterial initiator tRNAs to E . coli elongator methionine tRNA (tRNA(Met)) along with an anticodon sequence change and analyzed their activities in initiation in E . coli . Introduction of a C1.A72 mismatch at the end of the acceptor stem of tRNA(Met), which generates the minimal features necessary for formylation, produces a tRNA with very low activity in initiation . Subsequent introduction of three consecutive G.C base pairs at the bottom of the anticodon stem, which is necessary for ribosomal P site binding, produces a tRNA with significant activity in initiation . Furthermore, introduction of the features necessary for formylation and for ribosomal P site binding into E . coli elongator glutamine tRNA produces a tRNA that initiates protein synthesis in E . coli.

J Mol Biol, 1993 Mar 5, 230(1), 57 - 76
Gene organization, primary structure and RNA processing analysis of a ribosomal RNA operon in Lactococcus lactis; Chiaruttini C et al.; Southern blot analysis of genomic DNA of the mesophilic lactic bacterium Lactococcus lactis subsp . lactis strain IL1403, illuminated six rRNA gene clusters . Each cluster contains one copy each of three rRNA genes, displaying the typical eubacterial organization of physically linked 16 S, 23 S and 5 S rRNA genes . Five of the six rRNA clusters were cloned into plasmid pBR322 . One recombinant plasmid, pSLCM6, containing a 6500 base-pair genomic DNA fragment, was characterized by physical mapping and the sequences encoding rRNAs and tRNAs were localized by Southern hybridization . This fragment contains a single operon composed of one promoter, a leader sequence, a 16 S rRNA gene, a tRNA(Ala) gene, a 23 S rRNA gene, a 5 S rRNA gene and a tRNA(Asn) gene . S1 nuclease mapping and primer extension analysis of in vivo transcripts localized one transcriptional initiation site 150 base-pairs upstream from the start of the 16 S rRNA gene . These procedures also suggest that this transcript is processed by an RNAse III-like activity similar to Bacillus subtilis; i.e . the L . lactis nuclease might be sequence-specific . The chronology of specific cleavages occurring during the maturation process of the precursor transcript is described . One interesting observation is that the regions flanking the 16 S and 23 S rRNAs containing the primary processing sites are identical and contain sequences that could be involved in transcriptional antitermination . S1 mapping of the 3' ends of in vivo transcripts indicate that a terminator-like sequence a few base-pairs downstream from the distal tRNA(Asn) gene is inefficient in arresting transcription.

J Mol Biol, 1993 Mar 5, 230(1), 51 - 6
Unassigned or nonsense codons in Micrococcus luteus; Kano A et al.; We previously reported that in Micrococcus luteus, a Gram-positive eubacterium with high genomic G + C content, certain codons ending with A did not appear in coding frames, including termination sites, and tRNAs that translate these codons were not detected . These facts suggest that at least some of them are unassigned (nonsense) codons, i.e . not assigned to any amino acid or to any stop signal . We have investigated whether AGA and AUA, universal Arg and Ile codons, respectively, are really unassigned codons by using a cell-free extract prepared from M . luteus and synthetic messenger RNAs . Translation of synthetic mRNA containing in-frame AGA codons does not result in "read-through" to codons beyond the AGA codons, i.e . translation ceases at codon AGA . Essentially the same result was obtained with mRNA containing AUA in-frame . A sucrose-gradient centrifugation profile of the reaction mixture has shown that practically all of the peptides that have been synthesized are attached to 70 S ribosomes . When in-frame AGA or AUA codons are replaced by UGA codons in mRNA, no read-through occurs beyond UGA, just as in the case of AGA or AUA . However, the synthesized peptide is released from the 70 S ribosomes . These data suggest that AGA and AUA are unassigned codons and differ from UGA in that they are not used for termination.

J Biol Chem, 1993 Mar 5, 268(7), 4643 - 50
Cross-linked amino acids in the protein pairs L3-L19 and L23-L29 of Bacillus stearothermophilus ribosomes after treatment with diepoxybutane; Herwig S et al.; Treatment of native 50 S ribosomal subunits of Bacillus stearothermophilus with the homobifunctional cross-linking reagent diepoxybutane generated two cross-linked protein pairs, L3-L19 and L23-L29, which were isolated and identified . The analysis of the cross-linking sites at the amino acid level in both protein pairs is presented . Using a combination of sequence analysis and mass spectrometry it could be demonstrated that His-28 in protein L3 and the N-terminal amino acids Met-1, His-2, and His-3 in protein L19 are involved in forming the cross-link L3-L19 . Within the pair L23-L29 Met-1 in protein L23 and Lys-4 in protein L29 were identified as cross-linking sites employing a similar approach . Comparison of our data with results derived from other cross-linking experiments showed that in general the structural organization of the ribosomes in eubacteria (the Gram-positive B . stearothermophilus and the Gram-negative Escherichia coli) has been conserved to quite an extent during evolution but that the fine structures differ slightly . By mass spectrometry the specificity of diepoxybutane and its cleaving mechanism using sodium periodate could be examined . In addition the complete amino acid sequence of protein L19 of B . stearothermophilus has been determined and revealed 58% identical amino acid residues to the homologous E . coli protein L19.

FEMS Immunol Med Microbiol, 1993 Mar, 6(2-3), 213 - 7
Black-pigmented gram-negative anaerobes in endodontic infections; Haapasalo M; Necrotic dental root canal infections are polymicrobial infections dominated by anaerobic bacteria . The number of different species in one canal is usually low, approx . 4-7 species . The species isolated most frequently belong to the genera Prevotella, Porphyromonas, Fusobacterium, Peptostreptococcus, Eubacterium and Streptococcus . The frequency of isolation of black-pigmented Gram-negative anaerobes in endodontic infections varies from 25% to > 50% . Pr . intermedia is the most commonly found pigmented species, followed by Pr . denticola and two Porphyromonas species, P . gingivalis and P . endodontalis . Several studies have shown that P . gingivalis and P . endodontalis are closely related to the presence of acute symptoms in endodontic infections, whereas other black-pigmented Gram-negative anaerobes are not . However, several other species may also be involved in acute infections . Moreover, Porphyromonas species have occasionally been isolated from cases with no symptoms . Although Porphyromonas spp . are clearly related to symptoms at the beginning of therapy, they are not important for the prognosis of the treatment.

Plant Mol Biol, 1993 Mar, 21(5), 779 - 87
Secondary structure and phylogeny of the chloroplast 23S rRNA gene from the brown alga Pylaiella littoralis; Somerville CC et al.; The entire nucleotide sequence of a 23S rRNA gene from the brown alga Pylaiella littoralis (L.) Kjellm has been determined . The predicted length of the 23S rRNA is 2948 nucleotides, including the 4.5S rRNA-like region at the 3' end of the molecule . The putative transcript has been folded into a secondary structure by comparison to existing structure models, and the predicted helical regions were inspected by identifying compensatory downstream base changes . The 23S rRNA secondary structure presented here has features that are unique to P . littoralis (no other chromophyte or red algal 23S rRNA sequences are yet available), but has none of the features specific to the chloroplast rRNAs of green plants and green algae . The Pylaiella sequence was aligned with analogous plastidial and eubacterial gene sequences, and the alignment was used to construct a phylogenetic tree . The plastidial sequences formed a coherent cluster closely associated with the 23S rRNA of the cyanobacterium Anacystis nidulans . Within the plastid group, the P . littoralis sequence was most closely related to that of Euglena gracilis confirming earlier analyses based upon 16S rRNA sequences.

J Clin Microbiol, 1993 Mar, 31(3), 646 - 52
Elimination of contaminating DNA within polymerase chain reaction reagents: implications for a general approach to detection of uncultured pathogens; Meier A et al.; Analysis based on comparisons of 16S rRNA sequences provides a rapid and reliable approach to identifying human pathogens . By directing oligonucleotide primers at sequences conserved throughout the eubacterial kingdom, bacterial 16S ribosomal DNA sequences of virtually any member of the eubacterial kingdom can be amplified by polymerase chain reaction and subsequently analyzed by sequence determination . Indeed, automated systems for broad-range amplification, sequencing, and data analysis are now feasible and may form the basis of the next generation of automated microbial identification systems . However, identification of pathogens by this strategy is hampered by the frequent contamination of reagents used for the amplification reaction, in particular Taq polymerase, with exogenous bacterial DNA . Here, we describe detailed investigations on the use of 8-methoxypsoralen and long-wave UV light to eliminate contaminating DNA in polymerase chain reaction reagents . The clinical utility of the developed procedure was demonstrated in a case of paucibacillary osteomyelitis, for which no specific bacterial agent had been cultured.

J Bacteriol, 1993 Mar, 175(5), 1514 - 23
Cloning, characterization, and functional expression in Escherichia coli of chaperonin (groESL) genes from the phototrophic sulfur bacterium Chromatium vinosum; Ferreyra RG et al.; A recombinant lambda phage which was able to propagate in groE mutants of Escherichia coli was isolated from a Chromatium vinosum genomic DNA library . A 4-kbp SalI DNA fragment, isolated from this phage and subcloned in plasmid vectors, carried the C . vinosum genes that allowed lambda growth in these mutants . Sequencing of this fragment indicated the presence of two open reading frames encoding polypeptides of 97 and 544 amino acids, respectively, which showed high similarity to the molecular chaperones GroES and GroEL, respectively, from several eubacteria and eukaryotic organelles . Expression of the cloned C . vinosum groESL genes in E . coli was greatly enhanced when the cells were transferred to growth temperatures that induce the heat shock response in this host . Coexpression in E . coli of C . vinosum groESL genes and the cloned ribulose bisphosphate carboxylase/oxygenase genes from different phototrophic bacteria resulted in an enhanced assembly of the latter enzymes . These results indicate that the cloned DNA fragment encodes C . vinosum chaperonins, which serve in the assembly process of oligomeric proteins . Phylogenic analysis indicates a close relationship between C . vinosum chaperonins and their homologs present in pathogenic species of the gamma subdivision of the eubacterial division Proteobacteria.

Res Microbiol, 1993 Mar-Apr, 144(3), 187 - 200
The histidine operon of Azospirillum brasilense: organization, nucleotide sequence and functional analysis; Fani R et al.; A 3457-base pair fragment of Azospirillum brasilense DNA which complemented mutations in the hisA and hisF genes of Escherichia coli was sequenced . The sequence analysis revealed the presence of six major contiguous open reading frames (ORF) . The comparison of the predicted amino acid sequence of these ORF with those encoded by the eubacterial, archaebacterial and eukaryotic his genes sequenced thus far revealed that four of them have a significant degree of homology with the E . coli hisH, hisA, hisF and the C-terminal domain of the hisI gene products . S1 mapping experiments indicated that the putative transcription start site coincided with the AUG translational initiation codon of the hisBd gene, the first gene of the A . brasilense his operon . Downstream from the last ORF, a sequence was identified which functions as a Rho-independent transcription terminator . Comparison of amino acid sequences, gene order and organization and evolutionary aspects of the A . brasilense his cluster are discussed.

Curr Microbiol, 1993 Mar, 26(3), 161 - 6
The genome of the non-cultured, bacterial-like organism associated with citrus greening disease contains the nusG-rplKAJL-rpoBC gene cluster and the gene for a bacteriophage type DNA polymerase; Villechanoux S et al.; We have recently cloned three DNA fragments (In-2.6, In-1.0, and In-0.6) of the non-cultured, bacterial-like organism (BLO) associated with citrus greening disease . Nucleotide sequence determination has shown that fragment In-2.6 is part of the rplKAJL-rpoBC gene cluster, a well-known operon in eubacteria . The DNA fragment upstream of and partially overlapping with In-2.6 could be isolated and was shown to be the nusG gene . In Escherichia coli, nusG is also immediately upstream of rplKAJL-rpoBC . Fragment In-1.0 carries the gene for a bacteriophage type DNA polymerase . Fragment In-0.6 could not be identified . When In-2.6 was used, at high stringency, as a probe to detect greening BLO strains in infected plants, hybridization was obtained with all Asian strains tested, but not with the African strain examined . At lower stringencies, In-2.6 was able to detect also the African strain . The implications of these results in the taxonomical position of the greening BLO are discussed.

Curr Microbiol, 1993 Mar, 26(3), 129 - 32
Congruent evolution between whiteflies (Homoptera: Aleyrodidae) and their bacterial endosymbionts based on respective 18S and 16S rDNAs; Campbell BC; Whiteflies (family Aleyrodidae) possess heritable eubacterial endosymbionts sustained in specialized organ-like structure called mycetomes . Comparisons of distances between the ash whitefly, Siphoninus phillyreae, and two biotypes ("A" and "B") of the sweetpotato whitefly, Bemisia tabaci, based on sequence analysis of genes for 18S rRNAs (rDNAs), were equivalent to the distances represented by the 16S rDNAs of their respective endosymbionts . This finding indicates that evolutionary divergence in whitefly hosts and their endosymbionts is congruent . The nucleotide sequences of the 18S rDNAs and endosymbiont 16S rDNAs indicate the two biotypes of B . tabaci are the same species.

Appl Environ Microbiol, 1993 Mar, 59(3), 687 - 94
Polymerase chain reaction amplification of naphthalene-catabolic and 16S rRNA gene sequences from indigenous sediment bacteria; Herrick JB et al.; We report the amplification of bacterial genes from uninoculated surface and subsurface sediments by the polymerase chain reaction (PCR) . PCR amplification of indigenous bacterial 16S ribosomal DNA genes was unsuccessful when subsurface sediment containing approximately 10(7) cells.g-1 was added directly to a PCR mixture . However, when 10 mg of sediment was inoculated with approximately 10(5) cells of Pseudomonas putida G7, the nahAc naphthalene dioxygenase gene characteristic of the P . putida G7 NAH7 plasmid was detected by PCR amplification . Southern blotting of the PCR amplification product improved sensitivity to 10(3) to 10(4) cells from samples inoculated with P . putida G7, but controls with no sediment added showed that the PCR was partially inhibited by the sediments . Lysozyme-sodium dodecyl sulfate-freeze-thaw DNA extraction was combined with gel electrophoretic partial purification in the presence of polyvinylpyrrolidone to render DNA from indigenous bacteria in surface or subsurface sediment samples amplifiable by PCR using eubacterial 16S ribosomal DNA primers . The nahAc gene could also be amplified from indigenous bacteria by using nahAc-specific primers when PCR conditions were modified by increasing Taq and primer concentrations . Restriction digests of the nahAc amplification products from surface and subsurface sediments revealed polymorphism relative to P . putida G7 . The procedures for DNA extraction, purification, and PCR amplification described here demonstrate that the PCR is a potentially useful tool in studies of function- and taxon-specific DNA from indigenous microbial communities in sediment and groundwater environments.

FEBS Lett, 1993 Feb 22, 318(1), 61 - 4
Evidence for a Rieske-type FeS center in the thermoacidophilic archaebacterium Sulfolobus acidocaldarius; Anemuller S et al.; A high-potential iron-sulfur cluster with characteristics similar to a Rieske-type center was detected in the plasma membrane of Sulfolobus acidocaldarius by EPR spectroscopy . In the reduced form this center has g-values of gz = 2.031, gy = 1.890 and gx = 1.725 (gav = 1.88, rhombicity = 0.37) and its reduction potential at pH 7.4 was determined to be +325 +/- 10 mV . The The archaebacterial cluster exhibits some unique properties, in comparison to eubacterial and eukaryotic Rieske-type centers . First, the reduction potential is pH-dependent in the range from pH 6.7 to 8.2 . Second, the typical inhibitor of Rieske FeS centers, DBMIB, had no effect on the g-values of this cluster . The center is reducible by both NADH and succinate in the presence of cyanide, an inhibitor of the terminal oxidases . The possible role of a Rieske-type center in an organism lacking any c-type cytochromes is discussed.

Eur J Biochem, 1993 Feb 15, 212(1), 41 - 9
Glucose-6-phosphate dehydrogenase . Structure-function relationships and the Pichia jadinii enzyme structure; Jeffery J et al.; The primary structure of glucose-6-phosphate dehydrogenase from the yeast Pichia jadinii (formerly Candida utilis) has been determined . It consists of a 495-residue, N-terminally acetylated protein chain . The structure shows extensive differences from those of the corresponding mammalian, fruit fly, and bacterial enzymes (52-68% residue non-identities), but also from that of another yeast, Saccharomyces cerevisiae (38%) . A eubacterial type and a yeast type of glucose-6-phosphate dehydrogenase are discerned, in addition to the known mammalian type . They are distinguished from each other, from the mammalian type, and the insect enzyme, on the basis of both specific residues and pattern differences . The distribution of residues conserved in all forms locates short segments in which identities are closely grouped . Approximately 50% of these segments correspond to predicted turns and appear to mark the principal folds characteristic of the enzyme's tertiary structure . A region in the N-terminal part of the protein chain has characteristics suggestive of a coenzyme-binding site, while, in the middle third, another functionally important segment may be related to glucose-6-phosphate binding and catalysis.

Gene, 1993 Feb 14, 124(1), 67 - 74
Domain V of Giardia lamblia large-subunit rRNA: structure of the peptidyl transferase loop from an early-branching eukaryote and correlation with antibiotic sensitivity; Edlind TD et al.; Large subunit rRNA (LSR) sequences that have been implicated in peptide bond formation form a specific secondary structure called the peptidyl transferase loop (PTL) . Although well conserved, the PTLs of eubacteria, archaebacteria, and eukaryotes have several distinct differences . These differences correlate with different sensitivities to peptidyl transferase and translocase inhibitors . To shed light on the basis for these kingdom-specific differences in PTL structure and function, we have analyzed the sequence and secondary structure of LSR domain V, which contains the PTL, from Giardia lamblia . This parasitic protozoan derives from a very early branch in eukaryotic evolution, and its rRNA was previously shown to have bacteria-like features . In vitro and cell-free systems were also used to test the sensitivity of G . lamblia protein synthesis to specific PTL-targeted inhibitors . Our results indicate that the PTL structure and inhibitor sensitivity typical of higher eukaryotes is conserved in G . lamblia . However, several adjacent domain V sequences more closely resemble archaebacterial rRNA, confirming the 'primitive' nature of G . lamblia rRNA . Thus, the eukaryotic PTL has been conserved over a vast evolutionary period . We speculate that the eukaryotic PTL is primordial and employs specific RNA-RNA interactions to catalyze protein synthesis . Three potential interactions were identified.

Nucleic Acids Res, 1993 Feb 11, 21(3), 649 - 55
Zinc finger-like motifs in rat ribosomal proteins S27 and S29; Chan YL et al.; The primary structures of the rat 40S ribosomal subunit proteins S27 and S29 were deduced from the sequences of nucleotides in recombinant cDNAs and confirmed by determination of amino acid sequences in the proteins . Ribosomal protein S27 has 83 amino acids and the molecular weight is 9,339 . Hybridization of cDNA to digests of nuclear DNA suggests that there are 4-6 copies of the S27 gene; the mRNA for the protein is about 620 nucleotides in length . Ribosomal protein S29 has 55 amino acids and the molecular weight is 6,541 . There are 14-17 copies of the S29 gene and its mRNA is about 500 nucleotides in length . Rat ribosomal protein S29 is related to several members of the archaebacterial and eubacterial S14 family of ribosomal proteins . S27 and S29 have zinc finger-like motifs as do other proteins from eukaryotic, archaebacterial, eubacterial, and mitochondrial ribosomes . Moreover, ribosomes and ribosomal subunits appear to contain zinc and iron as well.

J Biol Chem, 1993 Feb 5, 268(4), 2828 - 35
Sarcina ventriculi synthesizes very long chain dicarboxylic acids in response to different forms of environmental stress; Jung S et al.; Changes in the composition of membrane lipids in a strictly anaerobic, facultative acidophilic eubacterium, Sarcina ventriculi, were studied in response to various forms of environmental stress . Changes in lipid composition and structure occurred in response to changes in environmental pH . At neutral pH, the predominant membrane fatty acids ranged in chain length from C14 to C18 . However, when cells were grown at pH 3.0, a family of unique very long chain fatty acids containing 32-36 carbon atoms was synthesized and accounted for 50% of the total membrane fatty acids . These acids were identified as very long chain alpha,omega-dicarboxylic acids ranging in length from 28 to 36 carbons by electron impact mass spectrometry of methyl and (perdeuterio) methyl ester derivatives . These methyl esters all bore a vicinal dimethyl group toward the center of the chain . The assignment of the structures was confirmed by isolating one of the very long chain unusual fatty acids as the ester form after methanolysis and performing further analyses including 1H and 13C NMR spectroscopy and Fourier transform infrared spectroscopy . Coupling this information with the data from gas chromatography/mass spectrometry analysis, the exact structure was confirmed as alpha,omega-15,16-dimethyltricotanedioate dimethyl ester . Addition of alcohols, either metabolic (0.25 M ethanol) or nonmetabolic (0.05 M butanol) to cells grown at pH 7.0, or thermal stress (growth temperature at pH 7.0 was raised from 37 to 45 or 55 degrees C) also resulted in the synthesis of these very long chain fatty acids . Synthesis of these very long chain alpha,omega-dicarboxylic acids was reversed by reducing the temperature back to 37 degrees C . S . ventriculi is also unusual in that the membrane components are not the usual phospholipid components but appear to be predominantly glycolipids.

J Biol Chem, 1993 Feb 5, 268(4), 2755 - 61
The primary structure of rat ribosomal protein L23a . The application of homology search to the identification of genes for mammalian and yeast ribosomal proteins and a correlation of rat and yeast ribosomal proteins; Suzuki K et al.; The amino acid sequence of the rat 60 S ribosomal subunit protein L23a was deduced from the sequence of nucleotides in a recombinant cDNA . Ribosomal protein L23a has 156 amino acids and a molecular weight of 17,684 . Hybridization of the L23a cDNA to digests of nuclear DNA suggests that there are 18-20 copies of the L23a gene . The mRNA for the protein is about 600 nucleotides in length . Rat L23a is related to the yeast Saccharomyces cerevisiae L25, to the archaebacterial Methanococcus vannielii L23, to eubacterial Escherichia coli L23, and to other members of the L23 family of ribosomal proteins . A novel application of a routine homology search procedure was employed to identify a nucleotide sequence that could be used to design an oligodeoxynucleotide probe to screen a library for a cDNA that encodes rat L23a; this same procedure uncovered a number of previously unidentified genes for yeast ribosomal proteins in the GenBank DNA data base . In a correlation of rat and yeast ribosomal proteins 48 pairs are shown to be related.

Mol Cell Probes, 1993 Feb, 7(1), 7 - 17
Amplification of rDNA loci to detect and type Neisseria meningitidis and other eubacteria; McLaughlin GL et al.; In 1991-92, Neisseria meningitidis group C was isolated from the blood of eight students in Urbana, Illinois, USA, and from the cerebrospinal fluid of one student from a nearby community, Decatur, Illinois . These and other bacterial species were analysed by PCR fingerprinting using primers selected from the ribosomal (r)DNA loci . A rDNA primer pair spanning a region within the 16S rDNA amplified a predicted 280 base pair (bp) DNA fragment from Neisseria spp . and fragments of different sizes for other genera . This primer pair specifically detected a carrier of N . meningitidis in a small clinical battery . Identity of the fragment was confirmed by restriction endonuclease analysis . A 600 bp fragment was also amplified from the 16S-23S internal transcribed spacer (ITS) of N . meningitidis; amplification from six other genera yielded different-sized fragments . Digestion of the ITS fragment from N . meningitidis with Alu I revealed three patterns; pattern I was found only for serogroup C isolates, and it was the dominant pattern among recent isolates with the exception of the one from Decatur . The isolate from Decatur yielded pattern III which suggested a non-clonal relationship to the seven isolates from Urbana . Patterns II and III were more prevalent in isolates from the 1960's and 1980's . PCR-based analysis of these loci can complement the techniques which are currently used for the detection and typing of these and other eubacteria.

J Bacteriol, 1993 Feb, 175(3), 669 - 73
Initial steps in the anaerobic degradation of 3,4,5-trihydroxybenzoate by Eubacterium oxidoreducens: characterization of mutants and role of 1,2,3,5-tetrahydroxybenzene; Haddock JD et al.; Chemical mutagenesis and antibiotic enrichment techniques were used to isolate five mutant strains of the obligate anaerobe Eubacterium oxidoreducens that were unable to grow on 3,4,5-trihydroxybenzoate (gallate) . Two strains could not transform gallate and showed no detectable gallate decarboxylase activity . Two other strains transformed gallate to pyrogallol and dihydrophloroglucinol but lacked the hydrolase activity responsible for ring cleavage . A fifth strain accumulated pyrogallol, although it contained adequate levels of the enzymes proposed for the complete transformation of gallate to the ring cleavage product . The conversion of pyrogallol to phloroglucinol by cell extract of the wild-type strain was dependent on the addition of 1,2,3,5-tetrahydroxybenzene or dimethyl sulfoxide . This activity was induced by growth on gallate, while the other enzymes involved in the initial reactions of gallate catabolism were constitutively expressed during growth on crotonate . The results confirm the initial steps in the pathway previously proposed for the metabolism of gallate by E . oxidoreducens, except for the conversion of pyrogallol to phloroglucinol.

J Bacteriol, 1993 Feb, 175(3), 583 - 8
Retrotransfer in Escherichia coli conjugation: bidirectional exchange or de novo mating?
Heinemann JA, Ankenbauer RG.
DNA can be transferred among eubacteria and to plants and fungi by related, plasmid-mediated processes collectively referred to as bacterial conjugation . Conjugation occurs between cells in contact with one another and results in the unidirectional delivery of DNA from a bacterial donor to a recipient . Recent experiments that have reexamined the directionality of DNA flow during conjugation have come to different conclusions, some suggesting that genetic material also flows from recipient cells into the donor and that this process, termed retrotransfer, is likewise directed by donor-encoded functions . Given that bacteria are perhaps united with all living creatures by conjugation, the possibility of gene flow into donor bacteria during conjugation raises interesting evolutionary and biocontainment issues . Here we report that plasmid transmission from bacterial recipients to donors is not a donor-mediated event . Movement of genetic material from recipients to donors was inhibited by streptomycin, which does not inhibit the conjugative donor, indicating that retrotransfer requires gene expression in recipients . Furthermore, retrotransfer was reduced in matings mediated by plasmids that encode strong entry exclusion, to a similar degree as matings between two donors . Therefore we suggest that retrotransfer is in fact newly initiated conjugation between transconjugants and donors.

Appl Environ Microbiol, 1993 Feb, 59(2), 417 - 21
Detection of Tn5-like sequences in kanamycin-resistant stream bacteria and environmental DNA; Leff LG et al.; Resistance to kanamycin and neomycin in the bacterial assemblage of a coastal plain stream was detected by growth of colonies on media containing antibiotics . Three of 184 kanamycin-resistant colonies hybridized with a probe containing the nptII gene from transposon Tn5; the nptII gene encodes the enzyme neomycin phosphotransferase and conveys resistance to kanamycin and neomycin . In one of these isolates, the homologous gene was cloned and shown to confer resistance to a kanamycin-sensitive Escherichia coli strain . Since enumeration of bacteria by acridine orange direct counts revealed that less than 0.2% of the bacteria present were cultivated, direct examination of environmental DNA was used to assess abundance of sequences that hybridize to the nptII gene . To examine the resistance potential of bacteria that were not cultured, total DNA was extracted from environmental samples and hybridized with specific probes . The relative amount of eubacterial DNA in each sample was determined by using a eubacterial specific rDNA probe . Then, the abundance of sequences that hybridize to the eubacterial neomycin phosphotransferase gene was determined by hybridization and expressed relative to the total eubacterial DNA in the assemblage . Relative gene abundance was significantly different among assemblages from different habitats (leaves, midchannel sediments, and bank sediments) but did not differ among stream sites.

Mol Microbiol, 1993 Feb, 7(4), 577 - 84
Identification and characterization of IS1138, a transposable element from Mycoplasma pulmonis that belongs to the IS3 family; Bhugra B et al.; Insertion sequence (IS) elements are mobile genetic elements found in prokaryotes . We have identified a repetitive element from Mycoplasma pulmonis, a murine pathogen, that is similar to eubacterial IS elements . By subcloning a single strain of M . pulmonis, we isolated a variant clone in which the IS element had undergone an apparent transposition event . The nucleotide sequences of the element, designated IS1138, and the target site into which it inserted were determined . IS1138 consists of 1288 bp with 18 bp perfect terminal inverted repeats . Sequence analysis of the target site before and after insertion of IS1138 identified a 3 bp duplication of target DNA flanking the element . The predicted amino acids encoded by the major open reading frame of IS1138 share significant similarity with the transposases of the IS3 family . Southern hybridization analysis indicates that repetitive sequences similar to IS1138 are present in most, if not all, strains of M . pulmonis, but IS1138-like sequences were not detected in other mycoplasmal species.

Gene, 1993 Jan 15, 123(1), 25 - 32
Nucleotide sequences of the trpI, trpB, and trpA genes of Pseudomonas syringae: positive control unique to fluorescent pseudomonads; Auerbach S et al.; A 904-bp probe from Pseudomonas aeruginosa was used to identify the trpB, trpA and trpI genes of Pseudomonas syringae . Transcription initiation at the P . syringae trpBA promoter in vitro was activated by the P . aeruginosa TrpI protein in the presence of indoleglycerol phosphate . Thus, trpB and trpA are regulated positively in three species of fluorescent pseudomonads, P . aeruginosa, P . putida, and P . syringae, but in no other eubacteria so far investigated {Crawford, Annu . Rev . Microbiol . 43 (1989) 567-600} . In addition to conservation of protein-coding sequences, there is a high degree of nucleotide sequence identity in the intergenic control region that includes the divergent trpI and trpBA promoters, especially in the binding sites for TrpI protein . Differences in patterns of codon usage distinguish the trpI genes of P . syringae and P . putida from P . aeruginosa trpI and from the trpB and trpA genes of all three species.

Gene, 1993 Jan 15, 123(1), 75 - 80
Allele-specific structure probing of plasmid-derived 16S ribosomal RNA from Escherichia coli; Powers T et al.; Biochemical analysis of Escherichia coli ribosomes containing mutant 16S or 23S (r)ribosomal RNAs, produced via cloned rDNA genes on multicopy plasmids, has been hindered by the background of wild-type (wt) ribosomes containing host-derived rRNA . Here, we describe a method for the construction of unique priming sites in 16S rRNA that allow allele-specific structure probing of ribosomes containing plasmid-encoded RNA . Phenotypically silent mutations, designed to mimic related eubacterial sequences, have been introduced into four phylogenetically variable regions in the 16S rDNA gene that allow inspection of several 16S rRNA functional sites . When oligodeoxyribonucleotides complementary to these altered sequences are used to prime cDNA synthesis in primer extension reactions using reverse transcriptase, only plasmid-derived 16S rRNA is used as a template, thus rendering the wt background invisible . Unexpectedly, we were unable to introduce silent mutations into one nonconserved helix in 16S rRNA, suggesting that constraints in addition to Watson-Crick pairing are important in this region.

Biochem Mol Biol Int, 1993 Jan, 29(1), 25 - 31
A small novel chloroplast ribosomal protein (S31) that has no apparent counterpart in the E . coli ribosome; Schmidt J et al.; Higher plant chloroplast ribosomes contain several novel protein components whose homologues are not present in the eubacterial E . coli ribosome, indicating a complex evolution of the chloroplast translational apparatus following the endosymbiotic event . Here we describe the isolation and characterization of a new small protein from spinach chloroplast ribosome which has, based on the amino acid sequence and immunological data, no counterpart in the E . coli ribosome . We suggest a nomenclature suitable for this protein and other novel proteins in the chloroplast ribosome.

Ital J Biochem, 1993 Jan-Feb, 42(1), 1 - 11
Properties of the purified elongation factor 2 in the thermoacidophilic archaebacterium Sulfolobus solfataricus; Raimo G et al.; The elongation factor 2 (aEF-2) has been purified to homogeneity from the extreme thermoacidophilic archaebacterium Sulfolobus solfataricus . It is the only target protein which is ADP-ribosylated by diphtheria toxin in presence of NAD and this modification abolishes its property to support poly(Phe) synthesis in vitro . The factor is constituted by a single polypeptide chain with a relative molecular mass of 78,000 and an isoelectric point of 5.9 . aEF-2 is resistant to heat denaturation as shown by the fact that its capability to be ADP-ribosylated was only 10% reduced after 4 h treatment at 80 degrees C . Its amino acid composition does not reveal significant differences with that of analogous factors in other sources; nevertheless, the deviation function indicates that aEF-2 is related to Sulfolobus acidocaldarius and eukaryotes EF-2 more than to eubacterial EF-G or other archaebacterial EF-2.

Vet Microbiol, 1993 Jan, 34(1), 89 - 95
Chromosomal DNA probes for the identification of Bacteroides tectum and Bacteroides fragilis from the oral cavity of cats; Love DN et al.; A dot-blot hybridisation assay using high molecular weight DNA as whole chromosomal probes was used to differentiate Bacteroides tectum from Bacteroides fragilis . 32P-labelled probes were compared with digoxigenin (DIG)-labelled probes . The whole chromosomal probes were specific--differentiating B . tectum from B . fragilis and both from a variety of other species (including other members of the genera Bacteroides, Fusobacterium, Eubacterium, and Prevotella) found in normal and abnormal mouths of cats and horses . However, even at very high stringencies, B . tectum homology groups I, II and III were not distinguishable from one another using either 32P-labelled or DIG-labelled probes . Thus, DIG-labelled whole chromosome probes directed against cellular DNA released directly onto nitrocellulose membranes is considered a useful method for diagnostic veterinary laboratories wishing to identify B . tectum and distinguish it from B . fragilis and other oral anaerobic flora of cats.

Mol Microbiol, 1993 Jan, 7(2), 159 - 65
SecA protein: autoregulated ATPase catalysing preprotein insertion and translocation across the Escherichia coli inner membrane; Oliver DB; Recent insight into the biochemical mechanisms of protein translocation in Escherichia coli indicates that SecA ATPase is required both for the initial binding of preproteins to the inner membrane as well as subsequent translocation across this structure . SecA appears to promote these events by direct recognition of the preprotein or preprotein-SecB complex, binding to inner-membrane anionic phospholipids, insertion into the membrane bilayer and association with the preprotein translocator, SecY/SecE . ATP binding appears to control the affinity of SecA for the various components of the system and ATP hydrolysis promotes cycling between its different biochemical states . As a component likely to catalyse a rate-determining step in protein secretion, SecA synthesis is co-ordinated with the activity of the protein export pathway . This form of negative regulation appears to rely on SecA protein binding to its mRNA and repressing translation if conditions of rapid protein secretion prevail within the cell . A precise biochemical scheme for SecA-dependent catalysis of protein export and the details of secA regulation appear to be close at hand . The evolutionary conservation of SecA protein among eubacteria as well as the general requirement for translocation ATPases in other protein secretion systems argues for a mechanistic commonality of all prokaryotic protein export pathways.

J Bacteriol, 1993 Jan, 175(1), 133 - 40
Proteins antigenically related to methyl-accepting chemotaxis proteins of Escherichia coli detected in a wide range of bacterial species; Morgan DG et al.; The four methyl-accepting chemotaxis proteins of Escherichia coli, often called transducers, are transmembrane receptor proteins that exhibit substantial identity among the sequences of their cytoplasmic domains . Thus, antiserum raised to one of these proteins recognizes the others and might be expected to recognize related proteins in other bacteria . We used antiserum raised to the transducer Trg in immunoblot experiments to probe a wide range of bacterial species for the presence of antigenically related proteins . Such proteins were detected in over 20 different species, representing 6 of the 11 eubacterial phyla defined by analysis of rRNA sequences as well as one archaebacterial group . Species containing proteins antigenically related to the transducers of E . coli included members of all four subdivisions of the phylum in which E . coli is placed, members of four of the six subdivisions of spirochetes, and two gliding bacteria . These observations provide substantial support for the notion that methyl-accepting taxis proteins are widely distributed among the diversity of bacterial species.

Protein Sci, 1993 Jan, 2(1), 20 - 30
Mammalian integral membrane receptors are homologous to facilitators and antiporters of yeast, fungi, and eubacteria; Reizer J et al.; We demonstrate that three integral membrane receptors of mammals--the ecotropic retroviral leukemia receptor (ERR), the human retroviral receptor (HRR), and the T-cell early activator (Tea)--are homologous to a family of transporters specific for amino acids, polyamines, and choline (APC), which catalyze solute uniport, solute:cation symport, or solute:solute antiport in yeast, fungi, and eubacteria . Interestingly, the ERR membrane protein was recently shown to function as a cation:amino acid cotransporter . A binary sequence similarity matrix and an evolutionary tree of the 14 members of this family, illustrating their sequence similarities and divergences, were constructed . Other proteins, including the developmentally controlled GerAII spore germination protein of Bacillus subtilis and the acetylcholine receptor of Drosophila melanogaster gave sequence comparison scores of a sufficiently large magnitude to suggest (but not to establish) a common evolutionary origin with members of the APC family . We report an extended and corrected Tea cDNA sequence and show that the mammalian Tea and ERR encoding genes are differentially expressed in tissues and cell lines . Furthermore, the two mammalian cDNA sequences hybridize with other vertebrate and yeast genomic DNAs under stringent conditions . These observations support the notion that cell surface receptor proteins in mammals are transport proteins that share a common origin with transport proteins of single-celled organisms . Thus, permeases of essential metabolites may function pathologically as viral receptors.

Antimicrob Agents Chemother, 1993 Jan, 37(1), 46 - 50
Coumermycin A1 inhibits growth and induces relaxation of supercoiled plasmids in Borrelia burgdorferi, the Lyme disease agent; Samuels DS et al.; Coumermycin A1 is an inhibitor of DNA gyrase, an enzyme that catalyzes supercoiling of DNA and is required for bacterial DNA replication . We have investigated the activity of this coumarin antibiotic on Borrelia burgdorferi, a spirochete and the causative agent of Lyme disease . B . burgdorferi was more susceptible than many other eubacteria to coumermycin as well as novobiocin, another coumarin antibiotic; this contrasted with its relative resistance to the DNA gyrase inhibitors nalidixic acid, oxolinic acid, and ciprofloxacin . Coumermycin at 0.2 micrograms/ml inhibited the growth of B . burgdorferi B31 in BSK II medium . A 100-fold-lower concentration induced the relaxation of two negatively supercoiled circular plasmids within 2 h . Plasmid supercoiling was restored within 2 h of removal of coumermycin . These results suggest that B . burgdorferi has a DNA gyrase and that this enzyme's activity is required for growth . Furthermore, structural analogs of coumermycin may be considered as treatments for Lyme disease.

Biosystems, 1993, 30(1-3), 113 - 35
Statistical analysis of L-tuple frequencies in eubacteria and organelles; Sitnikova TL et al.; This work is an attempt to study the structural features and evolutionary patterns of nucleotide sequences by analyzing their 1- through 4-plet frequencies and statistical relations between them . We present mathematical apparatus for this analysis . In particular, we introduce criteria to estimate the degree of homogeneity of L-plet composition in a given set of sequences and the dependence of the L-plet frequencies on the composition of lower orders . We apply these criteria to the study of eubacteria, mitochondria and chloroplasts . We demonstrate that L-plet frequencies are quite useful for revealing evolutionary relationship between DNA sequences and that the non-random distribution is more typical for doublets than to triplets . Non-randomness of triplet composition is more characteristic to coding than to non-coding regions, while no significant differences in dinucleotide composition can be observed . The obtained results can be used for revealing possible mechanisms of the codon usage phenomena.

Antonie Van Leeuwenhoek, 1993, 63(3-4), 275 - 88
Carbohydrate transport in bacteria under environmental conditions, a black box?
Lengeler JW.
A typical eubacterium carries a battery of substrate transport systems which are the ultimate pacemakers for growth . These systems reflect a billion year old selection for coping with rapidly changing conditions in the environment and each of them is optimised for specific growth conditions . Metabolic pathways in combination with transport systems can be interpreted as transient sensory systems, where a transport system corresponds to a sensor for external stimuli . Characteristics is a tightly linked common control between a carbohydrate metabolic pathway and the corresponding transport system . Many of the observed growth phenomena are a direct result of adaptation and regulation of transport capacity to rapid changes in environmental conditions . Some of the better understood examples are discusses . Nevertheless, knowledge on bacterial carbohydrate transport under environmental conditions as documented in the literature is still scarce.

Biosystems, 1993, 31(2-3), 127 - 30; discussion 130-3
Holobionts, hybrids, and cladistic classification; Zrzavy J et al.; David P . Mindell assumes that incorporation of the holobionts (evolutionarily stable symbiotic complexes) and hybrids into phylogenetic trees necessarily distorts the hierarchical structure of cladistic classification, and must lead to reformulation of some basic cladistic concepts (BioSystems, 27, 53-62, 1992) . He does not regard the Eukarya and Eubacteria as monophyletic taxa, the former being 'polyphyletic', the latter 'paraphyletic' . We attempt to show that the existence of holobionts/hybrids is not a cladistic problem . Cladistics is a systematic methodology, not a system of evolutionary hypotheses; cladograms are statements about distribution of shared characters (synapomorphies) and, consequently, the cladograms are always branching and hierarchically structured . A taxon is monophyletic if it has a single root in the cladogram, not a single ancestor 'in reality' . Cladistic methodology does not provide a clue for distinguishing whether a conflicting distribution of the potential synapomorphies is due to reticulation (e.g., symbiogenesis) or convergent evolution . Consequently, both Eubacteria and Eukarya either are or are not monophyletic taxa if they are or are not determined to be so during cladistic analysis.

Biosystems, 1993, 31(2-3), 111 - 9
Horizontal transfer of ATPase genes--the tree of life becomes a net of life; Hilario E et al.; An ancient gene duplication gave rise to the catalytic and non-catalytic subunits of each of the three types of proton pumping ATPases: vacuolar, archaebacterial and eubacterial . Previously, this gene duplication has been used to root the universal tree of life . However, recent findings of archaebacterial type ATPases in eubacteria and of eubacterial type in an archaebacterium suggested that both types of ATPases may have been already present in the last common ancestor . Here we show that a phylogenetic analysis of these ATPase subunits indicates that this conclusion is premature . We suggest that horizontal gene transfer can explain the data . In addition, we show that the analysis of glutamate dehydrogenases data neither affirm nor contradict any particular placement of the last common ancestor in the universal tree of life . The prevalence and the mode of horizontal gene transfer is discussed.

Reprod Nutr Dev, 1993, 33(6), 577 - 84
Effect of Eubacterium limosum, a ruminal hydrogenotrophic bacterium, on the degradation and fermentation of cellulose by 3 species of rumen anaerobic fungi; Bernalier A et al.; The degradation and fermentation of cellulose filter paper were studied in axenic cultures of 3 species of rumen anaerobic fungi, Neocallimastix frontalis, Piromyces communis and Caecomyces communis, and in cocultures containing 1 of these fungal strains and Eubacterium limosum, a hydrogenotrophic rumen bacterial species . When E limosum was introduced into fungal cultures a slight decrease in fungal cellulolytic activity was observed . The end products of the fermentation of cellulose found in the cocultures were different from those found in the fungal monocultures . E limosum used formate and part of the hydrogen produced by the fungi and probably created a shift in the metabolism of the fungi resulting in a reduction of lactate and ethanol production.

FASEB J, 1993 Jan, 7(1), 7 - 14
Recent studies of ribonuclease P; Altman S et al.; RNase P is an essential enzyme that is required for the biosynthesis of tRNA . It is composed of RNA and protein subunits . The RNA subunit of the enzyme derived from eubacterial sources can carry out the catalytic function by itself in vitro . Current studies of RNase P focus on structure-function relationships with respect to interactions of the RNA subunit with its substrates and with respect to the determination of the kinetic parameters of the reaction, the role of the protein component, and the rules governing recognition of substrates.

FEBS Lett, 1992 Dec 21, 314(3), 211 - 4
Nucleotide sequence of the genes for ribosomal proteins HS15 and HSH from Haloarcula marismortui: an archaeon-specific gene cluster; Arndt E et al.; The nucleotide sequences of the genes for two ribosomal proteins, HS15 and HSH, from the archaeon Haloarcula marismortui, have been determined . The genes were found in a cluster together with another open reading frame with a probable regulatory function . HS15 and HSH have counterparts in eucarya . HS15 is significantly homologous to S19 from frog (Xenopus laevis) . HSH is related to S37 from yeast (Saccharomyces cerevisiae) and S27 from fly (Drosophila melanogaster), as well as to other members of the S27 family . Eubacterial counterparts were not found, suggesting that these proteins are 'extra proteins' that are absent in eubacterial ribosomes.

J Biol Chem, 1992 Dec 15, 267(35), 25304 - 8
The primary structure of rat ribosomal protein S5 . A ribosomal protein present in the rat genome in a single copy; Kuwano Y et al.; The amino acid sequence of the rat 40 S ribosomal subunit protein S5 was deduced from the sequence of nucleotides in a recombinant cDNA and confirmed by the determination, directly from the protein, of 17 residues near the NH2 terminus . S5 has 204 amino acids; the molecular weight is 22,863 . The protein designated S5a has the same amino acid sequence as S5 except that it lacks the NH2-terminal 5 residues . It is not known whether the conversion of a portion of S5 to S5a is physiological or fortuitous . The mRNA for S5 has about 820 nucleotides . Hybridization of the S5 cDNA to digests of nuclear DNA indicates that the rat genome has only a single copy of the gene; this is in distinction to the mouse and human genomes which have three to six copies of the S5 gene . Rat ribosomal protein S5 is related to the eubacteria, the arachaebacteria, and the chloroplast family of S7 ribosomal proteins . There is a peptide of 16 residues at the carboxyl terminus of S5 that is highly conserved in 18 species spanning the three kingdoms and chloroplasts.

Nucleic Acids Res, 1992 Dec 11, 20(23), 6331 - 7
Analysis of the gene encoding the RNA subunit of ribonuclease P from cyanobacteria; Vioque A; The genes encoding the RNA subunit of ribonuclease P from the unicellular cyanobacterium Synechocystis sp . PCC 6803, and from the heterocyst-forming strains Anabaena sp . PCC 7120 and Calothrix sp . PCC 7601 were cloned using the homologous gene from Anacystis nidulans (Synechococcus sp . PCC 6301) as a probe . The genes and the flanking regions were sequenced . The genes from Anabaena and Calothrix are flanked at their 3'-ends by short tandemly repeated repetitive (STRR) sequences . In addition, two other sets of STRR sequences were detected within the transcribed regions of the Anabaena and Calothrix genes, increasing the length of a variable secondary structure element present in many RNA subunits of ribonuclease P from eubacteria . The ends of the mature RNAs were determined by primer extension and RNase protection . The predicted secondary structure of the three RNAs studied is similar to that of Anacystis and although some idiosyncrasies are observed, fits well with the eubacterial consensus.

Biochem Int, 1992 Dec, 28(4), 725 - 34
Relationship between substrate activity and pKa value of phenols on sulfotransferase from Eubacterium A-44; Konishi-Imamura L et al.; The relationship between the kinetics of the enzyme activity and the structural features of phenolic donor and of acceptor substrates was investigated with a sulfotransferase from Eubacterium A-44, a human intestinal bacterium . The enzyme catalyzed the transfer of the sulfate group from the sulfate esters of phenol having a lower pKa to phenols having a higher pKa . When the Km values for acceptor substrates were measured at their optimal pH, a linear plot for log10Km versus the pKa with a slope of 0.615 was obtained . In addition, it is considered that the effect of pH on the Km values for the various acceptors is due to ionization of free enzyme . The kinetic behavior of bacterial sulfotransferase differed from that of mammalian phenol sulfotransferase.

J Bacteriol, 1992 Dec, 174(24), 8008 - 15
Evidence for an S-layer protein pool in the peptidoglycan of Bacillus stearothermophilus; Breitwieser A et al.; Intact cells of Bacillus stearothermophilus PV72 revealed, after conventional thin-sectioning procedures, the typical cell wall profile of S-layer-carrying gram-positive eubacteria consisting of a ca . 10-nm-thick peptidoglycan-containing layer and a ca . 10-nm-thick S layer . Cell wall preparations obtained by breaking the cells and removing the cytoplasmic membrane by treatment with Triton X-100 revealed a triple-layer structure, with an additional S layer on the inner surface of the peptidoglycan . This profile is characteristic for cell wall preparations of many S-layer-carrying gram-positive eubacteria . Among several variants of strain PV72 obtained upon single colony isolation, we investigated the variant PV72 86-I, which does not exhibit an inner S layer on isolated cell walls but instead possesses a profile identical to that observed for intact cells . In the course of a controlled mild autolysis of isolated cell walls, S-layer subunits were released from the peptidoglycan of the variant and assembled into an additional S layer on the inner surface of the walls, leading to a three-layer cell wall profile as observed for cell wall preparations of the parent strain . In comparison to conventionally processed bacteria, freeze-substituted cells of strain PV72 and the variant strain revealed in thin sections a ca . 18-nm-wide electron-dense peptidoglycan-containing layer closely associated with the S layer . The demonstration of a pool of S-layer subunits in such a thin peptidoglycan layer in an amount at least sufficient for generating one coherent lattice on the cell surface indicated that the subunits must have occupied much of the free space in the wall fabric of both the parent strain and the variant . It can even be speculated that the rate of synthesis and translation of the S-layer protein is influenced by the packing density of the S-layer subunits in the periplasm of the cell wall delineated by the outer S layer and the cytoplasmic membrane . Our data indicate that the matrix of the rigid wall layer inhibits the assembly of the S-layer subunits which are in transit to the outside.

EMBO J, 1992 Dec, 11(12), 4369 - 78
Isolation and cloning of Omp alpha, a coiled-coil protein spanning the periplasmic space of the ancestral eubacterium Thermotoga maritima; Engel AM et al.; We have discovered a new oligomeric protein component associated with the outer membrane of the ancestral eubacterium Thermotoga maritima . In electron micrographs, the protein, Omp alpha, appears as a rod-shaped spacer that spans the periplasm, connecting the outer membrane to the inner cell body . Purification, biochemical characterization and sequencing of Omp alpha suggest that it is a homodimer composed of two subunits of 380 amino acids with a calculated M(r) of 43,000 and a pI of 4.54 . The sequence of the omp alpha gene indicates a tripartite organization of the protein with a globular NH2-terminal domain of 64 residues followed by a putative coiled-coil segment of 300 residues and a COOH-terminal, membrane-spanning segment . The predicted length of the coiled-coil segment (45 nm) correlates closely with the spacing between the inner and outer membranes . Despite sequence similarity to a large number of coiled-coil proteins and high scores in a coiled-coil prediction algorithm, the sequence of the central rod-shaped domain of Omp alpha does not have the typical 3.5 periodicity of coiled-coil proteins but rather has a periodicity of 3.58 residues . Such a periodicity was also found in the central domain of staphylococcal M protein and beta-giardin and might be indicative of a subclass of fibrous proteins with packing interactions that are distinct from the ones seen in other two-stranded coiled-coils.

Nucleic Acids Res, 1992 Nov 25, 20(22), 5919 - 25
Evolutionary conserved nucleotides within the E.coli 4.5S RNA are required for association with P48 in vitro and for optimal function in vivo; Wood H et al.; E.coli 4.5S RNA is homologous to domain IV of eukaryotic SPR7S RNA, the RNA component of the signal recognition particle . The 4.5S RNA is associated in vivo with a 48kD protein (P48), which is homologous to a protein component of the signal recognition particle, SRP54 . In addition to secondary structural features, a number of nucleotides are conserved between the 4.5S RNA and domain IV of all other characterised SRP-like RNAs from eubacteria, arachaebacteria and eukaryotes . This domain consists of an extended stem-loop structure; conserved nucleotides lie within the terminal loop and within single-stranded regions bulged from the stem immediately preceding the loop . This conserved region is a candidate for the SRP54/P48 binding site . To determine the functional importance of this region within the 4.5S RNA, mutations were introduced into the 4.5S RNA coding sequence . Mutated alleles were tested for their function in vivo and for the ability of the corresponding RNAs to bind P48 in vitro . Single point mutations in conserved nucleotides within the terminal tetranucleotide loop do not affect P48 binding in vitro and produce only slight growth defects . This suggests that the sequence of the loop may be important for the structure of the molecule rather than for specific interactions with P48 . On the other hand, nucleotides within the single-stranded regions bulged from the stem were found to be important both for the binding of P48 to the RNA and for optimal function of the RNA in vivo.

J Theor Biol, 1992 Nov 7, 159(1), 53 - 65
Macroevolution of plasmids: a model for plasmid speciation; Sykora P; A new evolutionary model for diversification in plasmid incompatibility groups (plasmid speciation) is suggested . The model is based on the formation of plasmid cointegrates from two compatible plasmids . The existence of plasmid cointegrates is well known, however, their potential key role in plasmid macroevolution has not yet been recognized . In a hypothesis presented here, one of the rep genes is supposed to be relaxed from selection in plasmid cointegrates and thus becomes free to accumulate mutations . These mutations can lead to a change in incompatibility specificity . Evidence supporting this hypothesis comes from the common occurrence of multi-replicon plasmids in nature as well as from experimental studies on plasmid cointegrate formation . A more speculative extension of this model hypothesizes an evolutionary scenario for origin of the eubacterial single-replicon genome and the eukaryotic multi-replicon genome, as well as the place of plasmids and viruses in this picture.

Gene, 1992 Nov 2, 121(1), 155 - 60
Isolation and characterization of the gene coding for the major sigma factor of Rickettsia prowazekii DNA-dependent RNA polymerase; Marks GL et al.; The gene coding for the major sigma factor of Rickettsia prowazekii, an obligate intracellular parasitic bacterium, has been isolated utilizing an oligodeoxyribonucleotide as a probe to a conserved region of major sigma factors . Nucleotide sequence analysis revealed an open reading frame of 1905 bp that could encode a protein of 635 amino acids (aa) with a calculated molecular size of 73 kDa (sigma 73) . R . prowazekii sigma 73 displayed extensive homology with major sigma factors from a variety of eubacteria . Comparison of the major sigma factors from Escherichia coli and R . prowazekii revealed 44.9% aa identity . R . prowazekii sigma 73 produced in E . coli minicells migrated as a 85-kDa protein when analyzed by sodium dodecyl sulfate-polyacrylamide-gel electrophoresis . This anomalous migration is characteristic of eubacterial major sigma factors and agrees with the migration noted for the purified rickettsial sigma protein . Despite a similarity to the E . coli sigma 70 encoded by rpoD, R . prowazekii sigma 73 did not complement E . coli rpoD temperature-sensitive mutants.

J Exp Biol, 1992 Nov, 172, 137 - 47
Evolution and isoforms of V-ATPase subunits; Gogarten JP et al.; The structure of V- and F-ATPases/ATP synthases is remarkably conserved throughout evolution . Sequence analyses show that the V- and F-ATPases evolved from the same enzyme that was already present in the last common ancestor of all known extant life forms . The catalytic and non-catalytic subunits found in the dissociable head groups of both V-ATPases and F-ATPases are paralogous subunits, i.e . these two types of subunits evolved from a common ancestral gene . The gene duplication giving rise to these two genes (i.e . those encoding the catalytic and non-catalytic subunits) pre-dates the time of the last common ancestor . Similarities between the V- and F-ATPase subunits and an ATPase-like protein that is implicated in flagellar assembly are evaluated with regard to the early evolution of ATPases . Mapping of gene duplication events that occurred in the evolution of the proteolipid, the non-catalytic and the catalytic subunits onto the tree of life leads to a prediction of the likely quaternary structure of the encoded ATPases . The phylogenetic implications of V-ATPases found in eubacteria are discussed . Different V-ATPase isoforms have been detected in some higher eukaryotes, whereas others were shown to have only a single gene encoding the catalytic V-ATPase subunit . These data are analyzed with respect to the possible function of the different isoforms (tissue-specific, organelle-specific) . The point in evolution at which the different isoforms arose is mapped by phylogenetic analysis.

Appl Environ Microbiol, 1992 Nov, 58(11), 3622 - 9
Hydrogen as an electron donor for dechlorination of tetrachloroethene by an anaerobic mixed culture; DiStefano TD et al.; Hydrogen served as an electron donor in the reductive dechlorination of tetrachloroethene to vinyl chloride and ethene over periods of 14 to 40 days in anaerobic enrichment cultures; however, sustained dechlorination for more extended periods required the addition of filtered supernatant from a methanol-fed culture . This result suggests a nutritional dependency of hydrogen-utilizing dechlorinators on the metabolic products of other organisms in the more diverse, methanol-fed system . Vancomycin, an inhibitor of cell wall synthesis in eubacteria, was found to inhibit acetogenesis when added at 100 mg/liter to both methanol-fed and hydrogen-fed cultures . The effect of vancomycin on dechlorination was more complex . Methanol could not sustain dechlorination when vancomycin inhibited acetogenesis, while hydrogen could . These results are consistent with a model in which hydrogen is the electron donor directly used for dechlorination by organisms resistant to vancomycin and with the hypothesis that the role of acetogens in methanol-fed cultures is to metabolize a portion of the methanol to hydrogen . Methanol and other substrates shown to support dechlorination in pure and mixed cultures may merely serve as precursors for the formation of an intermediate hydrogen pool . This hypothesis suggests that, for bioremediation of high levels of tetrachloroethene, electron donors that cause the production of a large hydrogen pool should be selected or methods that directly use H2 should be devised.

FEMS Microbiol Lett, 1992 Nov 1, 77(1-3), 161 - 8
Analysis of genomic DNA from Chlamydia trachomatis for Dam and Dcm methylation; Wagar EA et al.; Chlamydia trachomatis is a Gram-negative eubacterium with a dimorphic developmental cycle and obligate intracellular growth in the eucaryotic host . The Dam transmethylase of Escherichia coli methylates at the N6 position of adenine in the sequence 5'-GATC-3' and the Dcm transmethylase adds methyl groups to the C5 position of the internal cytosines in the sequences 5'-CCWGG-3' . In contrast to E . coli, C . trachomatis DNA appears to have unmethylated Dam sites and only low level Dcm methylation.

Biosci Biotechnol Biochem, 1992 Nov, 56(11), 1797 - 800
Purification and characterization of a DNA-dependent RNA polymerase from Pseudomonas putida; Fujita M et al.; DNA-dependent RNA polymerase (EC 2.7.7.6) was purified from Pseudomonas putida . The enzyme had the typical composition of beta',beta,alpha, and sigma subunits of eubacterial RNA polymerases . The molecular masses of the subunits were 156,000 Da, 151,000 Da, 87,000 Da, and 42,000 Da, respectively, as measured by polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate . The NH2-terminal amino acid residues of the alpha subunit had a marked homology with those of the alpha subunit of Escherichia coli RNA polymerase . The enzyme activity was dependent on ribonucleoside triphosphates, Mg2+, and a DNA template, and was inhibited in vitro by rifampicin . The enzyme activity was maximal in the presence of 10 mM MgCl2 . In an in vitro transcription assay using the tac promoter-controlled DNA as a template, the RNA polymerase of P . putida initiated transcription at the same site as that of E . coli.

Can J Microbiol, 1992 Nov, 38(11), 1091 - 6
Bacterial conjugation: everybody's doin' it; Frost LS; Recent evidence suggests that bacterial conjugation is ubiquitous in the bacterial world and that DNA transfer between different genera-kingdoms is possible . It has also been demonstrated that many bacterial gene transfer systems resemble each other at the molecular level and that others are a blend of two or more of these systems . Thus, in the absence of a sexual cycle, bacterial conjugation, along with bacteriophages, transposons, and natural transformation systems, forms a potent force for gene dissemination and repair in the eubacteria and simple eukaryotes.

J Endod, 1992 Nov, 18(11), 558 - 61
The relationship between clinical symptoms and anaerobic bacteria from infected root canals; Hashioka K et al.; The purpose of this study was to investigate the correlation between the composition of bacterial flora from infected root canals and clinical symptoms . The materials evaluated consisted of 28 teeth from 25 patients with apical periodontitis . Eubacterium were found to be significantly related to acute or chronic clinical symptoms and Peptococcus, Peptostreptococcus, and Porphyromonas gingivalis to subacute clinical symptoms . We suggested that Peptococcus, Peptostreptococcus, Eubacterium, Porphyromonas, and Bacteroides were significantly related to percussion pain; Porphyromonas and Bacteroides were significantly related to odor in the infected root canals . Many Bacteroides were isolated from most of the infected root canals.

Gene, 1992 Oct 12, 120(1), 33 - 41
The sequence of the gene encoding elongation factor Tu from Chlamydia trachomatis compared with those of other organisms; Cousineau B et al.; Nucleotide (nt) sequences encoding the elongation factor Tu (EF-Tu), tRNA(Thr) and tRNA(Trp) from Chlamydia trachomatis have been determined . The environment of the EF-Tu-encoding gene (tuf), between two tRNA gene sequences, suggests that it is part of a tufB locus . The nt sequence and the deduced amino acid (aa) sequence were aligned with comparable sequences from other organisms and the resulting data bases were used to infer phylogenies . Phylogenetic trees based on aa sequences and nt sequences are similar, but not completely congruent with rRNA gene-based phylogenies . Both the nt and aa sequence trees concur on the early divergence of Thermotoga and Chlamydia from the bacterial root . The aa alignment highlights the presence of four unique Cys residues in the chlamydial sequence which are found at strictly conserved positions in other sequences . Further peculiarities of the chlamydial and eubacterial sequences have been mapped to the X-ray crystallographic structure of the protein.

Oral Microbiol Immunol, 1992 Oct, 7(5), 285 - 90
Coaggregation studies of the Eubacterium species; George KS et al.; Eubacterium species are gram-positive anaerobic rods that are frequently isolated from subgingival plaque of periodontal pockets . Five Eubacterium species were tested for their ability to coaggregate with 33 oral bacterial strains . Using visual and turbidimetric assays, coaggregation was observed among Eubacterium brachy, Eubacterium nodatum, Eubacterium alactolyticum and Eubacterium limosum strains only when tested with Fusobacterium nucleatum strains; Eubacterium saburreum displayed only weak coaggregation ability . Coaggregation between F . nucleatum and the Eubacterium species was observed over a wide range of concentrations of each organism . The F . nucleatum strains contained a heat labile and the Eubacterium species a heat stabile coaggregation receptor . Arginine, histidine, lysine and glycine inhibited the coaggregation between F . nucleatum and the Eubacterium species . Sugars and other amino acids tested did not inhibit the observed coaggregation . Rabbit anti-F . nucleatum serum completely inhibited coaggregation, but anti-E . brachy serum and normal rabbit serum did not . As these anaerobic microorganisms are frequently isolated from the same oral lesions, the surface interactions observed may be important in the pathogenesis of these polymicrobic infections.

Oral Microbiol Immunol, 1992 Oct, 7(5), 257 - 62
Associations between microbial species in dental root canal infections; Sundqvist G; The existence of commensal or antagonistic relationships between microorganisms in the root canals of teeth with apical periodontitis was investigated . Samples were taken from 65 infected human root canals and were analysed according to species, frequency of occurrence and proportion of the total isolated flora . The most frequent species were Fusobacterium nucleatum, Prevotella intermedia, Peptostreptococcus micros, Peptostreptococcus anaerobius, Eubacterium alactolyticum, Eubacterium lentum and Wolinella recta . An odds ratio system was used to calculate positive or negative associations between the isolated bacteria . Strong positive associations were found between F . nucleatum and P . micros, Porphyromonas endodontalis, Selenomonas sputigena and W . recta . There was also a positive association between P . intermedia and P . micros, P . anaerobius and the eubacteria . In general, species of streptococci, Propionibacterium propionica, Capnocytophaga ochracea and Veillonella parvula showed no or negative associations with the other bacteria . The results are consistent with the concept of a special and selective environment occurring in the root canal that is due, in part, to the cooperative as well as antagonistic nature of the relationships between bacteria in the root canal.

Appl Environ Microbiol, 1992 Oct, 58(10), 3413 - 6
Amplification of DNA from native populations of soil bacteria by using the polymerase chain reaction; Bruce KD et al.; Specific DNA sequences from native bacterial populations present in soil, sediment, and sand samples were amplified by using the polymerase chain reaction with primers for either "universal" eubacterial 16S rRNA genes or mercury resistance (mer) genes . With standard amplification conditions, 1.5-kb rDNA fragments from all 12 samples examined and from as little as 5 micrograms of soil were reproducibly amplified . A 1-kb mer fragment from one soil sample was also amplified . The identity of these amplified fragments was confirmed by DNA-DNA hybridization.

Mol Microbiol, 1992 Oct, 6(19), 2797 - 804
Transcriptional regulation of the redD transcriptional activator gene accounts for growth-phase-dependent production of the antibiotic undecylprodigiosin in Streptomyces coelicolor A3(2); Takano E et al.; Transcription of redD, the activator gene required for production of the red-pigmented antibiotic undecylprodigiosin by Streptomyces coelicolor A3(2), showed a dramatic increase during the transition from exponential to stationary phase . The increase in redD expression was followed by transcription of redX, a biosynthetic structural gene, and the appearance of the antibiotic in the mycelium, and coincided with the intracellular appearance of ppGpp . However, ppGpp production elicited either by nutritional shift-down of, or addition of serine hydroxamate to, exponentially growing cultures had no stimulatory effect on redD transcription . The presence of redD on a multicopy plasmid resulted in elevated levels of the redD transcript and production of redX and undecylprodigiosin during exponential growth; the normal growth-phase-dependent production of undecylprodigiosin appeared to be mediated entirely through the redD promoter, which shows limited similarity to the consensus sequence for the major class of eubacterial promoters.

J Gen Virol, 1992 Oct, 73 ( Pt 10), 2763 - 6
Geminivirus replication proteins are related to prokaryotic plasmid rolling circle DNA replication initiator proteins; Koonin EV et al.; It is demonstrated, by means of computer-assisted analysis, that C1 protein involved in the replication of geminivirus DNA is related to the rolling circle replication initiator proteins of eubacterial plasmids, particularly the plasmids of the pMV158 family . Three sequence motifs conserved in the geminivirus and plasmid replication proteins were delineated, one of them encompassing the Tyr residue that presumably forms a covalent linkage to DNA . These findings are compatible with the results of recent analyses of geminivirus replicative intermediates suggesting a rolling circle mechanism for geminivirus DNA replication . It is hypothesized that C1 protein initiates the rolling circle replication of geminivirus DNA by nicking a specific site in the virus-sense DNA and covalently linking to the 5' side of the nick . The putative rolling circle replication initiator domain comprises the N-terminal portion of C1, whereas its C-terminal part is a putative helicase domain . By analogy with prokaryotic systems, it is speculated that the replication initiator domain and the helicase domain function coordinately . The possibility of the origin of geminiviruses from prokaryotic circular ssDNA replicons is discussed.

J Bacteriol, 1992 Oct, 174(19), 6317 - 20
Characterization of the gill symbiont of Thyasira flexuosa (Thyasiridae: Bivalvia) by use of polymerase chain reaction and 16S rRNA sequence analysis; Distel DL et al.; Comparative molecular sequence (16S rRNA) analysis methods were used to identify and characterize the symbionts of Thyasira flexuosa independently of pure culture techniques and to compare these symbionts with the previously reported putative symbiont isolate, Thiobacillus thyasiris TG-2 (A . P . Wood and D . P . Kelly, Arch . Microbiol . 152:160-166, 1989) . Polymerase chain reaction amplification using 16S rRNA primers specific for eubacteria was used to amplify a single unique sequence from the gill tissue of T . flexuosa . This sequence is phylogenetically most closely related to the 16S rRNA genes of known symbionts of lucinid clams and is distinct from those determined for strain TG-2 and other known bacteria . Strain TG-2 most closely resembles a free-living, chemolithoautotrophic bacterium known to be associated with the surfaces of thiotrophic bivalve shells, suggesting that this strain is a contaminant and not the authentic intracellular symbiont of T . flexuosa.

J Bacteriol, 1992 Oct, 174(19), 6103 - 8
Evidence that a plasmid from a hyperthermophilic archaebacterium is relaxed at physiological temperatures; Charbonnier F et al.; A plasmid of 3.45 kb (pGT5) was recently discovered in a strain of hyperthermophilic archaebacterium which was isolated from samples collected in a deep-sea hydrothermal vent . This strain (GE5) grows within a temperature range of 68 to 101.5 degrees C, and we show here that it contains a strong ATP-dependent reverse gyrase activity (positive DNA supercoiling) . By comparison with eubacterial plasmids of known superhelical densities, we estimated the superhelical density of the archaebacterial plasmid pGT5 to be -0.026 at 25 degrees C . The equation which relates the change of the rotation angle of the DNA double helix with temperature was validated at 95 degrees C, the optimal growth temperature of the GE5 strain . Considering these new data, the superhelical density of plasmid pGT5 was calculated to be -0.006 at the physiological temperature of 95 degrees C, which is close to the relaxed state . This finding shows that the DNA topology of a plasmid isolated from a hyperthermophilic archaebacterium containing reverse gyrase activity is strikingly different from that of typical eubacterial plasmids.

Eur J Biochem, 1992 Oct 1, 209(1), 351 - 5
Cloning and sequencing of the gene for the cytoplasmic inorganic pyrophosphatase from the thermoacidophilic archaebacterium Thermoplasma acidophilum; Richter OM et al.; The gene (ppa) from the thermoacidophilic archaebacterium Thermoplasma acidophilum, encoding the cytoplasmic pyrophosphatase, has been cloned . Two degenerate oligonucleotide probes, synthesized according to the N-terminal amino acid sequence of the isolated protein, were used to screen subgenomic libraries . The DNA-derived amino acid sequence of the archaebacterial enzyme allows, for the first time, comparative studies of cytoplasmic pyrophosphatases to be extended to all three urkingdoms . The archaebacterial pyrophosphatase more closely resembles the eubacterial enzymes on the basis of sequence similarity and subunit size . The majority of amino acid residues considered to be essential for hydrolysis of pyrophosphate seem to have been conserved throughout evolution, as inferred from the results of an alignment of sequences from all three urkingdoms.

Eur J Biochem, 1992 Oct 1, 209(1), 343 - 9
Purification and enzymic characterization of the cytoplasmic pyrophosphatase from the thermoacidophilic archaebacterium Thermoplasma acidophilum; Richter OM et al.; Cytoplasmic pyrophosphatase has been isolated from the thermoacidophilic archaebacterium Thermoplasma acidophilum . The enzyme was purified to electrophoretic homogeneity by combining ion-exchange and affinity-chromatographic separations . This soluble pyrophosphatase probably consists of six identical subunits, since SDS/PAGE gave an estimate of about 22 kDa for a single subunit and size-exclusion chromatography under non-denaturing conditions indicates a molecular mass of 110 +/- 5 kDa . The two most prominent catalytic features of this enzyme are the absolute requirement for divalent cations for catalytic action, Mg2+ conferring the highest activity, and the pronounced specificity for PPi . The catalytic behavior apparently follows simple Michaelis-Menten kinetics with a Km of about 7 microM for PPi and a specific activity of about 1200 U/mg at 56 degrees C . Surprisingly, maximum activity could be observed at 85 degrees C which is more than 20 degrees C above the temperature for optimal growth . Several cytoplasmic extracts of eubacteria and archaebacteria have been probed with a polyclonal antiserum raised against the purified archaebacterial protein . The only noticeable cross-reactivity could be detected with an extract from the methanogen Methanosarcina barkeri although this probably does not reflect the inferred phylogenetic relationship between methanogens and Thermoplasma acidophilum.

Gene, 1992 Sep 21, 119(1), 113 - 8
Sequence analysis of a DNA fragment from Buchnera aphidicola (an endosymbiont of aphids) containing genes homologous to dnaG, rpoD, cysE, and secB; Lai CY et al.; The aphid, Schizaphis graminum, contains a prokaryotic, obligately intracellular endosymbiont, Buchnera aphidicola, which is necessary for the survival of the host . A recent study of Bu . aphidicola 16S rRNA has indicated that it is a member of the gamma-3 subdivision of the eubacterial class, Proteobacteria, which includes Escherichia coli . In order to further characterize the endosymbiont and establish its similarity to free-living eubacteria and/or organelles, we have cloned and sequenced a 4534-bp DNA fragment containing dnaG-rpoD-cysE-secB . The deduced amino acid (aa) sequence identity to the homologous E . coli proteins ranged from 47 to 80% . The close proximity of the pair, dnaG-rpoD, to the pair, cysE-secB, on the Bu . aphidicola DNA, differed from E . coli, in which these two pairs of genes are 14 min apart on the bacterial chromosome . The results of past physiological studies of the endosymbiont were consistent with the presence and function of DNA primase (DnaG), sigma factor (RpoD) and components of the secretory system (SecB) . Comparison of the deduced aa sequence of Bu . aphidicola CysE (serine acetyltransferase, a key allosterically regulated enzyme in cysteine biosynthesis) with the E . coli wild-type enzyme and a mutant defective in feedback inhibition suggested that the endosymbiont CysE may not be regulated . By analogy with E . coli, the lack of feedback inhibition may lead to overproduction of cysteine by the endosymbiont . The results of this and previous investigations indicate that Bu . aphidicola has many of the properties of free-living bacteria and not of organelles.

Scand J Immunol, 1992 Sep, 36(3), 497 - 506
Immunohistology of joint inflammation induced in rats by cell wall fragments of Eubacterium aerofaciens; Kool J et al.; After a single intraperitoneal injection of cell wall fragments of Eubacterium aerofaciens, a main resident from the human intestinal flora, an acute arthritis develops within 2 days which is followed by a chronic arthritis that lasts at least 90 days . In an earlier report the histological appearance of the joint inflammation during this period has been described . In this study we investigated in more detail the cell types that are involved in the development of arthritis by using cell-type-specific monoclonal antibodies in an immunohistological assay . In the acute phase of arthritis, T-helper cells appeared in the synovial tissue together with ED1-positive (ED1+) and ED3-positive (ED3+) macrophages . After a temporary decline at day 12 all macrophage subsets, as well as T-helper cells, reappeared or increased again at day 33 . Later, in the chronic phase (days 47-90), an increased number of ED1-positive (ED1+) cells in the synovial tissue and a decreased number of ED2-positive (ED2+) cells in the synovial lining was the most prominent finding when compared with control rats . These results indicate that, apart from T lymphocytes, macrophages also play an important role in the development and continuation of chronic arthritis in this model.

Acta Stomatol Belg, 1992 Sep, 89(3), 155 - 62
Susceptibility of anaerobic microorganisms to hypothiocyanite produced by lactoperoxidase; Courtois P et al.; The susceptibility of Capnocytophaga ochracea, Eikenella corrodens, Eubacterium yurii, Fusobacterium nucleatum, Peptostreptococcus micros, Prevotella intermedia, Selenomonas sputigena, Wolinella recta to hypothiocyanite (OSCN-) produced by the lactoperoxidase system was tested . Results showed a decrease of bacterial survival rate after OSCN- exposure, with an intra- and inter-species variability from 0 to 95% for C . ochracea, 34-100% for E . corrodens, 0-83% for E . yurii, 1-15% for F . nucleatum, 8-61% for P . micros, 0-100% for P . intermedia, 0-44% for S . sputigena and 0-8% for W . recta . The survival rate did not correlate with the NADH/OSCN- oxidoreductase activity present in the lysed bacteria (r = -0.3248; N = 15; NS).

Plasmid, 1992 Sep, 28(2), 170 - 6
DNA sequence and analysis of a cryptic 4.2-kb plasmid from the filamentous cyanobacterium, Plectonema sp . strain PCC 6402; Perkins DR et al.; The 4194-bp plasmid, pRF1, from Plectonema sp . Strain PCC 6402 was completely sequenced and analyzed . Seven potential open reading frames were identified . The predicted amino acid sequence of open reading frame C (ORF C) had identities of 34, 29, and 25% with Rep B from the Staphylococcus aureus plasmid, pUB110; Rep from the Bacillus amyloliquefaciens plasmid, pFTB14; and protein A from the S . aureus plasmid, pC194, respectively . A 75-amino-acid region conserved in these proteins (Rep B, Rep, and protein A) also was highly conserved in ORF C with identities of 45, 37, and 40%, respectively . Significantly, 16 of the 21 amino acids conserved in Rep B, Rep, and protein A were found at the same positions in ORF C . This ORF may encode a replication protein that includes a region conserved in some eubacteria . Additional structural features include a 425-bp region that contains palindromes, tandem repeats, and short direct repeats which may correspond to the origin of replication . An 18-bp inverted repeat was located between two open reading frames, A and G.

J Gen Microbiol, 1992 Sep, 138 ( Pt 9), 1875 - 80
Cloning and physical mapping of the Staphylococcus aureus rplL, rpoB and rpoC genes, encoding ribosomal protein L7/L12 and RNA polymerase subunits beta and beta'; Aboshkiwa M et al.; Using segments of the Escherichia coli rpoB and rpoC genes as heterologous probes, we have identified and cloned an 8.3 kb PstI fragment from the Staphylococcus aureus genome containing the rpoB and rpoC genes, which respectively encode the beta and beta' subunits of DNA-directed RNA polymerase . This region is almost certainly equivalent to the rif locus, located near to fus at interval 12/13 on the S . aureus linkage map . Limited DNA sequencing revealed the gene order rpoB-rpoC (transcribed from left to right) and identified the rplL gene, encoding ribosomal protein L7/L12, upstream of rpoB . This and other evidence suggests that the rpoBC genes of S . aureus form part of a large gene cluster encoding components of the transcription and translation apparatus which is well-conserved in other eubacteria.

Curr Opin Dent, 1992 Sep, 2, 66 - 71
Bacteriology of periodontal disease; Russell RR; The microbial flora associated with the periodontal tissues in health and disease is extremely complex, and much research is being directed toward identifying those species that may be etiologic agents or that can be used as prognostic indicators . Recent work has resulted in changes in the taxonomic position of several periodontal species and the recognition that several others, particularly species of Eubacterium and Peptostreptococcus, as well as a novel oral spirochete, may be important in disease . New rapid techniques for identifying and enumerating the periodontal flora are being applied to cross-sectional and longitudinal studies to assess the significance of the various species; the DNA probe and immunologic detection methods demonstrate both advantages and limitations when compared with methods based on determining the predominant cultivable flora.

Biochem Biophys Res Commun, 1992 Aug 31, 187(1), 58 - 64
The primary structure of rat ribosomal protein L3; Kuwano Y et al.; The amino acid sequence of the rat 60S ribosomal subunit protein L3 was deduced from the sequence of nucleotides in a recombinant cDNA . Ribosomal protein L3 has 403 amino acids and has a molecular weight of 46,106 . Hybridization of the cDNA to digests of nuclear DNA suggests that there are 7 to 9 copies of the L3 gene . The mRNA for the protein is about 1,400 nucleotides in length . Rat L3 is homologous to ribosomal proteins from other eukaryotes and to proteins from eubacterial, archaebacterial, and chloroplast ribosomes.

Biochim Biophys Acta, 1992 Aug 17, 1132(1), 94 - 6
The complete nucleotide sequence of the gene (rpoD1) encoding the principal sigma factor of the RNA polymerase from the cyanobacterium Synechococcus sp . strain PCC7942; Tanaka K et al.; The complete nucleotide sequence of rpoD1 gene from Synechococcus PCC7942 has been determined . The nucleotide data have indicated the presence of an open reading frame of 1155 base pairs encoding a polypeptide which shares the framework structure for principal sigma factors of eubacterial strains.

Eur J Biochem, 1992 Aug 1, 207(3), 877 - 85
Primary structures of ribosomal proteins L3 and L4 from Bacillus stearothermophilus; Herwig S et al.; Ribosomal proteins L3 and L4 were purified to homogeneity from total protein isolated from the 50S subunit of Bacillus stearothermophilus by reversed-phase high-performance liquid chromatography (RP-HPLC) . Amino acid sequences of both proteins were determined by automated N-terminal sequence analysis and sequencing of internal peptides . Using oligonucleotides deduced from the N-terminal region of protein L3 as hybridization probes, a DNA fragment coding for proteins L3, L4 and the N-terminal part of protein L23 has been identified, cloned and sequenced . The organization of the genes is identical to that found in the S10 operon of Escherichia coli . Comparison of the sequences of proteins L3 and L4 with those of other organisms revealed that all proteins of the L3 family are highly conserved . On the other hand, the archaebacterial L4 proteins show no significant sequence similarity to the E . coli L4 protein whereas the L4 protein of B . stearothermophilus is significantly similar to all of the L4 proteins and thus justifies the membership of all the L4 proteins in one protein family . The results are discussed with respect to the phylogenetic relationship between eubacteria, archaebacteria and eukaryotes and possible functional domains of proteins L3 and L4.

J Dent Res, 1992 Aug, 71(8), 1509 - 15
Immunoglobulins in milk from cows immunized with oral strains of Actinomyces, Prevotella, Porphyromonas, and Fusobacterium; Takahashi N et al.; Immunization of pregnant cows with bacteria leads to the presence of high concentrations of specific antibodies in colostrum and milk . A total of 14 cows was immunized with single strains of heat-killed oral bacteria or pools of strains of Actinomyces, Porphyromonas, Prevotella, and Fusobacterium . Two cows were treated with adjuvant alone . The mean percentages of IgG1, IgG2, IgM, and IgA in all of the milks were 83.8, 3.8, 9.3, and 3.1, respectively . ELISA and whole cell agglutination assays demonstrated high titers in the milks from the cows immunized with either individual strains or the bacterial pools . The highest titers determined by ELISA belonged to the IgG1 isotype and in several milks were 64-fold greater than titers in milk from cows treated with adjuvant alone . The concentrations of all antibodies and the titers determined by ELISA and whole cell agglutination assays markedly decreased from the first to the sixth milkings . The functional specificity of the antibodies was demonstrated by agglutination tests against a wide range of bacteria including members of Actinomyces, Fusobacterium, Porphyromonas, Prevotella, Streptococcus, Eubacterium, Propionibacterium, Peptostreptococcus, Bacteroides, Actinobacillus, Haemophilus, Capnocytophaga, and Wolinella . Minimal cross-reactions with bacteria in other genera were observed with all of the milks . High-titer milk preparations have been obtained from immunized cows, and the capacity of the bovine antibodies to agglutinate target bacteria indicates their potential usefulness in oral passive immunization studies.

J Mol Evol, 1992 Aug, 35(2), 147 - 55
Phylogenetic analysis of the thiolase family . Implications for the evolutionary origin of peroxisomes; Igual JC et al.; The thiolase family is a widespread group of proteins present in prokaryotes and three cellular compartments of eukaryotes . This fact makes this family interesting in order to study the evolutionary process of eukaryotes . Using the sequence of peroxisomal thiolase from Saccharomyces cerevisiae recently obtained by us and the other known thiolase sequences, a phylogenetic analysis has been carried out . It shows that all these proteins derived from a primitive enzyme, present in the common ancestor of eubacteria and eukaryotes, which evolved into different specialized thiolases confined to various cell compartments . The evolutionary tree obtained is compatible with the endosymbiotic theory for the origin of peroxisomes.

Nucleic Acids Res, 1992 Jul 11, 20(13), 3279 - 85
Conserved sequence motifs in the initiator proteins for rolling circle DNA replication encoded by diverse replicons from eubacteria, eucaryotes and archaebacteria; Ilyina TV et al.; An amino acid motif was identified that consists of the sequence HisHydrHisHydrHydrHydr (Hydr--bulky hydrophobic residue) and is conserved in two vast classes of proteins, one of which is involved in initiation and termination of rolling circle DNA replication, or RCR (Rep proteins), and the other in mobilization (conjugal transfer) of plasmid DNA (Mob proteins) . Based on analogies with metalloenzymes, it is hypothesized that the two conserved His residues in this motif may be involved in metal ion coordination required for the activity of the Rep and Mob proteins . Rep proteins contained two additional conserved motifs, one of which was located upstream, and the other downstream from the 'two His' motif . The C-terminal motif encompassed the Tyr residue(s) forming the covalent link with nicked DNA . Mob proteins were characterized by the opposite orientation of the conserved motifs, with the (putative) DNA-linking Tyr being located near their N-termini . Both Rep and Mob protein classes further split into several distinct families . Although it was not possible to find a motif or pattern that would be unique for the entire Rep or Mob class, unique patterns were derived for large subsets of the proteins of each class . These observations allowed the prediction of the amino acid residues involved in DNA nicking, which is required for the initiation of RCR or conjugal transfer of single-stranded (ss) DNA, in Rep and Mob proteins encoded by a number of replicons of highly diverse size, structure and origin . It is conjectured that recombination has played a major part in the dissemination of genes encoding related Rep or Mob proteins among the replicons exploiting RCR . It is speculated that the eucaryotic small ssDNA replicons encoding proteins with the conserved RCR motifs and replicating via RCR-related mechanisms, such as geminiviruses and parvoviruses, may have evolved from eubacterial replicons.

Mol Biol Evol, 1992 Jul, 9(4), 753 - 69
Sample size for a phylogenetic inference; Churchill GA et al.; The objective of this work is to describe sample-size calculations for the inference of a nonzero central branch length in an unrooted four-species phylogeny . Attention is restricted to independent binary characters, such as might be obtained from an alignment of the purine-pyrimidine sequences of a nucleic acid molecule . A statistical test based on a multinomial model for character-state configurations is described . The importance of including invariable sites in models for sequence change is demonstrated, and their effect on sample size is quantified . The methods are applied to a four-species alignment of small-subunit rRNA sequences derived from two archaebacteria, a eubacteria and a eukaryote . We conclude that the information in these sequences is not sufficient to resolve the branching order of this tree . Estimates of the number of aligned nucleotide positions required to provide a reasonably powerful test are given.

Gene, 1992 Jul 1, 116(1), 21 - 6
'Stop-codon-specific' restriction endonucleases: their use in mapping and gene manipulation; Rowland GC et al.; Certain restriction endonucleases recognise target sequences that contain the stop triplet TAG and are commonly either 4 or 6 bp in length . Interestingly, these restriction targets do not occur at the frequency expected on the basis of base composition and size . For example, the tetranucleotide MaeI recognition sequence (CTAG) occurs considerably less commonly (5-8-fold) in the genome of Escherichia coli (and many other eubacteria) than expected from mononucleotide frequencies . This surprising rarity is particularly evident in protein-encoding genes and is largely dictated by codon usage . Thus, amber (TAG) nonsense mutations frequently give rise to novel MaeI (CTAG) sites which are unique within a translated region . Such amber/MaeI sites, whether arising spontaneously or created in vitro by site-directed mutagenesis, act as a useful physical marker for the presence of the nonsense mutation and are a convenient startpoint for a range of diverse procedures . These features provide a useful supplement to protein engineering methods which use nonsense suppression to mediate amino acid replacements.

J Bacteriol, 1992 Jul, 174(14), 4594 - 605
Cloning of the HSP70 gene from Halobacterium marismortui: relatedness of archaebacterial HSP70 to its eubacterial homologs and a model for the evolution of the HSP70 gene; Gupta RS et al.; Heat shock induces the synthesis of a set of proteins in Halobacterium marismortui whose molecular sizes correspond to the known major heat shock proteins . By using the polymerase chain reaction and degenerate oligonucleotide primers for conserved regions of the 70-kDa heat shock protein (HSP70) family, we have successfully cloned and sequenced a gene fragment containing the entire coding sequence for HSP70 from H . marismortui . HSP70 from H . marismortui shows between 44 and 47% amino acid identity with various eukaryotic HSP70s and between 51 and 58% identity with its eubacterial and archaebacterial homologs . On the basis of a comparison of all available HSP70 sequences, we have identified a number of unique sequence signatures in this protein family that provide a clear distinction between eukaryotic organisms and prokaryotic organisms (archaebacteria and eubacteria) . The archaebacterial (viz., H . marismortui and Methanosarcina mazei) HSP70s have been found to contain all of the signature sequences characteristic of eubacteria (particularly the gram-positive bacteria), which suggests a close evolutionary relationship between these groups . In addition, detailed analyses of HSP70 sequences that we have carried out have revealed a number of additional novel features of the HSP70 protein family . These include (i) the presence of an insertion of about 25 to 27 amino acids in the N-terminal quadrants of all known eukaryotic and prokaryotic HSP70s except those from archaebacteria and the gram-positive group of bacteria, (ii) significant sequence similarity in HSP70 regions comprising its first and second quadrants from organisms lacking the above insertion, (iii) highly significant similarity between a protein, MreB, of Escherichia coli and the N-terminal half of HSP70s, (iv) significant sequence similarity between the N-terminal quadrant of HSP70 (from gram-positive bacteria and archaebacteria) and the m-type thioredoxin of plant chloroplasts . To account for these and other observations, a model for the evolution of HSP70 proteins involving gene duplication is proposed . The model proposes that HSP70 from archaebacteria (H . marismortui and M . mazei) and the gram-positive group of bacteria constitutes the ancestral form of the protein and that all other HSP70s (viz., other eubacteria as well as eukaryotes) containing the insert have evolved from this ancient protein.

Ultramicroscopy, 1992 Jul, 42-44 ( Pt B), 1236 - 42
Scanning force microscopy studies of the S-layers from Bacillus coagulans E38-66, Bacillus sphaericus CCM2177 and of an antibody binding process; Ohnesorge F et al.; In many prokaryotic cells (eubacteria and archaebacteria) the outermost cell envelope component is composed of a regularly structured protein surface layer (S-layer) . The two-dimensional S-layer from Bacillus coagulans E38-66 and Bacillus sphaericus CCM2177 has been investigated by SFM at molecular resolution under physiological conditions (i.e., in buffer solution) . We find the E38-66 S-layer lattice to be oblique with lattice parameters of a = 9-10 nm, b = 7-8 nm and gamma = 80 degrees -90 degrees (E38-66) . The CCM2177 lattice is square with a = 12-14 nm, in good agreement with TEM data . We have used the unique possibility of the SFM to study the kinematics of biological processes and have performed experiments on the adhesion of polyclonal antibodies to the recrystallized E38-66 protein layer on a time scale of about two to ten seconds per image frame . This represents a first step in directly visualizing molecular recognition reactions.

Appl Environ Microbiol, 1992 Jul, 58(7), 2158 - 63
Dual staining of natural bacterioplankton with 4',6-diamidino-2-phenylindole and fluorescent oligonucleotide probes targeting kingdom-level 16S rRNA sequences; Hicks RE et al.; A method for quantifying eubacterial cell densities in dilute communities of small bacterioplankton is presented . Cells in water samples were stained with 4',6-diamidino-2-phenylindole (DAPI), transferred to gelatin-coated slides, and hybridized with rhodamine-labeled oligonucleotide probes specific for kingdom-level 16S rRNA sequences . Between 48 and 69% of the cells captured on membrane filters were transferred to gelatin-coated slides . The number of DAPI-stained cells that were visualized with eubacterial probes varied from 35 to 67% . Only 2 to 4% of these cells also fluoresced following hybridization with a probe designed to target a eukaryotic 16S rRNA sequence . Between 0.1 and 6% of the bacterioplankton in these samples were autofluorescent and may have been mistaken as cells that hybridized with fluorescent oligonucleotide probes . Dual staining allows precise estimates of the efficiency of transfers of cells to gelatin films and can be used to measure the percentage of the total bacterioplankton that also hybridize with fluorescent oligonucleotide probes, indicating specific phylogenetic groups.

Biosci Biotechnol Biochem, 1992 Jul, 56(7), 1113 - 7
Multiple rpoD-related genes of cyanobacteria; Tanaka K et al.; Genomes of many eubacterial strains have been shown to encode for multiple rpoD-related genes . In this report, we describe the identification of the multiple rpoD-related genes of cyanobacterial strains . DNAs of three cyanobacterial strains, Anabaena sp . PCC7120, Synechococcus sp . PCC7942, and Synechocystis sp . PCC6803, were examined by Southern hybridization, using a synthetic probe designed for detecting rpoD or rpoD-related genes . Four or five hybridization signals were found in each DNA . Four DNA regions of Synechococcus sp . PCC7942 corresponding to the hybridization signals were cloned and partially sequenced . The sequence data indicate the presence of genes, named rpoD1, rpoD2, rpoD3, and rpoD4, whose products are highly similar to the basic structure of the principal sigma factors of eubacterial strains . The rpoD1 gene showed the greatest similarity to the sigA gene of Anabaena sp . PCC7120.

Biochem Biophys Res Commun, 1992 Jun 15, 185(2), 539 - 47
The primary structure of rat ribosomal protein L8; Chan YL et al.; The amino acid sequence of the rat 60S ribosomal subunit protein L8 was deduced from the sequence of nucleotides in a recombinant cDNA . Ribosomal protein L8 has 257 amino acids and has a molecular weight of 28,007 . Hybridization of the cDNA to digests of nuclear DNA suggests that there are 4 or 5 copies of the L8 gene . The mRNA for the protein is about 950 nucleotides in length . Rat L8 is homologous to ribosomal proteins from other eukaryotes and to proteins from eubacterial, archaebacterial, and chloroplast ribosomes.

Proc Natl Acad Sci U S A, 1992 Jun 15, 89(12), 5655 - 9
The patA gene product, which contains a region similar to CheY of Escherichia coli, controls heterocyst pattern formation in the cyanobacterium Anabaena 7120; Liang J et al.; In Anabaena 7120, heterocysts (cells specialized for nitrogen fixation) develop at the ends of filaments and at intervals within each filament . We have isolated a mutant Anabaena strain that develops heterocysts mostly at the ends of filaments . This mutant, PAT-1, grows poorly under nitrogen-fixing conditions . The wild-type gene that complements the mutation in PAT-1, called patA, was cloned and sequenced . The predicted PatA protein contains 379 amino acids distributed among three "domains" based on predictions of hydropathy and flexibility . The carboxyl-terminal domain is very similar to that of CheY and other response regulators in two-component regulatory systems in eubacteria . The patA mutation suppresses the multiheterocyst phenotype produced by extra copies of the wild-type hetR gene described previously, suggesting that PatA and HetR are components of the same environment-sensing regulatory circuit in Anabaena.

FEMS Microbiol Lett, 1992 Jun 15, 72(3), 227 - 33
Rapid identification of bacteria of the Comamonadaceae with amplified ribosomal DNA-restriction analysis (ARDRA); Vaneechoutte M et al.; Ribosomal rRNA gene fragments (rDNA) encompassing the 16S rDNA, the 16S-23S rDNA spacer region and part of the 23S rDNA of 95 strains belonging to 13 well-described taxa of the eubacterial family Comamonadaceae (beta subclass of the Proteobacteria or rRNA superfamily III) were enzymatically amplified using conserved primers . The fragments of approximately 2400 base pairs were subjected to restriction analysis . Restriction fragment length patterns obtained with HinfI enabled us to distinguish 9 of the 13 taxa studied . Restriction with CfoI was necessary to differentiate Acidovorax delafieldii from A . temperans and Hydrogenophaga flava from H . pseudoflava . The results indicate that amplified rDNA restriction analysis is a simple and reliable tool for the identification of bacterial species.

Vet Microbiol, 1992 Jun 1, 31(2-3), 287 - 95
Chromosomal DNA probes for the identification of asaccharolytic anaerobic pigmented bacterial rods from the oral cavity of cats; Love DN et al.; A dot-blot hybridisation assay using isolated high molecular weight DNA as whole chromosomal probes of the cat pigmented asaccharolytic Bacteroides/Porphyromonas species was used against both purified high molecular weight DNA and DNA released on membranes from whole cells for the identification of B . salivosus and for its differentiation from the other anaerobic species isolated from normal and diseased mouths of cats and horses . 32P-labelled probes were compared with digoxigenin (DIG)-labelled probes (Boehringer-Mannheim) . The whole chromosomal probes were specific--differentiating B . salivosus from a variety of species (including members of the genera Bacteroides, Fusobacterium, Eubacterium, and Prevotella) found in normal and abnormal mouths of cats and horses . Likewise, asaccharolytic black pigmented Group 2 strains were distinguishable from all strains tested . However, cat strains of P . gingivalis which show 68-76% DNA-DNA homology with human strain P . gingivalis ATCC 33277T, were not distinguishable from each other using either 32P-labelled or DIG-labelled probes . The minimum amount of pure Bacteroides DNA which could be detected by the 32P-labelled probe was 100-300 pg, while the amount of pure DNA detected by the DIG system was 1-3 mg after room temperature colour development for 1 h and 100-300 pg after 6 h colour development.

Eur J Biochem, 1992 Jun 1, 206(2), 373 - 80
YmL9, a nucleus-encoded mitochondrial ribosomal protein of yeast, is homologous to L3 ribosomal proteins from all natural kingdoms and photosynthetic organelles; Graack HR et al.; The nuclear gene for mitochondrial ribosomal protein YmL9 (MRP-L9) of yeast has been cloned and sequenced . The deduced amino acid sequence characterizes YmL9 as a basic (net charge + 30) protein of 27.5 kDa with a putative signal peptide for mitochondrial import of 19 amino acid residues . The intact MRP-L9 gene is essential for mitochondrial function and is located on chromosome XV or VII . YmL9 shows significant sequence similarities to Escherichia coli ribosomal protein L3 and related proteins from various organisms of all three natural kingdoms as well as photosynthetic organelles (cyanelles) . The observed structural conservation is located mostly in the C-terminal half and is independent of the intracellular location of the corresponding genes {Graack, H.-R., Grohmann, L . & Kitakawa, M . (1990) Biol . Chem . Hoppe Seyler 371, 787-788} . YmL9 shows the highest degree of sequence similarity to its eubacterial and cyanelle homologues and is less related to the archaebacterial or eukaryotic cytoplasmic ribosomal proteins . Due to their high sequence similarity to the YmL9 protein two mammalian cytoplasmic ribosomal proteins {MRL3 human and rat; Ou, J.-H., Yen, T . S . B., Wang, Y.-F., Kam, W . K . & Rutter, W . J . (1987) Nucleic Acids Res . 15, 8919-8934} are postulated to be true nucleus-encoded mitochondrial ribosomal proteins.

Proc Natl Acad Sci U S A, 1992 Jun 1, 89(11), 5123 - 7
Restoration of motility to an Escherichia coli fliA flagellar mutant by a Bacillus subtilis sigma factor; Chen YF et al.; The activation of additional promoter sites by production of an alternative sigma subunit for RNA polymerase is a common strategy for the coordinate regulation of gene expression . Many alternative sigma factors control genes for specialized, and often narrowly distributed, functions . For example, most of the alternative sigma factors in Bacillus subtilis control genes necessary for endospore formation . In contrast, the B . subtilis sigma D protein controls the expression of genes important for flagellar-based motility and chemotaxis, a form of locomotion very broadly distributed in the eubacteria . A homologous sigma factor, sigma F, controls a similar group of motility genes in the enteric bacteria . The conservation of both promoter specificity and genetic function in these two regulons allowed us to test the ability of a B . subtilis sigma factor to function within an Escherichia coli host . We demonstrate that expression of the B . subtilis sigD gene restores motility to an E . coli strain mutant in the fliA locus encoding the sigma F factor . This result suggests that the B . subtilis sigma D protein can bind to the E . coli core RNA polymerase to direct transcription initiation from at least four of the late operon promoters, thereby leading to the synthesis of flagellin, motor, and hook-associated proteins . Conversely, expression of sigma D protein in a normally chemotactic strain of E . coli (fliA+) leads to a hyperflagellated, nonchemotactic phenotype.

J Mol Evol, 1992 Jun, 34(6), 471 - 7
Statistical evidence for remnants of the primordial code in the acceptor stem of prokaryotic transfer RNA; Moller W et al.; The specificity of interaction of amino acids with triplets in the acceptor helix stem of tRNA was investigated by means of a statistical analysis of 1400 tRNA sequences . The imprint of a prototypic genetic code at position 3-5 of the acceptor helix was detected, but only for those major amino acids, glycine, alanine, aspartic acid, and valine, that are formed by spark discharges of simple gases in the laboratory . Although remnants of the code at position 3-5 are typical for tRNAs of archaebacteria, eubacteria, and chloroplasts, eukaryotes do not seem to contain this code, and mitochondria take up an intermediary position . A duplication mechanism for the transposition of the original 3-5 code toward its present position in the anticodon stem of tRNA is proposed . From this viewpoint, the mode of evolution of mRNA and functional ribosomes becomes more understandable.

Trends Biochem Sci, 1992 Jun, 17(6), 215 - 8
Glutamyl-transfer RNA: a precursor of heme and chlorophyll biosynthesis; Jahn D et al.; In green plants, archaebacteria and many eubacteria, the porphyrin ring that is common to both chlorophyll and heme is synthesized from 5-aminolevulinic acid (ALA) via an interesting pathway . This two-step process involves the unusual enzymes glutamyl-tRNA reductase and glutamate-1-semialdehyde 2,1-aminomutase . Interest in this pathway has increased since it was discovered that a tRNA cofactor was required for the formation of ALA . This tRNA(Glu) is common to the biosyntheses of both porphyrins and proteins.

Oral Microbiol Immunol, 1992 Jun, 7(3), 182 - 6
Immunochemical and structural characterization of the antigenic polysaccharide from Eubacterium saburreum T18; Nakazawa F et al.; An antigenic surface polysaccharide produced by Eubacterium saburreum strain T18, isolated from human dental plaque, was purified from formamide extract of whole cells . Methylation analysis, Smith degradation, optical rotation data and nuclear magnetic resonance spectra demonstrated that the purified antigen was a homopolysaccharide composed of D-glycero-D-galacto-heptose (Hep.) residues . The structure of the repeating unit in the polysaccharide was: -{----6)-{alpha-Hep.furanosyl-(1----4)}-beta-Hep.pyranosyl- (1----6)-{alpha-Hep.furanosyl-(1----2), alpha-Hep.furanosyl-(1----4)}-beta- Hep.pyranosyl-(1-)4----6)-beta-Hep.pyranosyl-(1---- . No heptose residues were acetylated . Immunodiffusion reactions in agar gel suggested that the immunodeterminant of the antigenic polysaccharide was D-glycero-D-galacto-heptofuranosyl residues as branched nonreducing terminals.

Mol Microbiol, 1992 Jun, 6(12), 1673 - 80
PCR-amplified length polymorphisms in tRNA intergenic spacers for categorizing staphylococci; Welsh J et al.; The intergenic spacers between some adjacent tRNA genes were shown to be polymorphic in length when closely related Staphylococcus species were compared . A simple procedure was developed to detect and sequence these tRNA intergenic length polymorphisms (tRNA-ILPs) . A comparison of homologous tRNA gene sequences flanking these ILPs in three Staphylococcus species was used to derive primers for high-stringency amplification of the ILPs by the polymerase chain reaction (PCR) . The detection of tRNA-ILPs by PCR allowed the classification of virtually all strains from the five species of Staphylococcus that were examined . The procedure used to identify, sequence and derive primers for PCR detection of tRNA-ILPs in Staphylococcus should be applicable to many other genera of eubacteria . These primers could be used on uncultured material such as clinical samples.

J Bacteriol, 1992 Jun, 174(12), 3993 - 9
Cloning, sequencing, mapping, and transcriptional analysis of the groESL operon from Bacillus subtilis; Schmidt A et al.; Using a gene probe of the Escherichia coli groEL gene, a 1.8-kb HindIII fragment of chromosomal DNA of Bacillus subtilis was cloned . Upstream sequences were isolated as a 3-kb PstI fragment . Sequencing of 2,525 bp revealed two open reading frames in the order groES groEL . Alignment of the GroES and GroEL proteins with those of eight other eubacteria revealed 50 to 65% and 72 to 84% sequence similarity, respectively . Primer extension studies revealed one potential transcription start site preceding the groESL operon (S) which was activated upon temperature upshift . Northern (RNA) analysis led to the detection of two mRNA species of 2.2 and 1.5 kb . RNA dot blot experiments revealed an at least 10-fold increase in the amount of specific mRNA from 0 to 5 min postinduction, remaining at this high level for 10 min and then decreasing . A 9-bp inverted repeat within the 5' leader region of the mRNA might be involved in regulation of the heat shock response . By using PBS1 transduction, the groESL operon was mapped at about 342 degrees.

Biochem Biophys Res Commun, 1992 May 29, 185(1), 356 - 62
The primary structure of rat ribosomal protein L11; Chan YL et al.; The amino acid sequence of the rat 60S ribosomal subunit protein L11 was deduced from the sequence of nucleotides in a recombinant cDNA . Ribosomal protein L11 has 178 amino acids and a molecular weight of 20,239 . Hybridization of the cDNA to digests of nuclear DNA suggests that there are 6-8 copies of the L11 gene . The mRNA for the protein is about 800 nucleotides in length . Rat L11 is homologous to ribosomal proteins from other eukaryotes and is related to the L5 family of proteins from eubacterial and archaebacterial ribosomes.

Biochim Biophys Acta, 1992 May 20, 1100(2), 109 - 24
Evolution of organellar proton-ATPases; Nelson N; Proton ATPases function in biological energy conversion in every known living cell . Their ubiquity and antiquity make them a prime source for evolutionary studies . There are two related families of H(+)-ATPases; while the family of F-ATPases function in eubacteria chloroplasts and mitochondria, the family of V-ATPases are present in archaebacteria and the vacuolar system of eukaryotic cells . Sequence analysis of several subunits of V- and F-ATPases revealed several of the important steps in their evolution . Moreover, these studies shed light on the evolution of the various organelles of eukaryotes and suggested some events in the evolution of the three kingdoms of eubacteria, archaebacteria and eukaryotes.

J Biol Chem, 1992 May 5, 267(13), 9170 - 5
A thiocyanate hydrolase of Thiobacillus thioparus . A novel enzyme catalyzing the formation of carbonyl sulfide from thiocyanate; Katayama Y et al.; A thiocyanate hydrolase that catalyzes the first step in thiocyanate degradation was purified to homogeneity from Thiobacillus thioparus, an obligate chemolithotrophic eubacterium metabolizing thiocyanate to sulfate as an energy source . The thiocyanate hydrolase was purified 52-fold by steps involving ammonium sulfate precipitation, DEAE-Sephacel column chromatography, and hydroxylapatite column chromatography . The enzyme hydrolyzed 1 mol of thiocyanate to form 1 mol of carbonyl sulfide and 1 mol of ammonia as follows: SCN- + 2H2O----COS + NH3 + OH- . This is the first report describing the hydrolysis of thiocyanate to carbonyl sulfide by an enzyme . The enzyme had a molecular mass of 126 kDa and was composed of three different subunits: alpha (19 kDa), beta (23 kDa), and gamma (32 kDa) . The enzyme exhibited optimal activities at pH 7.5-8.0 and at temperatures ranging from 30 to 40 degrees C . The Km value for thiocyanate was approximately 11 mM . Immunoblot analysis with polyclonal antibodies against the purified enzyme suggested that it was induced in T . thioparus cells when the cells were grown with thiocyanate.

Curr Genet, 1992 May, 21(6), 485 - 97
Chloroplast RNA polymerase genes of Chlamydomonas reinhardtii exhibit an unusual structure and arrangement; Fong SE et al.; Nucleotide sequence analysis of a 17043 base-pair (bp) region of the Chlamydomonas reinhardtii plastome indicates the presence of three open reading frames (ORFs) similar to RNA polymerase subunit genes . Two, termed rpoB1 and rpoB2, are homologous to the 5'- and 3'-halves of the Escherichia coli beta subunit gene, respectively . A third, termed rpoC2, is similar to the 3'-half of the bacterial beta' subunit gene . These genes exhibit several unusual features: (1) all three represent chimeric structures in which RNA polymerase gene sequences are juxtaposed in-frame with long sequences of unknown identity; (2) unlike their counterparts in plants and eubacteria, rpoB1 and rpoB2 are separated from rpoC2 by a long (7 kilobase-pair, kbp) region containing genes unrelated to RNA polymerase; (3) DNA homologous to the 5' half of rpoC (termed rpoC1 in other species) is not present at the 5' end of rpoC2 and could not be detected in C . reinhardtii chloroplast DNA . RNA expression could not be detected for any of the RNA polymerase genes, suggesting that they are pseudogenes or genes expressed at stages of the C . reinhardtii life-cycle not investigated . The three genes are flanked by GC-rich repeat elements . We suggest that repeat DNA-mediated chloroplast recombination events may have contributed to their unusual arrangement.

J Bacteriol, 1992 May, 174(10), 3416 - 21
Phylogenetic relationships of chemoautotrophic bacterial symbionts of Solemya velum say (Mollusca: Bivalvia) determined by 16S rRNA gene sequence analysis; Eisen JA et al.; The protobranch bivalve Solemya velum Say (Mollusca: Bivalvia) houses chemoautotrophic symbionts intracellularly within its gills . These symbionts were characterized through sequencing of polymerase chain reaction-amplified 16S rRNA coding regions and hybridization of an Escherichia coli gene probe to S . velum genomic DNA restriction fragments . The symbionts appeared to have only one copy of the 16S rRNA gene . The lack of variability in the 16S sequence and hybridization patterns within and between individual S . velum organisms suggested that one species of symbiont is dominant within and specific for this host species . Phylogenetic analysis of the 16S sequences of the symbionts indicates that they lie within the chemoautotrophic cluster of the gamma subdivision of the eubacterial group Proteobacteria.

J Bacteriol, 1992 May, 174(10), 3220 - 6
Conserved gene arrangement in the origin region of the Streptomyces coelicolor chromosome; Calcutt MJ et al.; A 23-kb fragment of the Streptomyces coelicolor chromosome spanning the dnaA region has been isolated as a cosmid clone . Nucleotide sequence analysis of a 5-kb portion shows that the genes for the RNase P protein (rnpA), ribosomal protein L34 (rpmH), the replication initiator protein (dnaA), and the beta subunit of DNA polymerase III (dnaN) are present in the highly conserved gene arrangement found in all eubacterial genomes studied so far . The dnaA-dnaN intergenic region is approximately 1 kb and contains a cluster of at least 12 DnaA boxes with a consensus sequence of TTGTCCACA matching the consensus DnaA box in the phylogenetically related Micrococcus luteus . Two DnaA boxes precede the dnaA sequence . We propose that the chromosomal origin (oriC) of S . coelicolor lies between dnaA and dnaN . In related work, J . Zakrzewska-Czerwinska and H . Schrempf (J . Bacteriol . 174:2688-2693, 1992) have identified the homologous sequence from the closely-related Streptomyces lividans as capable of self-replication.

Lab Invest, 1992 May, 66(5), 618 - 23
Species-specific assessment of Mycobacterium leprae in skin biopsies by in situ hybridization and polymerase chain reaction; Arnoldi J et al.; Conventional histopathologic diagnosis of mycobacterial infections are limited to the determination of "acid-fast bacilli" . A species-specific diagnosis is thus far impossible . In addition, routine microbiologic assessments of mycobacteria suffer from the major drawback that a species-specific diagnosis is extremely time-consuming and in several cases even impossible . As Mycobacterium leprae cannot be cultured in vitro, we tried to specifically target this obligate intracellular parasite by in situ hybridization and polymerase chain reaction (PCR) techniques . For this purpose we used a 22 mer oligonucleotide probe recognizing a species-specific sequence of the 16S rRNA of Mycobacterium leprae . Using an immunoenzymatic detection method for in situ hybridization we were able to specifically assess Mycobacterium leprae (a) in long-term cultured macrophages in vitro infected with different mycobacteria species and (b) in frozen sections of skin biopsies obtained from patients suffering from lepromatous leprosy . These results could be confirmed and extended by PCR experiments in which we used conserved oligonucleotide primers for 16S rRNA to amplify bacterial DNA isolated from different eubacterial species and from fresh-frozen as well as from formalin-fixed, paraffin-embedded and routinely processed mycobacteria-infected tissues . Upon Southern blot analysis, the Mycobacterium leprae-specific oligonucleotide probe exclusively hybridized with PCR products obtained from Mycobacterium leprae-containing samples (including paraffin sections), but not with PCR products obtained from samples containing other mycobacterial species . As species-specific oligonucleotide probes targeted at rRNA are described for a variety of mycobacterial species, these methods may be generally applied for a rapid species-specific assessment of mycobacteria in histologic material.

Mol Gen Genet, 1992 May, 233(1-2), 293 - 301
A general system to integrate lacZ fusions into the chromosomes of gram-negative eubacteria: regulation of the Pm promoter of the TOL plasmid studied with all controlling elements in monocopy; Kessler B et al.; A new procedure is described to recombine plasmid-borne lacZ fusions into the chromosome of gram-negative eubacteria in order to study promoter activity in monocopy . The procedure is based upon the insertion into the chromosome of a target bacterium of a recombinant transposon that carries DNA sequence homology to the regions flanking lacZ fusions present in multicopy promotor-probe vectors, which can be mobilized via RP4-mediated transfer but are unable to replicate in non-enteric bacteria . Double recombination between the promoter-probe vectors and the chromosomal homology region of the transposon is genetically selected by reconstruction and expression of wild-type sequences from truncated lacZ and aadA (streptomycin/spectinomycin) resistance genes in the homology fragment and from an amber mutation carrying lacZ and aadA genes present in the plasmid vectors . The structure of desired clones is confirmed by screening for loss of the transposon-encoded kanamycin resistance marker . We have used this procedure to assemble in monocopy in Pseudomonas putida the regulatory elements controlling expression of the XylS-activated Pm promoter of the TOL catabolic plasmid pWWO . We show here that the Pm promoter undergoes a XylS-independent, strictly growth-phase-controlled activation by benzoate but not meta-toluate . In the presence of XylS, however, activation by both effectors involves a combination of growth phase-dependent and -independent controls.

Biochim Biophys Acta, 1992 Apr 17, 1120(3), 267 - 72
Characterization of an extremely thermostable glutamate dehydrogenase: a key enzyme in the primary metabolism of the hyperthermophilic archaebacterium, Pyrococcus furiosus; Robb FT et al.; Glutamate dehydrogenase (L-glutamate:NAD(P)+ oxidoreductase, deaminating, EC 1.4.1.3) from the hyperthermophilic Archeon Pyrococcus furiosus was purified to homogeneity by chromatography on anion-exchange, molecular-exclusion and hydrophobic-interaction media . The purified native enzyme had an M(r) of 270,000 +/- 15,000 and was shown to be a hexamer with identical subunits of M(r) 46,000 . The enzyme was exceptionally thermostable, having a half-life of 3.5 to more than 10 h at 100 degrees C, depending on the concentration of enzyme . The Km of the enzyme for ammonia was high (9.5 mM), indicating that the enzyme is probably active in the deaminating, catabolic direction . The coenzyme utilization of the enzyme resembled the equivalent enzymes from eukaryotes rather than eubacteria, since both NADH and NADPH were recognized with high affinity . The enzyme displayed a preference for NADP+ over NAD+ that was more pronounced at low assay temperatures (50-70 degrees C) compared with the optimal temperature for enzyme activity, 95 degrees C.

Biochim Biophys Acta, 1992 Apr 17, 1120(3), 289 - 96
An extremely stable inorganic pyrophosphatase purified from the cytosol of a thermoacidophilic archaebacterium, Sulfolobus acidocaldarius strain 7; Wakagi T et al.; A highly active inorganic pyrophosphatase was purified to electrophoretical homogeneity from the cytosol of Sulfolobus acidocaldarius strain 7, an extremely thermoacidophilic archaebacterium . The enzyme has an apparent molecular mass of 80 kDa as estimated by gel permeation chromatography, and showed a 21-kDa polypeptide on SDS-PAGE, suggesting that the archaebacterial enzyme is similar to most of the eubacterial pyrophosphatases rather than eukaryotic ones . The pI = 5.1 . The enzyme showed relatively high content of Pro and low content of Ser plus Thr . The optimal pH was 6.5 (at 56 degrees C) . From the Arrhenius plot an activation energy of 11.2 kcal/mol was obtained between 37-95 degrees C . The specific activity was 617 mumol Pi release min-1 mg-1 at 56 degrees C . The S . acidocaldarius pyrophosphatase was extremely stable . Complete activity remained after incubation at 100 degrees C for 10 min . No dissociation into subunit or unfolding of polypeptide chain occurred in the presence of 8 M urea . Experiments using guanidine-HCl suggested that the transition between a native tetrameric state and an unfolded state is completely reversible, and essentially independent of any additional factors such as divalent metal cation or dithiothreitol.

Gene, 1992 Apr 15, 113(2), 175 - 81
Genetic analysis of an aphid endosymbiont DNA fragment homologous to the rnpA-rpmH-dnaA-dnaN-gyrB region of eubacteria; Lai CY et al.; Buchnera aphidicola is a Gram- eubacterium with a DNA G+C content of 28-30 mol% . This organism is an obligate intracellular symbiont of aphids . To determine its similarity to or difference from other eubacteria, a 4.9-kb DNA fragment from B . aphidicola containing the gene homologous to Escherichia coli dnaA (a gene involved in the initiation of chromosome replication) was cloned into E . coli and sequenced . The order of genes on this fragment, 60K-10K-rnpA-rpmH-dnaA-dnaN-gyrB, was similar to that found in other eubacteria . The sole difference was the absence of recF between dnaN and gyrB . The deduced amino acid sequence of these proteins resembled those of E . coli by a 41 to 83% identity . Except for E . coli, in all the eubacteria so far examined, dnaA is preceded by multiple 9-nucleotide repeats known as a DnaA boxes . No DnaA boxes were detected in the endosymbiont DNA . The possibility that this observation is a consequence of the low G+C content of this DNA fragment (14 mol% G+C) is unlikely since in Mycoplasma capricolum this fragment (19 mol% G+C) has eight DnaA boxes (Fujita et al., 1992) . The presence of the sequence, GATC, recognized by the Dam methyl-transferase system, only within six regions coding for proteins suggests that methylation is not a factor in the regulation of the initiation of endosymbiont chromosome replication.

Eur J Biochem, 1992 Apr 15, 205(2), 853 - 66
13C-NMR study of autotrophic CO2 fixation pathways in the sulfur-reducing Archaebacterium Thermoproteus neutrophilus and in the phototrophic Eubacterium Chloroflexus aurantiacus; Strauss G et al.; The unresolved autotrophic CO2 fixation pathways in the sulfur-reducing Archaebacterium Thermoproteus neutrophilus and in the phototrophic Eubacterium Chloroflexus aurantiacus have been investigated . Autotrophically growing cultures were labelled with {1,4-13C1}succinate, and the 13C pattern in cell constituents was determined by 1H- and 13C-NMR spectroscopy of purified amino acids and other cell constituents . In both organisms succinate contributed to less than 10% of cell carbon, the major part of carbon originated from CO2 . All cell constituents became 13C-labelled, but different patterns were observed in the two organisms . This proves that two different cyclic CO2 fixation pathways are operating in autotrophic carbon assimilation in both of which succinate is an intermediate . The 13C-labelling pattern in T . neutrophilus is consistent with the operation of a reductive citric acid cycle and rules out any other known autotrophic CO2 fixation pathway . Surprisingly, the proffered {1,4-13C1}succinate was partially converted to double-labelled {3,4-13C2}glutamate, but not to double-labelled aspartate . These findings suggest that the conversion of citrate to 2-oxoglutarate is readily reversible under the growth conditions used, and a reversible citrate cleavage reaction is proposed . The 13C-labelling pattern in C . aurantiacus disagrees with any of the established CO2 fixation pathways; it therefore demands a novel autotrophic CO2 fixation cycle in which 3-hydroxypropionate and succinate are likely intermediates . The bacterium excreted substantial amounts of 3-hydroxypropionate (5 mM) and succinate (0.5 mM) at the end of autotrophic growth . Autotrophically grown Chloroflexus cells contained acetyl-CoA carboxylase and propionyl-CoA carboxylase activity . These enzymes are proposed to be the main CO2-fixing enzymes resulting in malonyl-CoA and methylmalonyl-CoA formation; from these carboxylation products 3-hydroxypropionate and succinate, respectively, can be formed.

J Biol Chem, 1992 Apr 15, 267(11), 7539 - 44
Isolation and characterization of a NADP-dependent glutamate dehydrogenase gene from the primitive eucaryote Giardia lamblia; Yee J et al.; Giardia lamblia is believed to be the earliest branching derivative from the eucaryotic lineage . Genomic and cDNA clones encoding the giardia NADP-dependent glutamate dehydrogenase have been isolated and characterized . Southern hydridization using genomic DNA indicates that the gene encoding this activity is unique and single copy . Primer extension, S1 nuclease protection, and genomic and cDNA sequence analysis demonstrate that gene transcripts are initiated within a conserved AT-rich sequence element immediately preceding the ATG translation initiation codon and the short 5' untranslated region is not extended by transsplicing . The open reading frame is 1350 nucleotides in length and encodes a protein of 449 amino acids . The reading frame is not interrupted by introns and the primary transcript is probably not subjected to RNA editing . In the strictly anaerobic metabolism of giardia, NADP-dependent glutamate dehydrogenase activity participates along with alanine aminotransferase, in the cyclic dissipation of reducing equivalents (NADPH) through the conversion of pyruvate to alanine . The deduced amino acid sequence of the giardia protein exhibits substantial homology to numerous fungal and eubacterial NADP-dependent glutamate dehydrogenases . Comparisons of alignment gap positions and amino acid identities indicate that the giardia sequence is at least as similar or more similar to the eubacterial sequence than it is to the fungal sequence . This supports the hypothesis that giardia diverged very early from the eucaryotic lineage.

Oral Microbiol Immunol, 1992 Apr, 7(2), 100 - 5
Microbiological evaluation of periapical infections in Egypt; Wasfy MO et al.; This study identifies and correlates proportions of bacteria in periapically involved anterior teeth of 85 adult Egyptian patients . Affected sites were free from caries and periodontal disease but had a history of trauma . The mean number of component bacterial species per specimen was 3.1 . Anaerobic bacteria were the dominant flora present in specimen cultures, comprising 73% (190/259) of cultivable bacteria . The most frequently isolated organisms were Eubacterium species (68%), black-pigmented Bacteroides (56%), Streptococcus morbillorum (47%) and non-pigmented Bacteroides (37%) . These organisms also showed the highest proportional values relative to total cultivable bacteria . The mean percentages of total viable counts of these isolates were 19.0%, 14.1%, 18.0% and 15.5%, respectively . Statistical analysis of data revealed a significant negative correlation between S . morbillorum and Bacteroides species.

J Mol Evol, 1992 Apr, 34(4), 351 - 7
Organization and evolution of bacterial and bacteriophage primase-helicase systems; Ilyina TV et al.; Amino acid sequences of primases and associated helicases involved in the DNA replication of eubacteria and bacteriophages T7, T3, T4, P4, and P22 were compared by computer-assisted methods . There are two types of such systems, the first one represented by distinct helicase and primase proteins (e.g., DnaB and DnaG proteins of Escherichia coli), and the second one by single polypeptides comprising both activities (gp4 of bacteriophages T7 and T3, and alpha protein of bacteriophage P4) . Pronounced sequence similarity was revealed between approximately 250 amino acid residue N-terminal domains of stand-alone primases and the primase-helicase proteins of T7(T3) and P4 . All these domains contain, close to their N-termini, a conserved Zn-finger pattern that may be implicated in template DNA recognition by the primases . In addition, they encompass five other conserved motifs some of which may be involved in substrate (NTP) binding . Significant similarity was also observed between the primase-associated helicases (DnaB, gp12 and P22 and gp41 of T4) and the C-terminal domain of T7(T3) gp4 . On the other hand the C-terminal domain of P-alpha of P4 is related to another group of DNA and RNA helicases . Tentative phylogenetic trees generated for the primases and the associated helicases showed no grouping of the phage proteins, with the exception of the primase domains of bacteriophages T4 and P4 . This may indicate a common origin for one-component primase-helicase systems . Two scenarios for the evolution of primase-helicase systems are discussed.(ABSTRACT TRUNCATED AT 250 WORDS)

J Bacteriol, 1992 Apr, 174(8), 2592 - 8
Activities of two dissimilar thioredoxins from the cyanobacterium Anabaena sp . strain PCC 7120; Gleason FK; Thioredoxin is a small redox protein that functions as a reducing agent and modulator of enzyme activity . A gene for an unusual thioredoxin was previously isolated from the cyanobacterium Anabaena sp . strain PCC 7120 and cloned and expressed in Escherichia coli . However, the protein could not be detected in Anabaena cells (J . Alam, S . Curtis, F . K . Gleason, M . Gerami-Nejad, and J . A . Fuchs, J . Bacteriol . 171:162-171, 1989) . Polyclonal antibodies to the atypical thioredoxin were prepared, and the protein was detected by Western immunoblotting . It occurs at very low levels in extracts of Anabaena sp . and other cyanobacteria . No antibody cross-reaction was observed in extracts of eukaryotic algae, plants, or eubacteria . The anti-Anabaena thioredoxin antibodies did react with another unusual thioredoxin-glutaredoxin produced by bacteriophage T4 . Like the T4 protein and other glutaredoxins, the unusual cyanobacterial thioredoxin can be reduced by glutathione . The Anabaena protein can also activate enzymes of carbon metabolism and has some functional similarity to spinach chloroplast thioredoxin f . However, it shows only 23% amino acid sequence identity to the spinach chloroplast protein and appears to be distantly related to other thioredoxins . The data indicate that cyanobacteria, like plant chloroplasts, have two dissimilar thioredoxins . One is related to the more common protein found in other prokaryotes, and the other is an unusual thioredoxin that can be reduced by glutathione and may function in glucose catabolism.

Obstet Gynecol, 1992 Apr, 79(4), 534 - 8
Eubacterium nodatum mimics Actinomyces in intrauterine device-associated infections and other settings within the female genital tract; Hill GB; Eubacterium nodatum is an obligately anaerobic, gram-positive, branching rod that markedly resembles Actinomyces, particularly Actinomyces israelii, in its cellular and colonial characteristics . Its isolation from the female genital tract was examined for a study period in which use of intrauterine contraceptive devices (IUDs) was common, and additional characteristics of the organism were investigated . Fifteen genital isolates of E nodatum were all associated with the presence of a foreign body, usually an IUD (12 patients) . Six of these 12 patients had presented with clinically severe pelvic inflammatory disease . The remaining six had signs and symptoms related to IUD use and/or had a report of probable Actinomyces (five patients) by a Papanicolaou smear, demonstrating that E nodatum can be mistaken for Actinomyces in a Papanicolaou-stained smear . The three other patients had different types of foreign bodies . The frequency of isolation from cultures associated with IUD use during the study period was five (6.4%) of 78 for Actinomyces versus the 12 (15.4%) of 78 for E nodatum . In vitro-prepared E nodatum was not demonstrated to cross-react with A israelii or A naeslundii antisera . Both E nodatum and A israelii were shown to adhere in vitro to an inanimate object, indicating their propensities to colonize a foreign body . The present data, with the previous reports of isolation of E nodatum from cases of lumpy jaw and severe periodontitis, suggest that it is an opportunistic pathogen very much like A israelii.

J Bacteriol, 1992 Apr, 174(7), 2065 - 71
The bile acid-inducible baiB gene from Eubacterium sp . strain VPI 12708 encodes a bile acid-coenzyme A ligase; Mallonee DH et al.; The baiB gene from Eubacterium sp . strain VPI 12708 was previously cloned, sequenced, and shown to be part of a large bile acid-inducible operon encoding polypeptides believed to be involved in bile acid 7 alpha-dehydroxylation . In the present study, the baiB gene was subcloned and expressed in Escherichia coli and shown to encode a bile acid-coenzyme A (CoA) ligase . This ligase required a C-24 bile acid with a free carboxyl group, ATP, Mg2+, and CoA for synthesis of the final bile acid-CoA conjugate . Product analysis by reverse-phase high-performance liquid chromatography revealed final reaction products that comigrated with cholyl-CoA and AMP . A putative bile acid-AMP intermediate was detected when CoA was omitted from the reaction mixture . The bile acid-CoA ligase has amino acid sequence similarity to several other polypeptides involved in the ATP-dependent linking of AMP or CoA to cyclic carboxylated compounds . The bile acid-CoA ligation is believed to be the initial step in the bile acid 7 alpha-dehydroxylation pathway in Eubacterium sp . strain VPI 12708.

J Mol Evol, 1992 Apr, 34(4), 292 - 303
ATP synthase subunit c/III/9 gene sequences as a tool for interkingdom and metaphytes molecular phylogenies; Recipon H et al.; The 38 sequences of the ATPase c/III/9 gene determined in bacteria, fungi, mammals, and higher plants have been used to construct phylogenetic trees by distance matrix and parsimony methods (checked by bootstrapping); alignments have been performed on the deduced amino-acid sequences and then transferred back to the nucleotide sequences . Three lineages stand out: (1) eubacteria (except cyanobacteria and alpha purple bacteria), (2) chloroplasts, together with cyanobacteria, and (3) mitochondria together with nuclei and alpha purple bacteria . The clear monophyly of the mitochondrial/nuclear lineage, taken all together, strongly suggests that the nuclear copies of the gene now residing in the eukaryotic nucleus originate from a mitochondrial transfer . Within this lineage, metaphytes emerge late and as a cohesive group, after fungi (as a dispersed group) and metazoa, yielding an order that markedly differs from that obtained through typical RNA nuclear molecules . The possible biphyletic origin of mitochondria based on mitochondrial rRNA sequences is not evidenced by these sequences . Internal branches within both the chloroplastic and the mitochondrial lineages are consistent with botanical evolutionary schemes based on morphological characters . In spite of its relatively small size, the ATPase c/III/9 gene therefore displays remarkable properties as a phylogenetic index and adds a new tool for molecular evolutionary reconstructions, especially within the metaphytes.

J Bioenerg Biomembr, 1992 Apr, 24(2), 193 - 200
Sensory rhodopsin I: receptor activation and signal relay; Spudich JL et al.; Recent progress is summarized on the mechanism of phototransduction by sensory rhodopsin I (SR-I), a phototaxis receptor in Halobacterium halobium . Two aspects are emphasized: (i) The coupling of retinal isomerization to protein conformational changes . Retinal analogs have been used to probe chromophore-apoprotein interactions during the receptor activation process . One of the most important results is the finding of a steric trigger deriving from the interaction of residues on the protein with a methyl group near the isomerizing bond of the retinal (at carbon 13) . Recent work on molecular genetic methods to further probe structure/function includes the synthesis and expression of an SR-I apoprotein gene designed for residue replacements by cassette mutagenesis, and transformation of an H . halobium mutant lacking all retinylidene proteins known in this species to SR-I+ and bacteriorhodopsin (BR)+ . (ii) The relay of the SR-I signal to a post-receptor component . A carboxylmethylated protein ("MPP-I") associated with SR-I and found in the H . halobium membrane exhibits homology with the signaling domain of eubacterial chemotaxis transducers (e.g., Escherichia coli Tar, Tsr, and Trg proteins), suggesting a model based on SR-I----MPP-I signal relay.

Chem Pharm Bull (Tokyo), 1992 Apr, 40(4), 1056 - 7
Sulfation of phenolic antibiotics by sulfotransferase obtained from a human intestinal bacterium; Kim DH et al.; Novel sulfotransferase which was isolated from Eubacterium A-44, a human intestinal bacterium, sulfated phenolic antibiotics, such as amoxicillin, ceaodroxil and cefoperazone . The Km values of sulfotransferase for these antibiotics were 6.9, 4.3 and 22.2 mM, respectively . The Vmax values were 8.3, 3.3 and 1.6 mumol/min/mg protein . The optimal pH of the enzyme was 9.0, a weakly alkaline region . The antibacterial activity of amoxicillin was not altered by enzymic sulfation of the phenolic hydroxyl group.

Biochimie, 1992 Apr, 74(4), 337 - 51
Comparison of dissimilarity patterns of E coli, yeast and mammalian tRNAs; Steinberg SV et al.; A number of experimental approaches have been developed for identification of recognition (identity) sites in tRNAs . Along with them a theoretical methodology has been proposed by McClain et al that is based on concomitant analysis of all tRNA sequences from a given species . This approach allows an evaluation of nucleotide combinations present in isoacceptor tRNAs specific for the given amino acid, and not present in equivalent positions in cloverleaf structure in other tRNAs of the same organism . These elements predicted from computer analysis of the databank could be tested experimentally for their participation in forming recognition sites . The correlation between theoretical predictions and experimental data appeared promising . The aim of the present work consisted of introducing further improvements into McClain's procedure by: i), introducing into analysis a variable region in tRNAs which had not been previously considered; to accomplish this, 'normalization' of variable nucleotides was suggested, based on primary and tertiary structures of tRNAs; ii), developing a new procedure for comparison of patterns for synonymous and non-synonymous tRNAs from different organisms; iii), analysis of 3- and 4-positional contacts between tRNAs and enzymes in addition to a formerly used 2-positional model . A systematic application of McClain's procedure to mammalian, yeast and E coli tRNAs led to the following results: i), imitancy patterns for non-synonymous tRNAs of any amino acid specificity and from any organisms analysed so far overlap by no more than 30%, providing a structural basis for discrimination with high fidelity between cognate and non-cognate tRNAs; ii), the predicted identity sites are non-randomly distributed within tRNA molecules; the dominant role is ascribed to only two regions--anticodon and amino acid stem which are located far apart from one another at extremes of all tRNA molecules; iii), the imitancy patterns for synonymous tRNAs in lower (yeast) and higher (mammalian) eukaryotes are similar but not identical; iv), distribution of predicted identity sites in the cloverleaf structure in prokaryotes and eukaryotes is essentially different: in eubacterial tRNAs the major role in recognition plays anticodon and/or amino acid acceptor stem, whereas in eukaryotic (both unicellular and multicellular) tRNAs the remaining part of the molecules is also involved in recognition; v), the imitancy patterns of synonymous tRNAs from prokaryotes and eukaryotes are dissimilar, this observation leads to the prediction that the tRNA identity sites for the same amino acid in prokaryotes and eukaryotes may differ.

Biochem Biophys Res Commun, 1992 Mar 31, 183(3), 952 - 7
Tandem repeat of the 23S and 5S ribosomal RNA genes in Borrelia burgdorferi, the etiological agent of Lyme disease; Fukunaga M et al.; The DNA fragments containing the rrl and rrf genes were subcloned from a EMBL3 recombinant phage of Borrelia burgdorferi strain B31 into pUC18 and were characterized by restriction analysis and Southern hybridization . A fine restriction map of the fragments was constructed and the organization of the genes was determined . The genomic hybridization using the gene probes from B . burgdorferi showed that there are two sets of rrl/rrf genes in that genome . The results also revealed the important fact that the gene sets are repeated directly by 3.2-kb long . This is the first report of this remarkable feature in the organization of the eubacterial rRNA genes.

Proc R Soc Lond B Biol Sci, 1992 Mar 23, 247(1320), 163 - 8
Molecular identification of Wolbachia, the agent of cytoplasmic incompatibility in Drosophila simulans, and variability in relation with host mitochondrial types; Rousset F et al.; Sequences of a segment of the 16S ribosomal DNA of Wolbachia, a rickettsia-like microorganism responsible for cytoplasmic incompatibility in Drosophila simulans, have been obtained after polymerase chain reaction (PCR) amplification . Their comparison with other eubacterial sequences allows us to assign these endosymbionts to the alpha subdivision of purple bacteria . Four related sequences have been obtained for microorganisms carried by eight isofemale lines representative of the three mitochondrial types of D . simulans . Their phylogeny and level of divergence do not parallel that of the mitochondrial DNA, suggesting that several independent infections occurred . There is no direct relation between bacterial phylogeny and formerly identified incompatibility types.

J Mol Biol, 1992 Mar 20, 224(2), 529 - 36
Chloroplast ATPase genes in the diatom Odontella sinensis reflect cyanobacterial characters in structure and arrangement; Pancic PG et al.; We have cloned and sequenced a 5200 base restriction fragment and an overlapping 3100 base fragment of the large single copy region of the chloroplast genome of the diatom Odontella sinensis, which hybridized to several ATPase gene probes . These fragments contain six closely linked reading frames that were identified as atpI, atpH, atpG, atpF, atpD, and atpA, coding for subunits IV, III, II, I, delta, and alpha, respectively . Remarkably, the genes atpG and atpD, which are nucleus-encoded in chlorophyll a + b plants, are present in the Odontella chloroplast gene cluster . They map at the same positions as in cyanobacteria . The genes atpD and atpF overlap by four base-pairs as in certain photosynthetic and heterotrophic eubacteria . Upstream from the atpA gene cluster an open reading frame coding for 251 amino acid residues was found, which shows sequence similarity to ATP-binding subunits of periplasmic prokaryotic and eukaryotic transport systems . No similar reading frame is present in the land plant chloroplast genomes analysed so far . Sequences and arrangement of the genes are discussed with respect to the peculiar evolution of the chlorophyll a + c-containing chromophytic plastids.

J Biol Chem, 1992 Mar 15, 267(8), 5162 - 70
Structure and function of MRP20 and MRP49, the nuclear genes for two proteins of the 54 S subunit of the yeast mitochondrial ribosome; Fearon K et al.; MRP20 and MRP49 are proteins of the large subunit of the mitochondrial ribosome in Saccharomyces cerevisiae . Their genes were identified through immunological screening of a genomic library in the expression vector lambda gt11 . Nucleotide sequencing revealed that MRP49 is tightly linked to TPK3 and encodes a 16-kDa, basic protein with no significant relatedness to any other known protein . MRP20 specifies a 263-amino-acid polypeptide with sequence similarity to members of the L23 family of ribosomal proteins . The levels of the mRNAs and proteins for both MRP20 and MRP49 were regulated in response to carbon source . In {rho0} strains lacking mitochondrial rRNA, the levels of the two proteins were reduced severalfold, presumably because the unassembled proteins are unstable . Null mutants of MRP20 converted to {rho-} or {rho0}, a characteristic phenotype of mutations in essential genes for mitochondrial translation . Inactivation of MRP49 caused a cold-sensitive respiration-deficient phenotype, indicating that MRP49 is not an essential ribosomal protein . The mrp49 mutants were defective in the assembly of stable 54 S ribosomal subunits at the nonpermissive temperature . With the results presented here, there are now published sequences for 14 yeast mitochondrial ribosomal proteins, only five of which bear discernable relationships to eubacterial ribosomal proteins.

Nucleic Acids Res, 1992 Mar 11, 20(5), 961 - 74
Compilation and analysis of DNA sequences associated with apparent streptomycete promoters; Strohl WR; The DNA sequences associated with 139 apparent streptomycete transcriptional start sites are compiled and compared . Of these, 29 promoters appeared to belong to a group which are similar to those recognized by eubacterial RNA polymerases containing sigma 70-like subunits . The other 110 putative promoter regions contain a wide diversity of sequences; several of these promoters have obvious sequence similarities in the -10 and/or -35 regions . The apparent Shine-Dalgarno regions of 44 streptomycete genes are also examined and compared . These were found to have a wide range of degree of complementarity to the 3' end of streptomycete 16S rRNA . Eleven streptomycete genes are described and compared in which transcription and translation are proposed to be initiated from the same or nearby nucleotide . An updated consensus sequence for the E sigma 70-like promoters is proposed and a potential group of promoter sequences containing guanine-rich -35 regions also is identified.

Plasmid, 1992 Mar, 27(2), 141 - 54
Heterologous gene expression in Bacteroides fragilis; Smith CJ et al.; Bacteroides fragilis and other gastrointestinal tract Bacteroides are unusual gram-negative eubacteria in that genes from other gram-negative eubacteria are not expressed when introduced into these organisms . To analyze gene expression in Bacteroides, expression vector and promoter probe (detection) vector systems were developed . The essential feature of the expression vector was the incorporation of a Bacteroides insertion sequence element, IS4351, which possesses promoter activity directed outward from its ends . Genes inserted into the multiple cloning site downstream from an IS4351 DNA fragment were readily expressed in B . fragilis . The chloramphenicol acetyltransferase (cat) structural gene from Tn9 was tested and conferred chloramphenicol resistance on B . fragilis . Both chloramphenicol resistance and CAT activity were shown to be dependent on the IS4351 promoters . Similar results were obtained with the Escherichia coli beta-glucuronidase gene (uidA) but activity was just 30% of the levels seen with cat . Two tetracycline resistance determinants, tetM from Streptococcus agalactiae and tetC from E . coli, also were examined . tetC did not result in detectable tetracycline resistance but the gram-positive tetM gene conferred high-level resistance to tetracycline and minocycline in Bacteroides hosts . Based on the cat results, promoter probe vectors containing the promoterless cat gene were constructed . These vectors were used to clone random B . fragilis promoters from partial genomic libraries and the recombinants displayed a range of CAT activities and chloramphenicol MICs in B . fragilis hosts . In addition, known E . coli promoters (Ptet, Ptac, Ptrc, Psyn, and P1P2rrnB) were tested for activity in B . fragilis . No chloramphenicol resistance or CAT activity was observed in B . fragilis with these promoters.

EMBO J, 1992 Mar, 11(3), 795 - 804
Fasciation induction by the phytopathogen Rhodococcus fascians depends upon a linear plasmid encoding a cytokinin synthase gene; Crespi M et al.; Rhodococcus fascians is a nocardiform bacteria that induces leafy galls (fasciation) on dicotyledonous and several monocotyledonous plants . The wild-type strain D188 contained a conjugative, 200 kb linear extrachromosomal element, pFiD188 . Linear plasmid-cured strains were avirulent and reintroduction of this linear element restored virulence . Pulsed field electrophoresis indicated that the chromosome might also be a linear molecule of 4 megabases . Three loci involved in phytopathogenicity have been identified by insertion mutagenesis of this Fi plasmid . Inactivation of the fas locus resulted in avirulent strains, whereas insertions in the two other loci affected the degree of virulence, yielding attenuated (att) and hypervirulent (hyp) bacteria . One of the genes within the fas locus encoded an isopentenyltranferase (IPT) with low homology to analogous proteins from Gram-negative phytopathogenic bacteria . IPT activity was detected after expression of this protein in Escherichia coli cells . In R.fascians, ipt expression could only be detected in bacteria induced with extracts from fasciated tissue . R.fascians strains without the linear plasmid but containing this fas locus alone could not provoke any phenotype on plants, indicating additional genes from the linear plasmid were also essential for virulence . These studies, the first genetic analysis of the interaction of a Gram-positive bacterium with plants, suggest that a novel mechanism for plant tumour induction has evolved in R.fascians independently from the other branches of the eubacteria.

Proc Natl Acad Sci U S A, 1992 Mar 1, 89(5), 1885 - 9
Direct repeat sequences are implicated in the regulation of two Streptomyces chitinase promoters that are subject to carbon catabolite control; Delic I et al.; We report the identification and partial characterization of the promoters for two chitinase genes from Streptomyces plicatus . Chitinases are a family of enzymes made by Streptomyces and other soil microbes to digest chitin, an abundant source of carbon and nitrogen in the soil . The promoter regions of two chitinases were defined by using transcriptional fusions to the xylE reporter gene . Transcription was shown to be glucose-sensitive and chitin-dependent . Each promoter contains a putative RNA polymerase binding site with a recognition sequence very similar to that observed in many eubacterial vegetatively expressed genes . In both promoters, a pair of 12-base-pair direct repeat sequences overlap the putative RNA polymerase binding sites . Further analysis of one of the promoters revealed that a single-base change within the direct repeat sequences resulted in glucose-resistant, chitin-independent expression in vivo . In addition, the promoter region that includes the direct repeat sequences was shown to interact with a sequence-specific DNA binding factor in vitro . Similar direct repeat sequences are present in other chitinase genes recently characterized, and we suggest that these repeats may be involved in repression and induction for this entire class of catabolite-controlled genes.

Eur J Biochem, 1992 Mar 1, 204(2), 679 - 83
Biosynthesis of vitamin B12 in anaerobic bacteria . Experiments with Eubacterium limosum on the incorporation of D-{1-13C}erythrose and {13C}formate into the 5,6-dimethylbenzimidazole moiety; Munder M et al.; Experiments on the incorporation of erythrose and formate into the 5,6-dimethylbenzimidazole moiety of vitamin B12 are described . In one experiment, a 1:1 mixture of D-{1-13C}erythrose and D-{1-13C}threose was added to a Eubacterium limosum fermentation . The vitamin B12 formed was methylated at N3 of its 5,6-dimethylbenzimidazole part and degraded to 1,5,6-trimethylbenzimidazole . The 13C-NMR spectrum of this compound exhibited a single prominent signal at 109.5 ppm due to 13C labeling in C7 . This shows that C1 of erythrose or threose was originally incorporated exclusively into C4 of the 5,6-dimethylbenzimidazole moiety of vitamin B12 . In another experiment, sodium {13C}formate was added to a culture of E . limosum . The vitamin B12 isolated was transformed into 1,5,6-trimethylbenzimidazole as before . The 13C-NMR spectrum also showed one prominent signal at 142.8 ppm, evoked by 13C at C2 . These results demonstrate that erythrose is incorporated into the base part of vitamin B12 regiospecifically and that formate is the precursor of the C2.

Microbiol Rev, 1992 Mar, 56(1), 100 - 22
Cyclic AMP in prokaryotes; Botsford JL et al.; Cyclic AMP (cAMP) is found in a variety of prokaryotes including both eubacteria and archaebacteria . cAMP plays a role in regulating gene expression, not only for the classic inducible catabolic operons, but also for other categories . In the enteric coliforms, the effects of cAMP on gene expression are mediated through its interaction with and allosteric modification of a cAMP-binding protein (CRP) . The CRP-cAMP complex subsequently binds specific DNA sequences and either activates or inhibits transcription depending upon the positioning of the complex relative to the promoter . Enteric coliforms have provided a model to explore the mechanisms involved in controlling adenylate cyclase activity, in regulating adenylate cyclase synthesis, and in performing detailed examinations of CRP-cAMP complex-regulated gene expression . This review summarizes recent work focused on elucidating the molecular mechanisms of CRP-cAMP complex-mediated processes . For other bacteria, less detail is known . cAMP has been implicated in regulating antibiotic production, phototrophic growth, and pathogenesis . A role for cAMP has been suggested in nitrogen fixation . Often the only data that support cAMP involvement in these processes includes cAMP measurement, detection of the enzymes involved in cAMP metabolism, or observed effects of high concentrations of the nucleotide on cell growth.

Biochim Biophys Acta, 1992 Feb 28, 1130(1), 113 - 6
The genes for ribosomal protein L15 and the protein equivalent to secY in the archaebacterium Haloarcula (Halobacterium) marismortui; Arndt E; The nucleotide sequences of the L15 gene and the secY gene, which form the last two genes of the S10/spc-operon region of Haloarcula marismortui, have been determined . The HmaL15 protein sequence translated from the DNA is 164 amino acids long, revealing 10 amino acids more at the C-terminus than the published protein sequence . The deduced HmasecY protein sequence has 487 amino acids and shows significant homology to its counterparts in Methanococcus vannielii and Escherichia coli . The gene order of the halobacterial gene cluster is similar to that in the methanogens and in eubacteria.

Eur J Biochem, 1992 Feb 15, 204(1), 231 - 40
Membrane protein acylation . Preference for exogenous myristic acid or endogenous saturated chains in Acholeplasma laidlawii; Nystrom S et al.; Mycoplasmas are small bacteria without a cell wall, often found as surface parasites on eukaryotic cells . Of the more than 200 membrane proteins from Acholeplasma laidlawii resolved by two-dimensional PAGE, 23 were covalently modified with acyl chains . These acyl proteins had lower pI values than average and were all labelled by different exogenously supplied radioactive fatty acids attached by O-ester bonds . The fatty acids were selectively incorporated in the order myristic acid (14:0) greater than palmitic acid (16:0) greater than stearic acid (18:0) greater than oleic acid (18:1) . However, endogenously synthesised saturated fatty acids, most of which were 16:0, were preferred over the supplied ones . A fraction of the exogenous 14:0 was elongated to 16:0 . Absence of saturated fatty acids increased the incorporation of 18:1 . The maximum extent of modification was one acyl chain for protein T2, on the exterior surface and two acyl chains for protein D12, spanning them membrane . Exogenously supplied fatty acids were incorporated into membrane lipids in proportion to their occurrence . However, the acylated proteins always contained 8-10 times more saturated chains than did the lipids . When exogenously supplied, all A . laidlawii polar membrane lipids could donate acyl chains to the acylated proteins but the neutral fraction (fatty acids and diacylglycerol) was most efficient . An incorporation into the acylated proteins of labelled cysteine, but not glucose or glycerol, was observed . Acylated proteins with different chains interacted similarly with a Triton X-114 detergent phase, and no full-size proteins (or acylated fragments) were released from cells by proteolytic enzymes . The results indicate an anchoring with peptide segments in addition to the acyl chains . Both 14:0 and 16:0 were attached at one end of both T2 and D12, but the N-terminal methionine of T2 was not acylated . The extent of modification and preference for saturated chains in the A . laidlawii membrane acylated proteins is more similar to eukaryotic than to eubacterial proteins.

J Bacteriol, 1992 Feb, 174(4), 1213 - 21
Mutagenesis of ribosomal protein S8 from Escherichia coli: defects in regulation of the spc operon; Wower I et al.; The structural features of Escherichia coli ribosomal protein S8 that are involved in translational regulation of spc operon expression and, therefore, in its interaction with RNA have been investigated by use of a genetic approach . The rpsH gene, which encodes protein S8, was first inserted into an expression vector under the control of the lac promoter and subsequently mutagenized with methoxylamine or nitrous acid . A screening procedure based on the regulatory role of S8 was used to identify mutants that were potentially defective in their ability to associate with spc operon mRNA and, by inference, 16S mRNA . In this way, we isolated 39 variants of the S8 gene containing alterations at 34 different sites, including 37 that led to single amino acid substitutions and 2 that generated premature termination codons . As the mutations were distributed throughout the polypeptide chain, our results indicate that amino acid residues important for the structural integrity of the RNA-binding domain are not localized to a single segment . Nonetheless, the majority were located within three short sequences at the N terminus, middle, and C terminus that are phylogenetically conserved among all known eubacterial and chloroplast versions of this protein . We conclude that these sites encompass the main structural determinants required for the interaction of protein S8 with RNA.

Cell Immunol, 1992 Feb, 139(2), 455 - 67
Arthritis by autoreactive T cell lines obtained from rats after injection of intestinal bacterial cell wall fragments; Klasen IS et al.; T cell lines (B13, B19) were isolated from the lymph nodes of Lewis rats 12 days after an arthritogenic injection of cell wall fragments of Eubacterium aerofaciens (ECW), a major resident of the human intestinal flora . These cell wall fragments consist of peptidoglycan polysaccharide complexes (PPC) . The cell lines that bear the helper phenotype were arthritogenic in knee or ankle joints upon intravenous injection into irradiated Lewis recipients . B13 was, however, not arthritogenic in irradiated F344 recipients that are largely RT1 identical . The arthritis induced in the knee joints of the irradiated Lewis rats was clearly shown by a 99mtechnetium-pertechnetate scanning technique and was confirmed histologically . In vitro the cell lines showed a proliferative response after stimulation with syngeneic spleen cells alone . The proliferation was significantly higher when bacterial PPC, isolated in soluble form from normal feces or ileostomy fluid were added . Recognition by B13 appeared to be MHC class II restricted . These results show that autoreactive T cell lines can be isolated from rats after injection of bacterial cell wall antigens and that these cell lines can be arthritogenic . This suggests a role for autoreactive T cells in the induction of bacterial cell wall arthritis and might give a clue for the arthritogenic properties of the normal human intestinal flora.

J Bacteriol, 1992 Feb, 174(3), 824 - 31
Immunocytochemical analysis of AlgP (Hp1), a histonelike element participating in control of mucoidy in Pseudomonas aeruginosa; Deretic V et al.; AlgP, a protein with an unusual carboxy-terminal domain resembling the tails of eukaryotic H1 histones, was detected in whole-cell extracts and within the cells of Pseudomonas aeruginosa by using immunoblotting and immunoelectron microscopy analyses . One known function of AlgP is its participation in the transcriptional activation of the algD gene . This is a pivotal step in the establishment of mucoidy in P . aeruginosa; mucoidy is a critical virulence factor expressed during respiratory infections in patients with cystic fibrosis . Polyclonal and monoclonal antibodies were raised against a synthetic 50-mer peptide containing two sets of six tandem repeats of the motif Lys-Pro-Ala-Ala (and its single-amino-acid substitution variants), based on the sequence of the algP gene from the standard genetic strain PAO . Western immunoblots with these antibodies and total protein extracts from P . aeruginosa revealed two polypeptides that reacted with the antibodies in all of the P . aeruginosa strains tested . The detected polypeptides displayed strain-dependent variability in their electrophoretic mobility, in accordance with the previously noted variability of the algP repeats at the DNA level . In strain PAO, the recognized polypeptides had apparent masses of 46.4 and 41.6 kDa . Immunoelectron microscopy revealed that AlgP is an intracellular protein with a wide distribution suggestive of its more general role . To indicate that fact, AlgP is given here an alternative name, Hp1 . Since AlgP (Hp1) is a eubacterial histonelike element displaying sequence and domanial similarity with eukaryotic H1 histones, these findings may have implications on the understanding of the organization of the prokaryotic nucleoid and its role in the control of gene expression and bacterial virulence.

Can J Microbiol, 1992 Feb, 38(2), 85 - 91
O-acetylated peptidoglycan: its occurrence, pathobiological significance, and biosynthesis; Clarke AJ et al.; Bacterial cell walls and their structural units, particularly peptidoglycan, induce a vast variety of biological effects in host organisms . The pathobiological effects of peptidoglycan are greatly enhanced by various modifications and substitutions to its basic composition and structure . One such modification is the presence of acetyl moieties at the C-6 hydroxyl group of N-acetylmuramyl residues, and to date, 11 species of eubacteria, including some important human pathogens, such as Neisseria gonorrhoeae, Proteus mirabilis, and Staphylococcus aureus, are known to possess O-acetylated peptidoglycan . This review addresses the influence of O-acetylation of peptidoglycan on its resistance to degradation both in vitro and in vivo, the clinical importance of the modification, and the currently held views on the pathway for its biosynthesis.

Int J Biol Macromol, 1992 Feb, 14(1), 41 - 4
Unfolding of the tertiary structure of specific tRNA and ribosomal 5S RNA from plants as studied with hydroxyl radicals; Barciszewska MZ et al.; Ribosomal 5S RNA is present in all eubacterial and eukaryotic ribosomes . Despite a large amount of experimental data on the primary and secondary structures of these types of molecules, details of their tertiary structure and their precise function in protein biosynthesis are still not known . Recently we have proposed a new model for the tertiary structure of plant 5S rRNA . In this study we applied the Fe(II)-mediated cleavage reaction to test the model . The data presented here provide experimental evidence that in the 5S rRNA molecule only a few nucleotides are buried in the tertiary structure . Similar experiments performed with methionine initiator tRNA gave results which imply the difference in its structure when compared with the X-ray structure of yeast tRNAPhe.

Biochemistry, 1992 Jan 21, 31(2), 328 - 33
Contributions of phylogenetically variable structural elements to the function of the ribozyme ribonuclease P; Darr SC et al.; Ribonuclease P (RNase P) is a ribonucleoprotein enzyme which participates in processing precursor tRNAs . The RNA subunit contains the catalytic site and is capable of catalysis in the absence of the protein subunit . RNase P RNAs from various eubacteria consist of a core of conserved sequence and secondary structure which is evolutionarily modified in different organisms by the presence of discrete helical elements at various sites in the RNAs . The variable occurrence of these helical elements suggests that they have no important functional role in the enzyme . The Escherichia coli RNase P RNA contains four such elements . It has been shown that simultaneous deletion of all four of them produces an RNA that is functional but has several significant defects which could arise from general disruption of the RNA or from the loss of element-specific functions . This paper describes a more detailed analysis of the role of the variable elements in E . coli RNase P RNA . Removal of one of the elements had no apparent effect on RNase P activity in vitro . Two other elements are required for correct folding of the RNA: their absence confers a requirement for extremely high monovalent salt concentrations, apparently to reduce intramolecular electrostatic repulsion . The fourth element that was tested participates in a long-range structural interaction (pseudoknot) which contributes to the structural stability of the enzyme and affects substrate binding affinity . In the absence of this helix, the RNA becomes temperature-sensitive, and the KM increases 100-fold.(ABSTRACT TRUNCATED AT 250 WORDS)

Eur J Biochem, 1992 Jan 15, 203(1-2), 81 - 7
The protein sequence of glutamate dehydrogenase from Sulfolobus solfataricus, a thermoacidophilic archaebacterium . Is the presence of N-epsilon-methyllysine related to thermostability?
Maras B, Consalvi V, Chiaraluce R, Politi L, De Rosa M, Bossa F, Scandurra R, Barra D.
The complete amino acid sequence of glutamate dehydrogenase from the thermoacidophilic archaebacterium Sulfolobus solfataricus has been determined . The sequence was reconstructed by automated sequence analysis of peptides obtained after cleavage by trypsin, cyanogen bromide, Staphylococcus aureus V8 protease and pepsin . The enzyme subunit is composed of 421 amino acid residues yielding a molecular mass of 46.078 kDa . The presence of N-epsilon-methyllysine in six positions of the sequence was observed . Comparison of the sequence of glutamate dehydrogenase from S . solfataricus with the other known primary structures of the corresponding enzyme from different sources, gives an overall identity of 9.2% and shows a symmetrical evolutionary distance of this archaebacterial protein from the two groups of vertebrate on one side and eubacterial and low eucaryote enzymes on the other side . The occurrence of specific substitutions and a possible role for N-epsilon-methylation of lysine residues are discussed in view of current hypotheses on the molecular basis of thermal adaptation of proteins.

J Endod, 1992 Jan, 18(1), 28 - 31
Antibodies in normal and diseased pulps reactive with microorganisms isolated from deep caries; Hahn CL et al.; Immunoglobulin molecules in the supernatant fluids (SF) from pulpal explant cultures have been observed to react with microorganisms implicated in infections of root canals . In this study, the reactivity of immunoglobulin molecules in the SF from normal and irreversible pulpitis pulps to six strains of predominant microorganisms isolated from the immediate layer of carious lesions above the pulps used for explant cultures was investigated using an enzyme-linked immunosorbent assay . Two ATCC strains of Eubacterium were also included in this assay . Specific antibodies to Lactobacillus casei subsp . casei, Lactobacillus casei subsp . rhamnosus, Lactobacillus acidophilus (I), (II), Streptococcus mutans, Bacteroides intermedius, Eubacterium brachy, and Eubacterium alactolyticum in the SF from the normal and irreversible pulpitis tissues were observed with a large variation of antibody levels in both groups . Immunodiffusion assays of the SF revealed that IgG was the major class of immunoglobulin in the normal as well as the irreversible groups . The presence of natural antibodies in the normal pulps suggested a possible protective role of antibodies during the invasive process of caries.

Gene, 1992 Jan 2, 110(1), 17 - 23
Structure of the dnaA and DnaA-box region in the Mycoplasma capricolum chromosome: conservation and variations in the course of evolution; Fujita MQ et al.; We have previously shown that the dnaA gene and the DnaA-box region were conserved in bacteria representative of all three major branches of the eubacterial phylogenic tree: high G + C Gram+, low-G + C Gram+ and Gram- . In the present work, we determined the structure of the dnaA region of Mycoplasma capricolum and found that the dnaA gene and at least two other genes, rpmH and dnaN, were conserved in this bacterium . An unusually high level of amino acid (aa) substitutions was observed in M . capricolum DnaA . It was the case even in those aa which were well conserved in other bacterial species . The nontranslatable region upstream from the dnaA gene was also conserved in this bacterium, as it was universally found in both Gram+ and Gram- bacteria . An additional nontranslatable region downstream from the dnaA gene, which is common to Gram+ bacteria, was also found in M . capricolum, consistent with the proposal that M . capricolum is Gram+ in origin . These regions were rich in A + T and contained ten DnaA-box-like sequences (9-mers that differ from TTATCCACA by one or two bases).

G Batteriol Virol Immunol, 1992 Jan-1993 Dec, 85(1-12), 3 - 11
The role of antibiotics in the evolution of microorganisms; Cavallo G; Bacteria were the first living beings to appear on our planet: the most ancient fossils available, all of them were procaryotic microorganisms, developed 4, 5 billion years ago . The paleomicrobiological studies made on that kind of fossils, which are by now several hundreds in each continent, proved bacteria to have constantly evolved and to have originated the modern Eubacteria as well as the Archebacteria and the Cianobacteria . These last appeared about 2 billion years ago and, having acquired the oxygen-type photosynthesis, have caused the formation of a large amount of organic material, afterwards used by the younger organisms, and have modified the atmosphere introducing oxygen in it and conditioning in this way the other living being's evolution . From Bacteriaceae and Cyanobacteria derive the eucaryotic microorganisms (algae, fungi, protozoa, mould) and, little by little, all the other organisms both vegetable and animal subjected to the evolutionary pressures . Nevertheless bacteria undergo more frequently than all the others the evolution law because of their short reproductive time; this is the reason why bacteria are favourite compared with the other organisms . In fact each species is subjected to a genetic mutation every 10(5)/10(6) generations: in the vegetables and animals the consequences of a genetic mutation will be evident after millennia whereas in the bacteria the mutation happens in a short time . We ourselves have witnessed the revolution which took place in the bacteria populations during the last half century when numerous antibiotic-resistant strains appeared.(ABSTRACT TRUNCATED AT 250 WORDS)

J Bacteriol, 1992 Jan, 174(1), 298 - 302
Cloning, expression, and sequencing of squalene-hopene cyclase, a key enzyme in triterpenoid metabolism; Ochs D et al.; The pentacyclic hopanoids, a class of eubacterial lipids, are synthesized by squalene-hopene cyclase and side chain-elongating enzymes . With the aid of DNA probes based on the amino-terminal sequence of purified squalene-hopene cyclase from Bacillus acidocaldarius, clones of Escherichia coli that express this enzyme in the cytoplasmic membrane were isolated . According to the DNA sequence, the cyclase contained 627 amino acids with a molecular mass of 69,473 Da . A high percentage of the amino acids were basic . No significant similarity to existing sequenced proteins was found.

C R Acad Sci III, 1992, 314(6), 253 - 8
{Purification, amino acid composition and N-terminal sequence of the major protein (protein H) of the outer membrane of Pasteurella multocida}; Chevalier G et al.; The major protein (protein H) of the outer membrane of Pasteurella multocida was purified by size-exclusion chromatography after selective extraction with detergents . The protein forms homotrimers which are stable in the presence of SDS at room temperature . Upon treatment at 100 degrees C, the protein is fully dissociated by the detergent into monomers exhibiting an apparent molecular mass of 37 kDa as estimated by electrophoresis . The amino acid composition of protein H is characterized by a low hydropathy index (HI = -0.40) and is strongly related to the compositions of bacterial porins, notably porins P2 (Haemophilus influenzae), PIA (Neisseria gonorrhoeae) and Cl.2 ("class 2 porin" of N . meningitidis) . The N-terminal amino acid sequence of protein H shares a strong homology with those of porins OmpC (Escherichia coli) and P2 . These data indicate that protein H of P . multocida is a porin belonging to the superfamily of the non-specific porins of Gram-negative eubacteria outer membrane.

FEMS Microbiol Lett, 1992 Jan 1, 69(2), 129 - 34
Correlation between glycosylation of flagellin proteins and sensitivity of flagellar filaments to Triton X-100 in methanogens; Faguy DM et al.; The flagellins of Methanospirillum hungatei strains JF1 and GP1, Methanococcus deltae, and Methanothermus fervidus are glycosylated . Isolated flagellar filaments from these organisms are dissociated by low concentrations (0.5% (v/v)) of Triton X-100 . Flagellar filaments from other methanogens (Methanococcus voltae, Methanococcus vannielii and Methanoculleus marisnigri) composed of non-glycosylated flagellins are resistant to Triton X-100 treatment . Consequently, the isolation techniques (employing Triton X-100) used to isolate basal body-hook-filament complexes in eubacteria may not be applicable to many methanogens.

J Periodontal Res, 1992 Jan, 27(1), 15 - 9
Predominant obligate anaerobes in human periodontal pockets; Uematsu H et al.; This study was carried out to investigate the predominant anaerobic bacteria of periodontal pockets in patients with advanced periodontitis, who had no previous treatment other than supragingival scaling, no history of recent or chronic systemic illness, nor any intake of antibiotics within 6 weeks prior to bacteriological sampling . Care was taken not to ignore tiny-colony-forming anaerobes, by means of a stereoscope and an anaerobic glove box system . Out of 422 (100%) isolates, 380 (90%) were obligate anaerobes, suggesting that the environment in periodontal pockets was anaerobic and favors the growth of obligate anaerobes . Among the 380 obligate anaerobes isolated, strains belonging to Eubacterium (54%) were predominant, and many of them occurred in tiny colonies . The other obligate anaerobes isolated were assigned to Wolinella (9%), unidentified motile rods which resemble Wolinella (7%), Peptostreptococcus (6%), Fusobacterium (5%), Bacteroides (2%; including those reclassified to Prevotella and Porphyromonas) and Selenomonas (0.5%) . Among the isolates, 67% were Gram-positive bacteria, including 59% of rods (mostly asaccharolytic Eubacterium), suggesting that these bacteria, particularly strains of the Eubacterium species, may play an important role in etiology of adult periodontitis.

Physiol Chem Phys Med NMR, 1992, 24(2), 109 - 17
Small acidic peptides from wheat germ chromatin . II . Regulatory activity in specific transcription systems reconstituted in vitro; Castigli E et al.; A variety of evidence suggests that a family of chromatin peptides (CPs), characterized by 1000D molecular weight, a pH dependent association to DNA and a prevailing presence of acidic amino acids in their structure, is involved in the regulation of genes expression . Nevertheless their action mechanism is still unknown . In our in vitro specific RNA transcription systems the CPs affect the initiation and not the elongation . Furthermore they inhibit the RNA transcription by interaction with the DNA rather than with the enzyme . The phagic in vitro specific RNA transcription is less affected by CPs than the eubacteric system, suggesting a kind of selectivity for target DNA sequences involved in the initiation of transcription.

Arch Microbiol, 1992, 157(6), 505 - 11
Purification and properties of a F420-nonreactive, membrane-bound hydrogenase from Methanosarcina strain Gö1; Deppenmeier U et al.; The distribution of the F420-reactive and F420-nonreactive hydrogenases from the methylotrophic Methanosarcina strain Go1 indicated a membrane association of the F420-nonreactive enzyme . The membrane-bound F420-nonreactive hydrogenase was purified 42-fold to electrophoretic homogeneity with a yield of 26.7% . The enzyme had a specific activity of 359 mumol H2 oxidized.min-1.mg protein-1 . The purification procedure involved dispersion of the membrane fraction with the detergent Chaps followed by anion exchange, hydrophobic and hydroxylapatite chromatography . The aerobically prepared enzyme had to be reactivated anaerobically . Maximal activity was observed at 80 degrees C . The molecular mass as determined by native gel electrophoresis and gel filtration was 77,000 and 79,000, respectively . SDS gel electrophoresis revealed two polypeptides with molecular masses of 60,000 and 40,000 indicating a 1:1 stoichiometry . The purified enzyme contained 13.3 mol S2-, 15.1 mol Fe and 0.8 mol Ni/mol enzyme . Flavins were not detected . The amino acid sequence of the N-termini of the subunits showed a higher degree of homology to eubacterial uptake-hydrogenases than to F420-dependent hydrogenases from other methanogenic bacteria . The physiological function of the F420-nonreactive hydrogenase from Methanosarcina strain Gol is discussed.

Arch Microbiol, 1992, 158(6), 394 - 7
Identification of sulphate-reducing ectosymbiotic bacteria from anaerobic ciliates using 16S rRNA binding oligonucleotide probes; Fenchel T et al.; The identity of ectosymbiotic bacteria of some marine, free-living anaerobic ciliates (Metopus contortus, Caenomorpha levanderi and Parablepharisma sp.) was studied using fluorescent-dye-conjugated oligonucleotides complementary to short sequence elements of 16S ribosomal RNA . The ectosymbiotic bacteria of all species hybridized with a eubacterial probe and those of the two former mentioned species hybridized with a general probe for sulphate-reducing bacteria, but not to a probe specific for Desulfobacter . The results support indirect evidence suggesting that ectosymbiotic bacteria of anaerobic ciliates are sulphate-reducers which depend on host metabolites for substrates.

Biochem Soc Symp, 1992, 58, 7 - 21
The enzymology of archaebacterial pathways of central metabolism; Danson MJ et al.; From a comparison of the pathways of central metabolism in the archaebacteria, eubacteria and eukaryotes, it is clear that the basic pathways were established before the divergence of the three kingdoms, but that the notable differences may provide important clues to their evolution . From these comparisons, enzymes found in all evolutionary groups have been chosen for detailed structural studies; given the range of extreme phenotypes found within the archaebacteria, these studies will be crucial to our understanding of the structural basis for protein stability and how such features may be engineered into a protein of choice.

Int Endod J, 1992 Jan, 25(1), 2 - 5
Bacterial invasion of non-exposed dental pulp; Hoshino E et al.; Anaerobic procedures were adopted to demonstrate the early bacterial invasion of non-exposed dental pulps, and to isolate and identify the bacteria . Of 19 freshly extracted teeth which originally exhibited deep dentinal lesions, clinical examination and electric pulp testing showed that nine of them had no pulpal exposure . Thus the pulps of these teeth were covered by clinically sound dentine beneath the carious lesion . Bacteria were found to have invaded the pulps of six of these nine teeth . The predominant bacteria were obligate anaerobes belonging to the genera Eubacterium, Propionibacterium and Actinomyces . Other obligate anaerobes were Lactobacillus, Peptostreptococcus, Veillonella and Streptococcus . The bacterial composition resembled that of the deep layers of dentinal lesions described previously, suggesting that the bacteria isolated in this study had passed through some individual dentinal tubules, to invade the dental pulp.

Biosystems, 1992, 27(1), 53 - 62
Phylogenetic consequences of symbioses: Eukarya and Eubacteria are not monophyletic taxa; Mindell DP; In the past systematists have not been concerned with distinguishing the different phylogenetic histories for symbiont taxa that have merged within a composite taxon, or holobiont . I suggest that symbionts can retain their status as discrete taxa and that their independent histories can be included in phylogenetic analyses intending to discover monophyletic groups . Use of reticulate branches to include independent histories for different symbionts, incorporates our improving understanding of evolution and provides greater accuracy in denoting monophyletic groups . In an expanded view, a monophyletic group includes only and all the descendants of the merged-symbionts' common ancestor . Holobiont taxa will have constituent symbionts included in different monophyletic groups and there will be a reduction in the number of monophyletic groups recognized, particularly at higher taxonomic levels . As a consequence of considering symbioses in phylogenetic analyses, the proposed taxa Eubacteria and Eukarya are seen to be non-monophyletic, and, thus, poor indicators of evolutionary history.

Biosystems, 1992, 27(1), 39 - 51
Biodiversity: molecular biological domains, symbiosis and kingdom origins; Margulis L; The number of extant species of organisms is estimated to be from fewer than 3 to more than 30 x 10(6) (May, 1992) . Molecular biology, comparative genetics and ultrastructural analyses provide new insights into evolutionary relationships between these species, including increasingly precise ideas of how species and higher taxa have evolved from common ancestors . Accumulation of random mutations and large macromolecular sequence change in all organisms since the Proterozoic Eon has been importantly supplemented by acquisition of inherited genomes ('symbiogenesis') . Karyotypic alterations (polyploidization and karyotypic fissioning) have been added to these other mechanisms of species origin in plants and animals during the Phanerozoic Eon . The new evolution concepts (coupled with current rapid rates of species extinction and ignorance of the extent of biodiversity) prompted this analysis of the field of systematic biology and its role in the reorganization of extant species into higher taxa . Two superkingdoms (= Domains: Prokaryotae and Eukaryotae) and five kingdoms (Monera = Procaryotae or Bacteria; Protoctista: algae, amoebae, ciliates, foraminifera, oomycetes, slime molds, etc.; Mychota: 'true' fungi; Plantae: one phylum (division) of bryophytes and nine phyla of tracheophytes; and Animalia) are recognized . Two subkingdoms comprise the monera: the great diverse lineages are Archaebacteria and Eubacteria . The criteria for classification using molecular, ultrastructural and genetic data for this scheme are mentioned . For the first time since the nineteenth century, logical, technical definitions for each group are given with their time of appearance as inferred from the fossil record in the primary scientific literature . This classification scheme, which most closely reflects the evolutionary history, molecular biology, genetics and ultrastructure of extant life, requires changes in social organization of biologists, many of whom as botanists and zoologists, still behave as if there were only two important kingdoms (plants and animals).

Sci Prog, 1992, 76(301-302 Pt 3-4), 565 - 80
Chemical anatomy of antibiotic resistance: chloramphenicol acetyltransferase; Shaw WV; The evolution of mechanisms of resistance to natural antimicrobial substances (antibiotics) was almost certainly concurrent with the development in microorganisms of the ability to synthesise such agents . Of the several general strategies adopted by bacteria for defence against antibiotics, one of the most pervasive is that of enzymic inactivation . The vast majority of eubacteria that are resistant to chloramphenicol, an inhibitor of prokaryotic protein synthesis, owe their resistance phenotype to genes for chloramphenicol acetyltransferase (CAT), which catalyses O-acetylation of the antibiotic, using acetyl-CoA as the acyl donor . The structure of CAT is known, as are many of the properties of the enzyme which explain its remarkable specificity and catalytic efficiency . Less clear is the evolutionary pathway which has produced the different members of the CAT 'family' of enzymes . Hints come from other acetyltransferases which share structure and mechanistic features with CAT, while not being strictly 'homologous' at the level of amino acid sequence . The 'super-family' of trimeric acetyltransferases appears to have in common a chemical mechanism based on a shared architecture.

Biosystems, 1992, 28(1-3), 15 - 32
The nature of the last universal ancestor and the root of the tree of life, still open questions; Forterre P et al.; The nature of the last universal ancestor to all extent cellular organisms and the rooting of the universal tree of life are fundamental questions which can now be addressed by molecular evolutionists . Several scenarios have been proposed during the last years, based on the phylogenies of ribosomal RNA and of duplicated proteins, which suggest that the last universal ancestor was either an RNA progenote or an hyperthermophilic prokaryote . We discuss these hypotheses in the light of new data on the evolution of DNA metabolizing enzymes and of contradictions between different protein phylogenies . We conclude that the last universal ancestor was a member of the DNA world already containing several DNA polymerases and DNA topoisomerases . Furthermore, we criticize current data which suggest that the rooting of the universal tree of life is located in the eubacterial branch and we conclude that both rooting the universal tree and the nature of the last universal ancestor are still open questions.

J Bacteriol, 1992 Jan, 174(1), 137 - 43
Hydrogen-oxidizing electron transport components in the hyperthermophilic archaebacterium Pyrodictium brockii; Pihl TD et al.; The hyperthermophilic archaebacterium Pyrodictium brockii grows optimally at 105 degrees C by a form of metabolism known as hydrogen-sulfur autotrophy, which is characterized by the oxidation of H2 by S0 to produce ATP and H2S . UV-irradiated membranes were not able to carry out the hydrogen-dependent reduction of sulfur . However, the activity could be restored by the addition of ubiquinone Q10 or ubiquinone Q6 to the UV-damaged membranes . A quinone with thin-layer chromatography migration properties similar to those of Q6 was purified by thin-layer chromatography from membranes of P . brockii, but nuclear magnetic resonance analysis failed to confirm its identity as a ubiquinone . P . brockii quinone was capable of restoring hydrogen-dependent sulfur reduction to UV-irradiated membranes . Hydrogen-reduced-minus-air-oxidized absorption difference spectra on membranes revealed absorption peaks characteristic of c-type cytochromes . A c-type cytochrome with alpha, beta, and gamma peaks at 553, 522, and 421 nm, respectively, was solubilized from membranes with 0.5% Triton X-100 . Pyridine ferrohemochrome spectra confirmed its identity as a c-type cytochrome, and heme staining of membranes loaded on sodium dodecyl sulfate gels revealed a single heme-containing component of 13 to 14 kDa . Studies with the ubiquinone analog 2-n-heptyl-4-hydroxyquinoline-N-oxide demonstrated that the P . brockii quinone is located on the substrate side of the electron transport chain with respect to the c-type cytochrome . These first characterizations of the strictly anaerobic, presumably primitive P . brockii electron transport chain suggest that the hydrogenase operates at a relatively high redox potential and that the H2-oxidizing chain more closely resembles those of aerobic eubacterial H2-oxidizing bacteria than those of the H2-metabolizing systems of anaerobes or the hyperthermophile Pyrococcus furiosus.

Ciba Found Symp, 1992, 171, 64 - 80; discussion 80-7
Origins of secondary metabolism; Cavalier-Smith T; Secondary metabolites generally benefit their producers as poisons that protect them against competitors, predators or parasites . They are produced from universally present precursors (most often acetyl-CoA, amino acids or shikimate) by specific enzymes that probably arose by the duplication and divergence of genes originally coding for primary metabolism . Most secondary metabolites are restricted to single major taxa on the universal phylogenetic tree and so probably originated only once . But different secondary metabolic pathways have originated from different ancestral enzymes at radically different times in evolution . Secondary metabolites are most abundantly produced by microorganisms in crowded habitats and by plants, fungi and sessile animals like sponges, where chemical defence and attack rather than physical escape or fighting are at a premium . The first secondary metabolites were probably antibiotics produced in microbial mats over 3500 million years ago . These first ecosystems probably consisted entirely of eubacteria: archaebacteria and eukaryotes arose much later . As a phylogenetic context for considering the earliest origins of antibiotics I summarize a cladistic analysis of the explosive eubacterial primary diversification . This suggests that the most primitive surviving cells are the photosynthetic heliobacteria . Study of these and of the nearly as primitive chloroflexibacteria, spirochaetes and deinobacteria may provide the best evidence on the origins of secondary and primary metabolism.

Nucleic Acids Res, 1991 Dec 11, 19(23), 6491 - 7
A mixed group II/group III twintron in the Euglena gracilis chloroplast ribosomal protein S3 gene: evidence for intron insertion during gene evolution; Copertino DW et al.; The splicing of a 409 nucleotide intron from the Euglena gracilis chloroplast ribosomal protein S3 gene (rps3) was examined by cDNA cloning and sequencing, and northern hybridization . Based on the characterization of a partially spliced pre-mRNA, the intron was characterized as a 'mixed' twintron, composed of a 311 nucleotide group II intron internal to a 98 nucleotide group III intron . Twintron excision is via a 2-step sequential splicing pathway, with removal of the internal group II intron preceding excision of the external group III intron . Based on secondary structural analysis of the twintron, we propose that group III introns may represent highly degenerate versions of group II introns . The existence of twintrons is interpreted as evidence that group II introns were inserted during the evolution of Euglena chloroplast genes from a common ancestor with eubacteria, archaebacteria, cyanobacteria, and other chloroplasts.

Proc Natl Acad Sci U S A, 1991 Dec 1, 88(23), 10783 - 7
The plastid genome of Cryptomonas phi encodes an hsp70-like protein, a histone-like protein, and an acyl carrier protein; Wang SL et al.; The plastid genome of Cryptomonas phi, a cryptomonad alga, contains three genes that have not previously been found in any organellar genome . Each of these genes encodes a functional class of organellar gene product not previously reported . The first gene, dnaK, encodes a polypeptide of the hsp70 heat shock protein family . The predicted amino acid sequence of the DnaK protein is 54% identical to that of the Escherichia coli hsp70 protein (DnaK), 50-53% identical to that of two nucleus-encoded mitochondrial hsp70 proteins, and 43-46% identical to that of several eukaryotic cytoplasmic members of the hsp70 protein family . The second gene, hlpA, encodes a polypeptide resembling bacterial histone-like proteins . The predicted amino acid sequence of the HlpA protein is 25-53% identical to that of several bacterial histone-like proteins, and the identity increases to 39-76% over a conserved region corresponding to the long arm that binds DNA . The third gene, acpA, encodes an acyl carrier protein, which is a key cofactor in the synthesis and metabolism of fatty acids . Its predicted amino acid sequence is 36-59% identical to that of eubacterial and plant chloroplast (nucleus-encoded) acyl carrier proteins.

Curr Opin Genet Dev, 1991 Dec, 1(4), 457 - 63
Early evolution and the origin of eukaryotes; Sogin ML; Our understanding of evolutionary relationships in the eukaryotic world has been revolutionized by molecular systematics . Phylogenies based upon comparisons of rRNAs define five major eukaryotic assemblages plus a series of paraphyletic protist lineages . Comparison of conserved genes that were duplicated prior to the divergence of eubacteria, archaebacteria, and eukaryotes, positions the root of the universal tree within the eubacterial line of descent . In this review a novel model is presented which uses the rRNA and protein based phylogenies to describe the evolutionary origins of eukaryotes.

Glycobiology, 1991 Dec, 1(6), 545 - 51
Bacterial surface layer glycoproteins; Messner P et al.; Crystalline cell surface layers (S-layers) are ubiquitously present in bacterial species from almost all phylogenetic branches . Recent investigations have shown that the S-layer proteins of many archaebacteria and eubacteria contain covalently linked carbohydrate chains . This evidence clearly shows that the ability for protein glycosylation is present as a common biosynthetic pathway in prokaryotic organisms.

Antimicrob Agents Chemother, 1991 Dec, 35(12), 2655 - 7
Characterization of unusual tetracycline-resistant gram-positive bacteria; Roberts MC et al.; Tetracycline-resistant Tet M-negative isolates of Actinomyces viscosus, Eubacterium lentum, Mobiluncus curtisii, and Mobiluncus mulieris were screened with the Tet K, Tet L, and Tet O DNA probes . Ten (71%) of the resistant Mobiluncus strains hybridized with the Tet O probe, two of the three E . lentum strains hybridized with the Tet K probe, and the A . viscosus isolate hybridized with the Tet L probe.

Gene, 1991 Dec 1, 108(1), 133 - 7
A dnaK homolog in the archaebacterium Methanosarcina mazei S6; Macario AJ et al.; A fragment of genomic DNA cloned from the methanogenic archaebacterium, Methanosarcina mazei strain S6, was found to contain an 1857-bp open reading frame (ORF) . A sequence matching the consensus ribosome-binding sequence determined for other methanogens was found upstream from the ORF . The amino acid (aa) sequence encoded by the ORF was compared with reference sequences and was found to be related to six DnaK sequences determined for five species of eubacteria (none exist for archaebacteria) . The M . mazei S6 aa sequence was over 61% identical and over 77% similar (identities plus conservative substitutions) to the closest four reference sequences, which were all DnaKs . The gene described here is therefore proposed to be the first member of the dnaK family sequenced from the archaebacterial kingdom (Archaea) . This finding confirms that DnaK proteins are highly conserved, occurring not only in eubacteria (Bacteria) and eukaryotes (Eucaria), but also in archaebacteria (Archaea).

J Med Microbiol, 1991 Dec, 35(6), 338 - 44
Surface layers of Eubacterium yurii subsp . yurii and their possible role in test-tube brush formation and iron acquisition; Krywolap GN et al.; Eubacterium yurii subsp . yurii is an anaerobic, gram-positive rod . On isolation E . yurii forms cellular arrangements resembling test-tube brushes (TTB) . Although TTB decreased in size and number on repeated laboratory subculture in enriched media, media poor in available iron enhanced formation of these . Plasmids were not demonstrated, even after chloramphenicol enhancement . To characterise the nature and possible physiological roles of the structures of the TTB, they were examined by transmission electronmicroscopy (TEM) with thin-section, negative-staining, shadow-casting, freeze-etching and freeze-fracturing techniques, and by scanning electronmicroscopy (SEM) . Previous studies by phase-contrast microscopy revealed an amorphous core, the size of which varied in direct proportion to the number of associated bacterial cells . Thin sections of the TTB showed a gram-positive cell wall with additional surface layers . Negative staining, shadow casting and freeze etching revealed a surface layer comprising subunits in tetragonal array (P4 symmetry) . Shadow casting showed also that the outermost layer of the cells was composed of fibrillar structures closely associated with but distinct from, the tetragonal layer . The fibrils extended from the cell surface in clumps or strands . The presence of these fibrils was confirmed by the freeze-fracture technique and SEM . Chemical analysis of the core material of the TTB showed it to be low in carbohydrate (0.06%) and protein (0.2%) . Energy-dispersive X-ray spectrometry showed that the core was composed mostly of iron.(ABSTRACT TRUNCATED AT 250 WORDS)

J Biol Chem, 1991 Nov 15, 266(32), 21578 - 83
Protein topography of Sulfolobus solfataricus ribosomes by cross-linking with 2-iminothiolane . Sso L12e, Sso L10e, and Sso L11e are neighbors; Casiano C et al.; Large ribosomal subunits from Sulfolobus solfataricus were cross-linked with 2-iminothiolane in order to investigate the arrangement of proteins in the region containing the multicopy acidic protein Sso L12e, the protein homologous to Escherichia coli L7/L12 . Proteins from cross-linked 50 S subunits were extracted and fractionated by chromatography on CM-cellulose . Fractions containing Sso L12e were analyzed by "diagonal" (two-dimensional reducing/nonreducing) dodecyl sulfate polyacrylamide gel electrophoresis . Sso L12e appeared in cross-linked homodimers and also in cross-linked complexes that contained Sso L10e, the protein equivalent to E . coli L10 . In addition, Sso L12e was found in cross-links to L4, L6a, L26, and L29 . N-terminal sequences obtained for L6a and L26 showed them to have significant homologies to E . coli proteins L11 and L23, respectively . The results indicate the presence in this archaebacterial ribosome of Sso L12e dimers and their location near Sso L10e and Sso L11e . The Sso L12e-L29 (Sso L23e) cross-link suggests proximity between components of the factor-binding and peptidyltransferase domains, since E . coli L23 is a protein affinity-labeled by puromycin . The (Sso L12e)4-Sso L10 pentameric complex, identified previously from studies in solution, appears to represent correctly the arrangement of these proteins in the ribosome . The occurrence in the archaebacterial ribosome of this unique structural element, similar to those shown previously in eubacteria and eukaryotes, reinforces the concept that the protein quaternary structure of the ribosomal factor-binding domain is highly conserved.

Eur J Biochem, 1991 Nov 15, 202(1), 85 - 93
Purification and characterization of a monomeric isocitrate dehydrogenase with dual coenzyme specificity from the photosynthetic bacterium Rhodomicrobium vannielii; Leyland ML et al.; An isocitrate dehydrogenase able to function with either NADP or NAD as coenzyme was purified to homogeneity from cell-free extracts of the purple photosynthetic eubacterium Rhodomicrobium vannielii using a rapid two-step procedure involving dye-ligand affinity chromatography . The enzyme was obtained in 60% yield with specific activities of 23 U.mg protein-1 (NADP-linked reaction) and 18.5 U.mg protein-1 (NAD-linked reaction) . The purified enzyme was monomeric and migrated with an approximate Mr of 75,000-80,000 on both SDS/PAGE and non-denaturing PAGE . Affinity constants (Km values) of 2.5 microM for NADP and 0.77 mM for NAD and values for kcat/Km of 981,200 min-1.mM-1 (NADP) and 2455 min-1.mM-1 (NAD) indicated a greater specificity for NADP compared to NAD . A number of metabolites were examined for possible differential regulatory effects on the NADP- and NAD-linked reactions, using a dual-wavelength assay . Oxaloacetate was found to be an effective inhibitor of both reactions and the enzyme was also sensitive to concerted inhibition by glyoxylate and oxaloacetate . The amino-acid composition and the identity of 39 residues at the N-terminus were determined and compared to other isocitrate dehydrogenases . The results suggested a relationship between the Rm . vannielii enzyme and the monomeric isocitrate dehydrogenase isoenzyme II from Vibrio ABE-1.

Science, 1991 Nov 8, 254(5033), 853 - 6
Long-range structure in ribonuclease P RNA; Haas ES et al.; Phylogenetic-comparative and mutational analyses were used to elucidate the structure of the catalytically active RNA component of eubacterial ribonuclease P (RNase P) . In addition to the refinement and extension of known structural elements, the analyses revealed a long-range interaction that results in a second pseudoknot in the RNA . This feature strongly constrains the three-dimensional structure of RNase P RNA near the active site . Some RNase P RNAs lack this structure but contain a unique, possibly compensating, structural domain . This suggests that different RNA structures located at different positions in the sequence may have equivalent architectural functions in RNase P RNA.

Mol Biol (Mosk), 1991 Nov-Dec, 25(6), 1643 - 9
{Mutual location of the rRNA operon and tuf gene in the Mycoplasma gallisepticum strain S6 genome}; Skamrov AV et al.; The genomic library of Mycoplasma gallisepticum was constructed and two clones, selected by hybridization on E . coli 16S rRNA, were analyzed . The restriction map of the clones indicate that both clones belong to the same region of the M . gallisepticum genome . The results of Southern hybridization with either E . coli 16S rRNA, or E . coli 23S rRNA, or oligonucleotide synthesized as a part of M . gallisepticum 5S rRNA, led to the conclusion that the unspliced rRNA operon was cloned . The order of genes in the operon is common for eubacteria: 16S-23S-5S . The tuf gene of M . gallisepticum was mapped inside the cloned region 5 kb upstream of 16S gene by hydridization on E . coli tufA gene and oligonucleotide synthesized on the basis of M . gallisepticum tuf gene sequence . The direction of transcription of the gene and the expected direction of transcription of the rRNA operon coincide.

Chem Pharm Bull (Tokyo), 1991 Nov, 39(11), 2994 - 8
Effect of enzymatic sulfation on biochemical and pharmacological properties of catecholamines and tyrosine-containing peptides; Konishi-Imamura L et al.; Substrate specificity of a novel sulfotransferase produced by Eubacterium A-44 isolated from human feces has been studied . Phenolic drugs, catecholamines, were good acceptors of this bacterial enzyme . With regard to dopamine, sulfation mostly occurred at the 4-aromatic hydroxy group . We also investigated the effects of enzymatic sulfation on pharmacologically active phenolic compounds . Sulfation of phenolic compounds generally led to inactivation (e.g . tyramine and Leu-enkephalin), with the exception of cholecystokinin (CCK) and some gastrointestinal peptides . Proteolytic hydrolysis in vitro did not occur at the C-terminal of the sulfated tyrosine residues of peptides such as Leu-enkephalin and kyotorphin . These results suggest that the sulfation by bacterial enzyme plays an important role in detoxification, activation and stability of phenolic compounds in the human body.

J Gen Microbiol, 1991 Nov, 137 ( Pt 11), 2631 - 41
Membrane fatty acids as phenotypic markers in the polyphasic taxonomy of methylotrophs within the Proteobacteria; Guckert JB et al.; A polyphasic approach to bacterial taxonomy attempts to integrate phylogenetic relationships with phenotypic marker analysis . This study describes the application of membrane fatty acids as a phenotypic marker for methylotrophs . Detailed phospholipid, ester-linked fatty acid (PLFA) profiles are reported for 17 methylotrophic eubacterial strains . These profiles included verification of double bond positions and geometries, both critical features for this analysis . Multivariate cluster analysis was used to indicate groupings of these strains along with literature values of both methylotrophs and non-methylotrophs based on the PLFA phenotype . Like many phenotypic characteristics, PLFA profiles were influenced by environmental conditions . The instabilities displayed, however, were predictable from physiological studies including increased trans/cis and cyclopropyl/cis ratios . Cluster analysis of PLFA profiles generated by separate investigators with different culture conditions indicated reproducibility by strain and species . The PLFA phenotype relationships compare favourably with phylogenetic associations based on 16S rRNA data for methylotrophs and will continue to be a valuable phenotypic marker for Proteobacteria taxonomy.

Appl Environ Microbiol, 1991 Nov, 57(11), 3127 - 34
Isolation of a bacterium resembling Pirellula species from primary tissue culture of the giant tiger prawn (Penaeus monodon); Fuerst JA et al.; During attempts to establish tissue cultures from hepatopancreas, heart, and hemolymph of the giant tiger prawn (Penaeus monodon), using a medium including penicillin, streptomycin, and amphotericin B, bacterial contamination in the form of a sheet of growth attached to the tissue culture vessel was a persistent problem . Contaminant bacteria were teardrop-shaped cells arranged in rosettes, and electron microscopy revealed buds, crateriform structures, and the absence of a peptidoglycan layer in the cell wall, features characteristic of bacteria in the Planctomyces-Pirellula group, a phylogenetically distinct group of eubacteria . Two strains of contaminant bacteria were isolated in pure culture . Both exhibited morphology and antibiotic resistance consistent with their membership in the Planctomyces-Pirellula group (order Planctomycetales) of eubacteria . Tissue culture media for marine invertebrates may select for such bacteria if high concentrations of cell wall synthesis-inhibiting antibiotics are included.

Mol Microbiol, 1991 Nov, 5(11), 2589 - 97
Structure and function of DnaA and the DnaA-box in eubacteria: evolutionary relationships of bacterial replication origins; Yoshikawa H et al.; DnaA protein (a trans-acting element) and its binding sequence, DnaA-box: (a cis-acting element) are two elements essential for the initiation of chromosomal replication in Escherichia coli and other enteric bacteria . Recently these two elements have been found to be conserved in three Gram-positive bacteria (Bacillus subtilis, Micrococcus luteus and Mycoplasma capricolum) as well as in Gram-negative pseudomonads . DnaA protein was also found to be essential in the initiation of the replication of the B . subtilis chromosome, and regions containing multiple repeats of DnaA-box (DnaA-box region) are found to be active as autonomously replicating elements both in B . subtilis and pseudomonads . In this MicroReview we compare first the structures of these DnaA-box regions and their locations on the chromosome and then functional aspects of DnaA protein and DnaA-box regions in the initiation and regulation of chromosomal replication . From these observations we propose evolutionary relationships between replication origins of eubacteria.

Biochem J, 1991 Oct 15, 279 ( Pt 2), 601 - 4
Acyltransferase activities of the high-molecular-mass essential penicillin-binding proteins; Adam M et al.; The high-molecular-mass penicillin-binding proteins (HMM-PBPs), present in the cytoplasmic membranes of all eubacteria, are involved in important physiological events such as cell elongation, septation or shape determination . Up to now it has, however, been very difficult or impossible to study the catalytic properties of the HMM-PBPs in vitro . With simple substrates, we could demonstrate that several of these proteins could catalyse the hydrolysis of some thioesters or the transfer of their acyl moiety on the amino group of a suitable acceptor nucleophile . Many of the acyl-donor substrates were hippuric acid or benzoyl-D-alanine derivatives, and their spectroscopic properties enabled a direct monitoring of the enzymic reaction . In their presence, the binding of radioactive penicillin to the PBPs was also inhibited.

FEBS Lett, 1991 Oct 7, 291(1), 71 - 4
Mycoplasma gallisepticum strain S6 genome contains three regions hybridizing with 16 S rRNA and two regions hybridizing with 23 S and 5 S rRNA; Scamrov A et al.; Southern hybridization and cloning experiments revealed existence of 3 regions hybridizing with 16 S rRNA and 2 regions hybridizing with 23 S and 5 S rRNA in Mycoplasma Gallisepticum strain S6 genome thus forming 4 separate contiguous regions . One set of a putative rRNA genes is organized classically for eubacteria order 16 S-23 S-5 S . The other two 16 S rRNA and the other one 23 S-5 S rRNA hybridizing regions are separated from each other and from the complete rRNA operon for a distance of more than 6 kb.

J Bacteriol, 1991 Oct, 173(20), 6321 - 4
Evidence for the establishment of aphid-eubacterium endosymbiosis in an ancestor of four aphid families; Munson MA et al.; Aphids (superfamily Aphidoidea) contain eubacterial endosymbionts localized within specialized cells (mycetocytes) . The endosymbionts are essential for the survival of the aphid hosts . Sequence analyses of the 16S rRNAs from endosymbionts of 11 aphid species from seven tribes and four families have indicated that the endosymbionts are monophyletic . Furthermore, phylogenetic relationships within the symbiont clade parallel the relationships of the corresponding aphid hosts . Our findings suggest that this endocytobiotic association was established in a common ancestor of the four aphid families with subsequent diversification into the present species of aphids and their endosymbionts.

Oral Microbiol Immunol, 1991 Oct, 6(5), 310 - 2
Predominant cultivable flora in pericoronitis; Wade WG et al.; The predominant cultivable flora in pericoronitis was investigated by culturing pus from affected sites in 20 patients . Twenty colonies were picked at random from non-selective plates and identified using conventional biochemical and physiological tests, analysis of metabolic end-products by gas chromatography and protein profile analysis . The most frequently isolated organisms were Prevotella (Bacteroides) intermedia, Peptostreptococcus micros, Veillonella species, Fusobacterium nucleatum and Streptococcus mitis . Porphyromonas (Bacteroides) gingivalis was not isolated and asaccharolytic Eubacterium species were virtually absent . The predominant cultivable microflora in pericoronitis was found to be highly anaerobic in nature and superficially similar to that found in chronic periodontitis, although proposed marker organisms of severe periodontitis were absent.

Kansenshogaku Zasshi, 1991 Oct, 65(10), 1344 - 54
{Bacteriological examination of infections in the field of oral surgery}; Higashitsutsumi M et al.; Etiology of bacterial infections in the field of oral surgery was studied . A total of 270 samples collected from patients with encapsulated abscess in their oral cavities was examined and bacteria were isolated from the 244 samples (90.4%) . The following results were found; 1) Organisms more than one from one sample were frequently isolated from cases with parodontitis, pericoronitis and gnathitis . Isolation of anaerobic bacteria was common (54.2%) . 2) Streptococcus milleri and Streptococcus sanguis and Capnocytophaga species were the most common isolates among aerobic gram-positive and gram-negative bacteria, respectively . 3) Peptostreptococcus micros and Eubacterium lentum were most frequent isolates among gram-negative anaerobic bacteria . Among gram-negative bacteria, Oral Group Bacteroides, especially Bacteroides gingivalis, Bacteroides intermedius, Bacteroides buccae and Bacteroides oralis were most prominent . 4) Isolation frequency of bacteria (both species and strains) was high from samples obtained from patients before antibiotic chemotherapy . 5) Most strains were sensitive to Midecamycin acetate and Josamycin . Minimum inhibitory concentration of 80% isolates (MIC80) against these antibiotics was 0.39 microgram/ml.

FEMS Microbiol Lett, 1991 Oct 1, 67(2), 165 - 8
Direct amplification and sequencing of the 16S ribosomal DNA of an intracellular Legionella species recovered by amoebal enrichment from the sputum of a patient with pneumonia; Fry NK et al.; DNA coding for the 16S rRNA of an intracellular bacterium was directly amplified from lysed cells of a host amoebae using the polymerase chain reaction and primers specific for eubacteria . The amoebae had been used to recover an uncultured bacterium observed in the sputum of a patient with pneumonia . The amplified DNA was sequenced directly and compared with published 16S rRNA sequences . The analysis revealed that the intracellular bacterium is a member of the genus Legionella and that it is different from species, including L . pneumophila, for which 16S ribosomal RNA sequence data are available.

J Gen Microbiol, 1991 Oct, 137 ( Pt 10), 2321 - 9
Positive selection of antibiotic-producing soil isolates; Huck TA et al.; Stepwise discriminant analysis was used to identify the most powerful selective substrates which could be used to formulate media capable of enriching for antibiotic-producing soil isolates . This was achieved by characterizing a collection of 74 soil bacteria, including eubacteria and actinomycetes, according to their ability to produce antibacterial antibiotics and their growth responses to 43 physiological and nutritional tests . The characters which were selective for actinomycetes relative to eubacteria included growth on proline (1%, w/v) and humic acid (0.1%) as sole sources of both carbon and nitrogen, growth on nitrate as a nitrogen source, and growth at pH 7.7-8.0 . Growth on proline (1%) and humic acid (0.1%) as sole carbon/nitrogen sources, growth on asparagine as a nitrogen source, and growth in the presence of vitamins were among the characteristics which allowed antibiotic-producing actinomycetes to be differentiated from non-antibiotic-producing strains . Several simple isolation media which incorporated the selective substrates identified by discriminant analysis succeeded in increasing the proportion of actinomycetes isolated from soil samples . Furthermore, the percentage of isolates capable of antibiotic production was considerably increased.

Appl Environ Microbiol, 1991 Oct, 57(10), 2847 - 57
Biodegradation of dichloromethane and its utilization as a growth substrate under methanogenic conditions; Freedman DL et al.; Biodegradation of dichloromethane (DCM) to environmentally acceptable products was demonstrated under methanogenic conditions (35 degrees C) . When DCM was supplied to enrichment cultures as the sole organic compound at a low enough concentration to avoid inhibition of methanogenesis, the molar ratio of CH4 formed to DCM consumed (0.473) was very close to the amount predicted by stoichiometric conservation of electrons . DCM degradation was also demonstrated when methanogenesis was partially inhibited (with 0.5 to 1.5 mM 2-bromoethanesulfonate or approximately 2 mM DCM) or completely stopped (with 50 to 55.5 mM 2-bromoethanesulfonate) . Addition of a eubacterial inhibitor (vancomycin, 100 mg/liter) greatly reduced the rate of DCM degradation . 14CO2 was the principal product of {14C}DCM degradation, followed by 14CH4 (when methanogenesis was uninhibited) or 14CH3COOH (when methanogenesis was partially or completely inhibited) . Hydrogen accumulated during DCM degradation and then returned to background levels when DCM was consumed . These results suggested that nonmethanogenic organisms mediated DCM degradation, oxidizing a portion to CO2 and fermenting the remainder to acetate; acetate formation suggested involvement of an acetogen . Methanogens in the enrichment culture then converted the products of DCM degradation to CH4 . Aceticlastic methanogens were more easily inhibited by 2-bromoethanesulfonate and DCM than were CO2-reducing methanogens . When DCM was the sole organic-carbon and electron donor source supplied, its use as a growth substrate was demonstrated . The highest observed yield was 0.085 g of suspended organic carbon formed per g of DCM carbon consumed . Approximately 85% of the biomass formed was attributable to the growth of nonmethanogens, and 15% was attributable to methanogens.

FEBS Lett, 1991 Sep 23, 290(1-2), 235 - 8
Folding intermediates of hyperthermophilic D-glyceraldehyde-3-phosphate dehydrogenase from Thermotoga maritima are trapped at low temperature; Schultes V et al.; D-Glyceraldehyde-3-phosphate dehydrogenase from the hyperthermophilic eubacterium, Thermotoga maritima, is extremely thermostable showing a thermal transition beyond 105 degrees C . At low temperature, 'cold denaturation' becomes detectable only in the presence of destabilizing agents . Reconstitution after preceding denaturation depends on temperature . At 0 degree C, no significant recovery of activity is detectable, whereas between 30 and 100 degrees C reactivation reaches up to 85% . Shifting the temperature from low values to the range of optimum reconstitution releases the trapped intermediate in a fast reaction . Evidence from ultra-centrifugal analysis and far-UV circular dichroism proves the intermediate to be partially assembled to the tetramer, with most of its native secondary structure restored in a fast reaction . Fluorescence emission exhibits at least biphasic kinetics with the rate-limiting step(s) reflecting local adjustments of aromatic residues involved in tertiary contacts in the native state of the enzyme.

J Mol Biol, 1991 Sep 20, 221(2), 387 - 401
Novel anticodon composition of transfer RNAs in Micrococcus luteus, a bacterium with a high genomic G + C content . Correlation with codon usage; Kano A et al.; The number and relative amount of isoacceptor tRNAs for each amino acid in Micrococcus luteus, a Gram-positive bacterium with high genomic G + C content, have been determined by sequencing their anticodon loop and its adjacent regions and by selective labelling of tRNAs . Thirty-one tRNA species with 29 different anticodon sequences have been detected . All the tRNAs have G or C at the anticodon first position except for tRNA(ICGArg) and tRNA(NGASer), in response to the abundant usage of NNC and NNG codons . No tRNA with the anticodon UNN capable of translating codon NNA has been detected, in accordance with a very low or zero usage of NNA codons . The relative amount of isoacceptor tRNAs for an amino acid determined by selective labelling strongly correlates with usage of the corresponding codons . On the basis of these and other observations in this and other eubacterial species, we conclude that the relative amount and anticodon composition of isoacceptor tRNA species are flexible, and their changes are mainly adaptive phenomena that have been primarily affected by codon usage, which in turn is affected by directional mutation pressure.

J Steroid Biochem Mol Biol, 1991 Sep, 39(3), 367 - 74
Properties of a 4-ene-3-ketosteroid-5 alpha-reductase in cell extracts of the intestinal anaerobe, Eubacterium sp . strain 144; Glass TL et al.; When Eubacterium sp . 144 was grown in the presence of progesterone, extracts of these cells contained a 4-ene-3-ketosteroid-5 alpha-reductase (5 alpha-reductase) . No evidence for the presence of a 5 beta-steroid-reductase or a 5 alpha to 5 beta-steroid-isomerase was found . 5 alpha-Reductase activity was dependent on reduced methyl viologen as the electron donor and this could be generated biologically by adding pyruvate or H2 to cell extracts or chemically by adding sodium dithionite . NADH or NADPH with or without flavin nucleotides were not electron donors for 5 alpha-reductase . Most of the 5 alpha-reductase activity (60-65%) of crude extracts was located in the membrane fraction and the enzyme was solubilized by treatment with 1% Triton X-100 . Optimum 5 alpha-reductase activity occurred at pH 7.0-7.5 in potassium phosphate buffer but was stimulated by Tris-HCl buffer (pH 8.0-9.0) . 5 alpha-Reductase activity was highest at 10% (v/v) methanol and was progressively inhibited by higher methanol concentrations . Sulfhydryl reagents strongly inhibited 5 alpha-reductase but the enzyme was not affected by other metabolic inhibitors . Extracts prepared from cells induced with 16-dehydroprogesterone and grown without hemin contained 5 alpha-reductase and 16-dehydroprogesterone-reductase activities equivalent to those found in extracts of induced cells grown with hemin . This indicates that hemin is not required for the synthesis of active steroid double bond-reductases in strain 144.

J Bacteriol, 1991 Sep, 173(18), 5837 - 42
Structural features of methyl-accepting taxis proteins conserved between archaebacteria and eubacteria revealed by antigenic cross-reaction; Alam M et al.; A number of eubacterial species contain methyl-accepting taxis proteins that are antigenically and thus structurally related to the well-characterized methyl-accepting chemotaxis proteins of Escherichia coli . Recent studies of the archaebacterium Halobacterium halobium have characterized methyl-accepting taxis proteins that in some ways resemble and in other ways differ from the analogous eubacterial proteins . We used immunoblotting with antisera raised to E . coli transducers to probe shared structural features of methyl-accepting proteins from archaebacteria and eubacteria and found substantial antigenic relationships . This implies that the genes for the contemporary methyl-accepting proteins are related through an ancestral gene that existed before the divergence of arachaebacteria and eubacteria . Analysis by immunoblot of mutants of H . halobium defective in taxis revealed that some strains were deficient in covalent modification of methyl-accepting proteins although the proteins themselves were present, while other strains appeared to be missing specific methyl-accepting proteins.

J Periodontal Res, 1991 Sep, 26(5), 409 - 14
Bacteriological study of juvenile periodontitis in China; Han NM et al.; The predominant cultivable bacteria associated with juvenile periodontitis (JP) in China were studied for the first time . Subgingival plaque samples were taken on paper points from 23 diseased sites in 15 JP patients and from 7 healthy sites in 7 control subjects . Serially diluted plaque samples were plated on nonselective blood agar and on MGB agar, a selective medium for the isolation of Actinobacillus actinomycetemcomitans . Fifteen or more isolated colonies from each sample (in sequence without selection) were purified for identification . The results indicated that the microflora in healthy sulci of the 7 control subjects was significantly different from that in diseased sites of JP patients . The predominant species in healthy sulci were Streptococcus spp . and Capnocytophaga gingivalis . In JP patients, Eubacterium sp . was found in significantly higher frequency and proportion . Actinobacillus actinomycetemcomitans was not detected in any samples . It appears that this species is not associated with juvenile periodontitis in China.

Res Microbiol, 1991 Sep-Oct, 142(7-8), 851 - 9
Initiation of chromosome replication: structure and function of oriC and DnaA protein in eubacteria; Ogasawara N et al.; Recent advances in DNA technology have made it possible to analyse the structure and function of the replication origin region of the chromosomes of various bacteria . Comparative studies have shown that 2 basic elements, the replicator and initiator, involved in initiation of chromosome replication are common to most eubacteria but with differences in the fine organization of these elements . In this article, we first review studies of the structural analysis of the origin regions of bacterial chromosomes, and then we summarize our recent work on the function of the 2 elements in Bacillus subtilis as compared to Escherichia coli, in order to show how organization of the elements is related to the differences in regulation of the initiation of replication in the 2 bacteria . Remarkable conservation of genes and their organization in the replication origin region was found in 5 bacteria representative of 3 major branches of the bacterial phylogenic tree . It was concluded that the conserved region containing the dnaA gene is the replication origin of the ancestral bacterium . Conservation of DnaA protein and its binding sequence (DnaA box) is remarkable, suggesting that they are the initiator and replicator of the chromosomes of most eubacteria . We have recently isolated an autonomously replicating sequence (ars) from B . subtilis . The essential features of ars, the presence of DnaA boxes and repeats of an AT rich 15-mer, are the same as E . coli oriC . However, 2 DnaA-box regions flanking the dnaA gene are both required for B . subtilis ars . The function of DnaA protein in vivo was studied in detail using a temperature-sensitive dnaA mutant in B . subtilis.(ABSTRACT TRUNCATED AT 250 WORDS)

Appl Environ Microbiol, 1991 Sep, 57(9), 2683 - 6
Light sensitivity of methanogenic archaebacteria; Olson KD et al.; Representatives of four families of methanogenic archaebacteria (archaea), Methanobacterium thermoautotrophicum delta H, Methanobacterium thermoautotrophicum Marburg, Methanosarcina acetivorans, Methanococcus voltae, and Methanomicrobium mobile, were found to be light sensitive . The facultative anaerobic eubacteria Escherichia coli and Salmonella typhimurium, however, were tolerant of light when grown anaerobically under identical light conditions . Interference filters were used to show that growth of the methanogens is inhibited by light in the blue end of the visible spectrum (370 to 430 nm).

J Periodontol, 1991 Sep, 62(9), 576 - 85
Clinical documentation and occurrence of putative periodontopathic bacteria in human immunodeficiency virus-associated periodontal disease; Gornitsky M et al.; Human immunodeficiency virus (HIV)-associated gingivitis (HIV-G) and HIV-associated periodontitis (HIV-P) are two intraoral lesions manifested by patients with HIV infection . Periodontal indices were measured for 87 subjects in 5 study groups: HIV-seropositive patients with healthy periodontium (HIV-H), with HIV-G, or with HIV-P; and non-HIV-infected subjects with healthy periodontium (H) or with adult chronic periodontitis (P) . The quantitative clinical parameters were compared and statistically significant intergroup differences were noted . The mean scores on PI and PD do not discriminate between HIV-seropositive and non-HIV-infected seronegative cohorts, but a significant difference in the GI between HIV-H and H was noted . When categories of PD and AL are examined, some differences become apparent . Generally, the PD and AL of HIV-P are not as great as those of P . PI correlates well with GI (r = 0.86) in P, but does not (r = 0.33) in HIV-P . In addition, the occurrence of selected putative periodontopathic bacteria (Porphyromonas gingivalis, spirochetes, and motile eubacteria) in these lesions was determined by brightfield (after staining), darkfield and immunofluorescent microscopy . No difference in microbiological profile in the bacterial groups monitored was found between P and HIV-P . Spirochetes were found to be more abundant than P . gingivalis in the lesions of P and HIV-P . In marked contrast, P . gingivalis was found to be in highest numbers in samples from the gingival crevice of H as determined by indirect immunofluorescence.

J Clin Invest, 1991 Sep, 88(3), 750 - 4
Penicillin-binding protein inactivation by human neutrophil myeloperoxidase; Rakita RM et al.; Myeloperoxidase (MPO), H2O2, and chloride comprise a potent antimicrobial system believed to contribute to the antimicrobial functions of neutrophils and monocytes . The mechanisms of microbicidal action are complex and not fully defined . This report describes the MPO-mediated inactivation, in Escherichia coli, Staphylococcus aureus, and Pseudomonas aeruginosa, of a class of cytoplasmic membrane enzymes (penicillin-binding proteins, PBPs) found in all eubacteria, that covalently bind beta-lactam antibiotics to their active sites with loss of enzymatic activity . Inactivation of "essential" PBPs, including PBP1-PBP3 of E . coli, leads to unbalanced bacterial growth and cell death . MPO treatment of bacteria was associated with loss of penicillin binding by PBPs, strongly suggesting PBP inactivation . In E . coli, PBP inactivation was most rapid with PBP3, where the rate of decline in binding activity approximated but did not equal loss of viability . Changes in E . coli morphology (elongation), observed just before bacteriolysis, were consistent with early predominant inactivation of PBP3 . We conclude that inactivation of essential PBPs is sufficient to account for an important fraction of MPO-mediated bacterial action . This feature of MPO action interestingly recapitulates an antibacterial strategy evolved by beta-lactam-producing molds that must compete with bacteria for limited ecologic niches.

FEBS Lett, 1991 Aug 19, 288(1-2), 72 - 6
Expression and functional assembly into bacterial ribosomes of a nuclear-encoded chloroplast ribosomal protein with a long NH2-terminal extension; Giese K et al.; Chloroplast ribosomal protein L13 is encoded in the plant nucleus and is considerably larger than its eubacterial homologue by having NH2- and COOH-terminal extensions with no homology to any known sequences (Phua et al., J Biol . Chem . 264, 1968-1971, 1989) . We made two gene constructs of L13 cDNA using the polymerase chain reaction (PCR) and expressed them in Escherichia coli . Analysis of the ribosomes and polysomes from these cells, using an antiserum specific to chloroplast L13, shows that the expressed proteins are incorporated, in the presence of the homologous E . coli L13, into functional ribosomes which participate in protein synthesis (i.e . polysomes) . Evidence is obtained that the large NH2-terminal extension probably lies on the surface of these 'mosaic ribosomes.' This first report of the assembly into E . coli ribosomes of nuclear-coded chloroplast ribosomal protein with terminal extensions thus suggest an extraordinary conservation in the function of eubacterial type ribosomal proteins, despite the many changes in protein structure during their evolution inside a eukaryotic system.

J Biol Chem, 1991 Aug 5, 266(22), 14151 - 4
Unique characteristics of superoxide dismutase of a strictly anaerobic archaebacterium Methanobacterium thermoautotrophicum; Takao M et al.; The superoxide dismutase (SOD) gene of Methanobacterium thermoautotrophicum (Takao, M., Oikawa, A., and Yasui, A . (1990) Arch . Biochem . Biophys . 283, 210-216), a strictly anaerobic archaebacterium, was expressed in Escherichia coli . The gene product accounted for more than 30% of the host's soluble protein . The purified protein was an active iron-containing tetrameric SOD with specific activity similar to known manganese-containing SODs (MnSODs) of aerobic archaebacteria . Although M . thermoautotrophicum SOD is an iron-containing SOD (FeSOD), it resembles MnSODs in amino acid sequence as judged by criteria distinguishing FeSODs from MnSODs . Moreover, M . thermoautotrophicum SOD is resistant to azide and hydrogen peroxide as MnSODs are, suggesting that its evolution is distinct from known eubacterial FeSODs.

Protein Seq Data Anal, 1991 Aug, 4(2), 75 - 9
Comparison of the structure of archaebacterial ribosomal proteins equivalent to proteins L11 and L1 from Escherichia coli ribosomes; Ramirez C et al.; The sequences of two ribosomal proteins from two widely divergent species of archaebacteria, Halobacterium cutirubrum and Sulfolobus solfataricus, have been deduced from the structure of their respective genes . These two proteins were found to be equivalent to the L11 and L1 ribosomal proteins of the eubacterium Escherichia coli . Sequence comparison revealed that the archaebacterial L11e (equivalent to E . coli L11) proteins are longer than the eubacterial protein due to a C-terminal extension of about 30 residues . The archaebacterial L11e proteins, like the E . coli L11, are rich in proline residues; most of these are conserved . L11 is the most highly methylated protein in the E . coli ribosome . However, sites of methylation are generally not conserved in the archaebacterial L11e proteins . The region of highest sequence similarity between L11 and the archaebacterial L11e proteins is the N-terminal domain . This domain is believed to interact with release factor 1 during termination of translation . The amino acid sequences of the archaebacterial L1e proteins were compared to the eubacterial E . coli L1 and Bacillus stearothermophilus L1e sequences . The archaebacterial L1e proteins are slightly shorter at both their N- and C-termini . A region of high sequence similarity (7 of 14 residues) occurs near the center of the proteins.

J Bacteriol, 1991 Aug, 173(15), 4558 - 69
Cloning, sequencing, and expression of the gene coding for bile acid 7 alpha-hydroxysteroid dehydrogenase from Eubacterium sp . strain VPI 12708; Baron SF et al.; Southern blot analysis indicated that the gene encoding the constitutive, NADP-linked bile acid 7 alpha-hydroxysteroid dehydrogenase of Eubacterium sp . strain VPI 12708 was located on a 6.5-kb EcoRI fragment of the chromosomal DNA . This fragment was cloned into bacteriophage lambda gt11, and a 2.9-kb piece of this insert was subcloned into pUC19, yielding the recombinant plasmid pBH51 . DNA sequence analysis of the 7 alpha-hydroxysteroid dehydrogenase gene in pBH51 revealed a 798-bp open reading frame, coding for a protein with a calculated molecular weight of 28,500 . A putative promoter sequence and ribosome binding site were identified . The 7 alpha-hydroxysteroid dehydrogenase mRNA transcript in Eubacterium sp . strain VPI 12708 was about 0.94 kb in length, suggesting that it is monocistronic . An Escherichia coli DH5 alpha transformant harboring pBH51 had approximately 30-fold greater levels of 7 alpha-hydroxysteroid dehydrogenase mRNA, immunoreactive protein, and specific activity than Eubacterium sp . strain VPI 12708 . The 7 alpha-hydroxysteroid dehydrogenase purified from the pBH51 transformant was similar in subunit molecular weight, specific activity, and kinetic properties to that from Eubacterium sp . strain VPI 12708, and it reached with antiserum raised against the authentic enzyme on Western immunoblots . Alignment of the amino acid sequence of the 7 alpha-hydroxysteroid dehydrogenase with those of 10 other pyridine nucleotide-linked alcohol/polyol dehydrogenases revealed six conserved amino acid residues in the N-terminal regions thought to function in coenzyme binding.

Mol Gen Genet, 1991 Aug, 228(1-2), 70 - 80
Organization and nucleotide sequence of ten ribosomal protein genes from the region equivalent to the spectinomycin operon in the archaebacterium Halobacterium marismortui; Scholzen T et al.; The nucleotide sequence has been determined of a 4700 bp region from a ribosomal protein gene cluster of Halobacterium marismortui (Haloarcula marismortui), which is equivalent to part of the spectinomycin operon of Escherichia coli . The genes were localized on the recombinant lambda EMBL3 clone PP*7, which also contains several other ribosomal protein genes from the DNA region in H . marismortui equivalent to the linked S10/spc operon . The genes analysed encode ten ribosomal proteins, namely HmaL5, HmaS14, HmaS8, HmaL6, HL5, HL24, HmaL18, HmaS5, HmaL30 and HmaL15 . The gene organization of the archaebacterial cluster is similar to that in eubacteria but has two additional genes, namely those encoding HL5 and HL24, which were identified as extra proteins that are apparently not present in E . coli . These correspond to the gene products of orfd and orfe in Methanococcus vannielii and also have eukaryotic counterparts.

Mol Cell Biol, 1991 Aug, 11(8), 3949 - 59
Binding sites of the 9- and 14-kilodalton heterodimeric protein subunit of the signal recognition particle (SRP) are contained exclusively in the Alu domain of SRP RNA and contain a sequence motif that is conserved in evolution; Strub K et al.; The mammalian signal recognition particle (SRP) is a small cytoplasmic ribonucleoprotein required for the cotranslational targeting of secretory proteins to the endoplasmic reticulum membrane . The heterodimeric protein subunit SRP9/14 was previously shown to be essential for SRP to cause pausing in the elongation of secretory protein translation . RNase protection and filter binding experiments have shown that binding of SRP9/14 to SRP RNA depends solely on sequences located in a domain of SRP RNA that is strongly homologous to the Alu family of repetitive DNA sequences . In addition, the use of hydroxyl radicals, as RNA-cleaving reagents, has revealed four distinct regions in this domain that are in close contact with SRP9/14 . Surprisingly, the nucleotide sequence in one of these contact sites, predicted to be mostly single stranded, was found to be extremely conserved in SRP RNAs of evolutionarily distant organisms ranging from eubacteria and archaebacteria to yeasts and higher eucaryotic cells . This finding suggests that SRP9/14 homologs may also exist in these organisms, where they possibly contribute to the regulation of protein synthesis similar to that observed for mammalian SRP in vitro.

Gene, 1991 Jul 22, 103(2), 139 - 45
Conservation and evolution of the nucleus-encoded and chloroplast-specific ribosomal proteins in pea and spinach; Carol P et al.; Two cDNA clones have been isolated from a lambda g11 cDNA library constructed with poly(A)+ mRNAs prepared from spinach seedlings . These nuclear cDNAs encode chloroplast (cp) ribosomal (r) proteins designated L24 and L40 . These r-proteins have been identified in the cp 50S r-subunit by immunoblot analysis, amino acid (aa) composition and N-terminal aa sequencing . The L24 r-protein contains a central eubacterial homologous core with the N- and C-terminal polypeptide extensions . The L40 r-protein has no homologous counterpart in bacterial ribosomes . The two nuclear encoded r-proteins have their homologues in pea, a legume, showing that specific elements of cp ribosomes are conserved in higher plants . Surprisingly, the cp-specific r-protein L40 has a higher aa substitution rate than that of other eubacterial-like cp r-proteins identified previously in pea and spinach.

Proc Natl Acad Sci U S A, 1991 Jul 15, 88(14), 6308 - 12
Detection of a key tertiary interaction in the highly conserved GTPase center of large subunit ribosomal RNA; Ryan PC et al.; Searches of ribosomal RNA sequences for compensatory base changes preserving Watson-Crick base pairing have led to detailed models of the conserved secondary structures of these RNAs . In principle, tertiary interactions can also be detected by searches for phylogenetically covariant bases . Within a highly conserved region of the large subunit ribosomal RNA termed the "GTPase center," the bases G-1056-U-1082.A-1086 are found in all eubacteria (Escherichia coli numbering), while A-1056.C-1082.G-1086 are found at the homologous positions in eukaryotes; archaebacteria fall into either category with some exceptions . Either sequence can potentially form a similar set of hydrogen bonds connecting the 3 bases . To determine the contribution of these 3 bases to RNA tertiary structure, sequence variants were made in RNA fragments covering the GTPase center . Correct folding of the RNA fragments was assayed by measuring the binding affinities of two different ligands that recognize the RNA tertiary structure: the highly conserved ribosomal protein L11, which is normally associated with the GTPase center RNA, and the peptide antibiotic thiostrepton, which inhibits the GTPase activity of eubacterial and some archaebacterial ribosomes . The results strongly support the existence of a base pair between positions 1082 and 1086: single mutations at either position weaken both L11 and thiostrepton binding by approximately 10-fold or more, while compensatory double mutations bind the ligands nearly as well as the wild-type E . coli sequence . Variants at position 1056 have little effect on either L11 or thiostrepton binding; a 3-base interaction is therefore not supported by these experiments . A base pair between positions 1082 and 1086 strongly constrains the geometry with which three helical segments join in the middle of the GTPase center.

J Bacteriol, 1991 Jul, 173(14), 4371 - 8
Analysis of a marine picoplankton community by 16S rRNA gene cloning and sequencing; Schmidt TM et al.; The phylogenetic diversity of an oligotrophic marine picoplankton community was examined by analyzing the sequences of cloned ribosomal genes . This strategy does not rely on cultivation of the resident microorganisms . Bulk genomic DNA was isolated from picoplankton collected in the north central Pacific Ocean by tangential flow filtration . The mixed-population DNA was fragmented, size fractionated, and cloned into bacteriophage lambda . Thirty-eight clones containing 16S rRNA genes were identified in a screen of 3.2 x 10(4) recombinant phage, and portions of the rRNA gene were amplified by polymerase chain reaction and sequenced . The resulting sequences were used to establish the identities of the picoplankton by comparison with an established data base of rRNA sequences . Fifteen unique eubacterial sequences were obtained, including four from cyanobacteria and eleven from proteobacteria . A single eucaryote related to dinoflagellates was identified; no archaebacterial sequences were detected . The cyanobacterial sequences are all closely related to sequences from cultivated marine Synechococcus strains and with cyanobacterial sequences obtained from the Atlantic Ocean (Sargasso Sea) . Several sequences were related to common marine isolates of the gamma subdivision of proteobacteria . In addition to sequences closely related to those of described bacteria, sequences were obtained from two phylogenetic groups of organisms that are not closely related to any known rRNA sequences from cultivated organisms . Both of these novel phylogenetic clusters are proteobacteria, one group within the alpha subdivision and the other distinct from known proteobacterial subdivisions . The rRNA sequences of the alpha-related group are nearly identical to those of some Sargasso Sea picoplankton, suggesting a global distribution of these organisms.

Eur J Biochem, 1991 Jul 1, 199(1), 89 - 94
Purification and properties of an iron-sulfur-containing and pyridoxal-phosphate-independent L-serine dehydratase from Peptostreptococcus asaccharolyticus; Grabowski R et al.; L-Serine dehydratase with a specific activity of 15 nkat/mg protein was present in the anaerobic eubacterium Peptostreptococcus asaccharolyticus grown either on L-glutamate or L-serine . The enzyme was highly specific for L-serine with the lowest Km = 0.8 mM ever reported for an L-serine dehydratase . L-Threonine (Km = 22 mM) was the only other substrate . V/Km for L-serine was 500 times higher than that for L-threonine . L-Cysteine was the best inhibitor (Ki = 0.3 mM, competitive towards L-serine) . The enzyme was purified 400-fold to homogeneity under anaerobic conditions (specific activity 6 mukat/mg) . PAGE in the presence of SDS revealed two subunits with similar intensities (alpha, 30 kDa; beta, 25 kDa) . The molecular mass of the native enzyme was estimated as 200 +/- 20 kDa (gel filtration) and 180 kDa (gradient PAGE) . In the absence of oxygen the enzyme was moderately stable even in the presence of sodium borohydride or phenylhydrazine (5 mM each) . However, by exposure to air the activity was lost, especially when the latter agent was added . The enzyme was reactivated by ferrous ion under anaerobic conditions . The inability of several nucleophilic agents to inactivate the enzyme indicated the absence of pyridoxal phosphate . This was confirmed by a microbiological determination of pyridoxal phosphate . However, the enzyme contained 3.8 +/- 0.2 mol Fe and 5.6 +/- 0.3 mol inorganic sulfur/mol heterodimer (55 kDa) indicating the presence of an {Fe-S} center . The enzyme was successfully applied to measure L-serine concentrations in bacterial media and in human sera.

J Bacteriol, 1991 Jul, 173(13), 4234 - 6
Novel tRNA gene organization in the 16S-23S intergenic spacer of the Streptococcus pneumoniae rRNA gene cluster; Bacot CM et al.; Isoleucine and alanine tRNAs are encoded tandemly within the 16S-23S intergenic spacer of some eubacterial rRNA gene clusters . Southern hybridization analysis and DNA sequence analysis demonstrated a novel gene organization for an rRNA gene cluster on the Streptococcus pneumoniae chromosome . A sequence specifying an alanine tRNA was found within the intergenic spacer, but no sequence specifying an isoleucine tRNA was found there . Southern hybridization analysis indicated that the location of the isoleucine tRNA gene was near the 5S rRNA gene in two of four rRNA gene clusters.

Infect Immun, 1991 Jul, 59(7), 2502 - 4
Somnogenic activity of pseudomurein in rabbits; Johannsen L et al.; Pseudomurein is a major cell wall component of some archaebacteria that chemically differs from but morphologically, functionally, and structurally resembles eubacterial peptidoglycan . Eubacterial cell wall components, e.g., peptidoglycan, induce changes in sleep and body temperature . We now report that intravenous injections of rabbits with a suspension of pseudomurein from Methanobacterium thermoautotrophicum also induce similar central nervous system effects.

J Clin Periodontol, 1991 Jul, 18(6), 441 - 6
Dental bacterial plaques . Nature and role in periodontal disease; Christersson LA et al.; Antony van Leeuwenhoek first described oral bacteria . However, not until almost 200 years later was the famous Koch postulate introduced . Since then, research has extensively been performed regarding the development and microbiology of dental plaques . In spite of the complexity of the developing flora of supragingival plaque, culture studies have shown a remarkably orderly succession of organisms . Lately, the concept of microbial specificity in the etiology of periodontal diseases has been widely suggested, i.e., that different forms of periodontal disease are associated with qualitatively distinct dental plaques . Cross-sectional and longitudinal studies of the predominant cultivable microflora reveal that only a small number of the over 300 species found in human subgingival plaques are associated with periodontal disease . Among the commonly mentioned are: Actinobacillus actinomycetemcomitans, Bacteroides gingivalis, Bacteroides intermedius, Capnocytophaga sp., Eikenella corrodens, Fusobacterium nucleatum, and Wolinella recta, as well as certain gram-positive bacteria such as Eubacterium species . Anti-infective therapy for many systemic infections equals the use of antimicrobial drugs . However, for localized infections like periodontal diseases, treatment may consist of a combination of mechanical wound debridement and the application of an antimicrobial agent . The general effectiveness of mechanical anti-infective therapy and successful oral hygiene in the management of periodontal disease is well established in the literature and has met the test of success in clinical practice for most cases of periodontitis in adults . The definition of periodontal pathogens as either opportunistic pathogens, or as exogenous pathogens carries with it significant implications.(ABSTRACT TRUNCATED AT 250 WORDS)

Biochimie, 1991 Jul-Aug, 73(7-8), 1067 - 88
The function of the translating ribosome: allosteric three-site model of elongation; Rheinberger HJ; During the last decade, a new model for the ribosomal elongation cycle has emerged . It is based on the finding that eubacterial ribosomes possess 3 tRNA binding sites . More recently, this has been confirmed for archaebacterial and eukaryotic ribosomes as well, and thus appears to be a universal feature of the protein synthetic machinery . Ribosomes from organisms of all 3 kingdoms harbor, in addition to the classical P and A sites, an E site (E for exit), into which deacylated tRNA is displaced during translocation, and from which it is expelled by the binding of an aminoacyl-tRNA to the A site at the beginning of the subsequent elongation round . The main features of the allosteric 3-site model of ribosomal elongation are the following: first, the third tRNA binding site is located 'upstream' adjacent to the P site with respect to the messenger, ie on the 5'-side of the P site . Second, during translocation, deacylated tRNA does not leave the ribosome from the P site, but co-translocates from the P site to the E site--when peptidyl-tRNA translocates from the A site to the P site . Third, deacylated tRNA is tightly bound to the E site in the post-translocational state, where it undergoes codon--anticodon interaction . Fourth, the elongating ribosome oscillates between 2 main conformations: (i), the pre-translocational conformer, where aminoacyl-tRNA (or peptidyl-tRNA) and peptidyl-tRNA (or deacylated tRNA) are firmly bound to the A and P sites, respectively; and (ii), the post-translocational conformer, where peptidyl-tRNA and deacylated tRNA are firmly bound to the P and E sites, respectively . The transition between the 2 states is regulated in an allosteric manner via negative cooperatively . It is modulated in a symmetrical fashion by the 2 elongation factors Tu and G . An elongating ribosome always maintains 2 high-affinity tRNA binding sites with 2 adjacent codon--anticodon interactions . The allosteric transition from the post- to the pre-translocational state is involved in the accuracy of aminoacyl-tRNA selection, and the maintenance of 2 codon--anticodon interactions helps to keep the messenger in frame during translation.

J Gen Microbiol, 1991 Jul, 137 ( Pt 7), 1523 - 8
Cloning, characterization and taxonomic significance of genes for the 5S ribosomal RNA of Leptonema illini strain 3055; Fukunaga M et al.; The genes encoding the 5S ribosomal RNA (rRNA) for Leptonema illini strain 3055 were isolated and sequenced . The 5S RNA molecule encoded was 117 nucleotides long . The genome of strain 3055 contained two genes for 5S rRNA that were located close together . The nucleotide sequences of the Leptonema illini genes exhibited less similarity to the rRNA gene of Leptospira interrogans strain Moulton and also to those of typical eubacterial genes than did the rRNA genes of other leptospires . However, the overall secondary structure of the 5S rRNA encoded exhibited a strong similarity to that of typical eubacterial 5S rRNA . Southern hybridization of the 5S rRNA gene probe with the genomic DNA of strain 965, which is currently classified as Leptospira biflexa, showed the latter to have close similarity to that of strain 3055 . The physical map of strain 965 was quite similar to that of strain 3055 and was greatly different from that of any other strains of L . biflexa . In the organization of 5S rRNA genes, strain 965 is sufficiently different from other members of the genus Leptospira to be regarded as a member of the genus Leptonema.

Biokhimiia, 1991 Jul, 56(7), 1155 - 89
{Immunochemical study of tryptophanyl-tRNA-synthetase}; Beresten' SF et al.; The immunochemical approach used extensively in medicine and in some fields of biology has not yet been systematically applied to enzymology, in particular, to the study of a large, functionally significant group of enzymes, such as aminoacyl-tRNA synthetases (EC 6.1.1) . The present investigation was aimed at the analysis of applicability of polyclonal and monoclonal antibodies against bovine tryptophanyl-tRNA synthetase in the study of functional properties of this enzyme as well as of its distribution inside the cell and among organs and tissues of various animals . The general conclusions one may draw from these data are as follows . i) Tryptophanyl-tRNA synthetase of eukaryotes, eubacteria and archaebacteria share one common structural element (antigenic determinant) that is not essential for the catalytic activity . The evolutionary conservative nature of this element suggests that the enzyme may implement functions other than catalysis of tryptophanyl-tRNA formation . ii) Tryptophanyl-tRNA synthetase shows an anomalous distribution among mammalian organs: its content is far greater in the exocrine part of the ruminant animal pancreas in comparison with their other organs (liver) or with other mammalian orders . This finding suggests that the enzyme or its fragments may play a role in the digestive function of ruminant animals . iii) Tryptophanyl-tRNA synthetase was found in considerable quantities in diffuse chromatin of mammalian cell nuclei . This fact indicates that the enzyme may participate in such processes in the nucleus as transcription, processing, transport, etc . It may thus be concluded that tryptophanyl-tRNA synthetase of higher organisms, besides catalyzing the formation of aminoacyl-tRNAs can exert some other, yet unknown, noncanonical functions.

Mol Microbiol, 1991 Jul, 5(7), 1687 - 93
A gene in the archaebacterium Sulfolobus solfataricus that codes for a protein equivalent to the alpha subunits of the signal recognition particle receptor in eukaryotes; Ramirez C et al.; We have sequenced a gene in the archaebacterium Sulfolobus solfataricus that codes for a protein that shows sequence similarity to the alpha subunit of the signal recognition particle receptor or docking protein in eukaryotes and the product of the ftsY gene in Escherichia coli . Comparison of the Sulfolobus 'docking protein' with its eukaryotic and eubacterial counterparts showed that the region of highest sequence similarity corresponds to a GTP-binding site . The presence of this gene in archaebacteria suggests that some of the components involved in protein transport have been conserved in the three kingdoms.

Biol Chem Hoppe Seyler, 1991 Jul, 372(7), 513 - 7
Microbial metabolism of quinoline and related compounds . X . The molybdopterin cofactors of quinoline oxidoreductases from Pseudomonas putida 86 and Rhodococcus spec . B1 and of xanthine dehydrogenase from Pseudomonas putida 86; Hettrich D et al.; The bis(carboxamidomethyl) derivatives of the molybdenum cofactors in three eubacterial molybdo-iron/sulphur-flavoproteins were examined . The quinoline oxidoreductases from Pseudomonas putida 86 and Rhodococcus spec . B1 contain molybdopterin cytosine dinucleotide . In xanthine dehydrogenase from Pseudomonas putida 86, however, only molybdopterin was found . The bis(carboxamidomethyl) derivatives of all three enzymes were treated with nucleotide pyrophosphatase, but only those of the quinoline oxidoreductases were cleaved into {bis(carboxamidomethyl)}molybdopterin and CMP, whereas that of xanthine dehydrogenase remained unchanged . Dephosphorylation by alkaline phosphatase yielded dephospho-{bis(carboxamidomethyl)}molybdopterin and cytidine from the cleaved molybdopterin cytosine dinucleotide . The bis(carboxamidomethyl) derivative from xanthine dehydrogenase was converted to dephospho-{bis(carboxamidomethyl)}molybdopterin by alkaline phosphatase . Acid hydrolysis of the purified enzymes and analysis of the hydrolysate by HPLC confirmed that compared with the xanthine dehydrogenase both quinoline oxidoreductases contain CMP.

Gene, 1991 Jun 30, 102(2), 283 - 8
Isolation and analysis of a human cDNA highly homologous to the yeast gene encoding L17A ribosomal protein; Berchtold MW et al.; A cDNA from human brain poly(A)+RNA with significant similarity to the gene encoding yeast L17A large subunit ribosomal (r) protein (L17A) was isolated using the polymerase chain reaction . The deduced amino acid (aa) sequence of 140 aa (calculated pI of 10.79) exhibits a 78% similarity to that of the yeast L17A r protein (88% when conservative aa replacements are considered as well) . This indicates that L17A is one of the best conserved r-proteins and therefore may play a critical role in ribosome function . In contrast to its eubacterial and chloroplast counterparts, human L17A contains an N-terminal extension of 19 aa which may be involved in nuclear targeting of the r-protein . Approximately five to seven genes in mammalian genomes give strong hybridization signals when probed with the human L17A homologue cDNA . Whereas the L17A homologue was found to be expressed at similar levels in several human tissues as a transcript of 600 nucleotides, a several-fold higher transcript level was detected in the rapidly growing neuroblastoma cell line, SK-N-BE.

FEBS Lett, 1991 Jun 17, 284(1), 51 - 6
Extended N-terminal sequencing of proteins of the large ribosomal subunit from yeast mitochondria; Grohmann L et al.; We have determined the N-termini of 26 proteins of the large ribosomal subunit from yeast mitochondria by direct amino acid micro-sequencing . The N-terminal sequences of proteins YmL33 and YmL38 showed a significant similarity to eubacterial ribosomal (r-) proteins L30 and L14, respectively . In addition, several proteins could be assigned to their corresponding yeast nuclear genes . Based on a comparison of the protein sequences deduced from the corresponding DNA regions with the N-termini of the mature proteins, the putative leader peptides responsible for mitochondrial matrix-targeting were compiled . In most leader sequences a relative abundance of aromatic amino acids, preferentially phenylalanine, was found.

Genes Dev, 1991 Jun, 5(6), 1032 - 41
The phage T4 nrdB intron: a deletion mutant of a version found in the wild; Eddy SR et al.; Bacteriophage T4 possesses three self-splicing group I introns . Two of the three introns are mobile elements; the third, in the gene encoding a subunit of the phage nucleotide reductase (nrdB), is not mobile . Because intron mobility offers a reasonable explanation for the paradoxical occurrence of large intervening sequences in a space-efficient eubacterial phage, it is puzzling that the nrdB intron is not mobile like its compatriots . We have discovered a larger nrdB intron in a closely related phage, and we infer from comparative sequence data that the T4 intron is a deletion mutant derived from this larger intron . This larger nrdB intron encodes an open reading frame of 269 codons, which we have cloned and overexpressed . The overexpressed protein shows a dsDNA endonuclease activity specific for the intronless nrdB gene, typical of mobile introns . Thus, we believe that all three introns of T4 are or were mobile "infectious introns" and that they have entered into and been maintained in the phage population by virtue of this efficient mobility.

J Bacteriol, 1991 Jun, 173(11), 3446 - 55
Evolutionary relationships among eubacteria, cyanobacteria, and chloroplasts: evidence from the rpoC1 gene of Anabaena sp . strain PCC 7120; Bergsland KJ et al.; RNA polymerases of cyanobacteria contain a novel core subunit, gamma, which is absent from the RNA polymerases of other eubacteria . The genes encoding the three largest subunits of RNA polymerase, including gamma, have been isolated from the cyanobacterium Anabaena sp . strain PCC 7120 . The genes are linked in the order rpoB, rpoC1, rpoC2 and encode the beta, gamma, and beta' subunits, respectively . These genes are analogous to the rpoBC operon of Escherichia coli, but the functions of rpoC have been split in Anabaena between two genes, rpoC1 and rpoC2 . The DNA sequence of the rpoC1 gene was determined and shows that the gamma subunit corresponds to the amino-terminal half of the E . coli beta' subunit . The gamma protein contains several conserved domains found in the largest subunits of all bacterial and eukaryotic RNA polymerases, including a potential zinc finger motif . The spliced rpoC1 gene from spinach chloroplast DNA was expressed in E . coli and shown to encode a protein immunologically related to Anabaena gamma . The similarities in the RNA polymerase gene products and gene organizations between cyanobacteria and chloroplasts support the cyanobacterial origin of chloroplasts and a divergent evolutionary pathway among eubacteria.

Biochimie, 1991 Jun, 73(6), 679 - 82
The structure of the gene for ribosomal protein L5 in the archaebacterium Sulfolobus acidocaldarius; Yang D et al.; The gene for the ribosomal protein L5 from the archaebacterium Sulfolobus acidocaldarius has been isolated and sequenced . The gene codes for a basic protein of molecular weight 29 165 Da . This protein shows substantial similarity to the equivalent protein from other archaebacteria as well as from yeast, and considerably less similarity to the equivalent eubacterial protein . These results support the concept of the archaebacteria as a monophyletic kingdom more closely related to eukaryotes than to eubacteria.

Biochimie, 1991 Jun, 73(6), 639 - 45
Mutants lacking individual ribosomal proteins as a tool to investigate ribosomal properties; Dabbs ER; We have isolated and characterized mutants which lack one or two of sixteen of the proteins of the Escherichia coli ribosome . The mutation responsible in each case mapped close to, and probably in, the corresponding gene . A conditional lethal phenotype and a variable degree of impairment in growth was observed . The missing protein was readily restored to the organelle if E coli or other eubacterial ribosomal proteins were added to a suspension of the mutant particles . The mutants have been used to investigate the role of individual proteins in ribosome function and assembly . They have also aided in the topographic pinpointing of proteins on the surface of the organelle.

Biochimie, 1991 Jun, 73(6), 797 - 804
The binding site of ribosomal protein L10 in eubacteria and archaebacteria is conserved: reconstitution of chimeric 50S subunits; Stoffler-Meilicke M et al.; It has been shown by electron microscopy that the selective removal of the stalk from 50S ribosomal subunits of two representative archaebacteria, namely Methanococcus vaniellii and Sulfolobus solfataricus, is accompanied by loss of the archaebacterial L10 and L12 proteins . The stalk was reformed if archaebacterial core particles were reconstituted with their corresponding split proteins . Next, structurally intact chimeric 50S subunits have been reconstituted in vitro by addition of Escherichia coli ribosomal proteins L10 and L7/L12 to 50S core particles from M vaniellii or S solfataricus, respectively . In the reverse experiment, using core particles from E coli and split proteins from M vaniellii, stalk-bearing 50S particles were also obtained . Analysis of the reconstituted 50S subunits by immunoblotting revealed that E coli L10 was incorporated into archaebacterial core particles in both presence or absence of E coli L7/L12 . In contrast, incorporation of E coli L7/L12 into archaebacterial cores was only possible in the presence of E coli L10 . Our results suggest that in archaebacteria - as in E coli - the stalk is formed by archaebacterial L12 proteins that bind to the ribosome via L10 . The structural equivalence of eubacterial and archaebacterial L10 and L12 proteins has thus for the first time been established . The chimeric reconstitution experiments provide evidence that the domain of protein L10 that interacts with the ribosomal particle is highly conserved between eubacteria and archaebacteria.

Biochimie, 1991 Jun, 73(6), 683 - 8
Presence of a gene in the archaebacterium Methanococcus vannielii homologous to secY of eubacteria; Auer J et al.; The nucleotide sequence of a gene located at the promoter-distal side of the 'spectinomycin-operon' homologue of the archaebacterium Methanococcus vannielii was determined . Its derived amino acid sequence displayed 20% (identical positions) or 52% (including conservative exchanges) similarity, respectively, to SECY from E coli . An alignment of the Methanococcus SECY with eubacterial SECY sequences showed the existence of 10 membrane-associated primary structure domains in equivalent positions . The 5' and 3' ends of the secY transcript were mapped and the gene was expressed in the T7 promoter/polymerase system in E col . The temperature-sensitive growth of the E coli mutant IQ292 which harbours a secYts mutation could be complemented by the secY gene from Methanococcus . This indicates that a protein integral to an archaebacterial ether-lipid membrane can be inserted into a eubacterial phospholipid membrane without apparent loss of function.

Biochimie, 1991 Jun, 73(6), 657 - 68
Primary structures of ribosomal proteins from the archaebacterium Halobacterium marismortui and the eubacterium Bacillus stearothermophilus; Arndt E et al.; Approximately 40 ribosomal proteins from each Halobacterium marismortui and Bacillus stearothermophilus have been sequenced either by direct protein sequence analysis or by DNA sequence analysis of the appropriate genes . The comparison of the amino acid sequences from the archaebacterium H marismortui with the available ribosomal proteins from the eubacterial and eukaryotic kingdoms revealed four different groups of proteins: 24 proteins are related to both eubacterial as well as eukaryotic proteins . Eleven proteins are exclusively related to eukaryotic counterparts . For three proteins only eubacterial relatives-and for another three proteins no counterpart-could be found . The similarities of the halobacterial ribosomal proteins are in general somewhat higher to their eukaryotic than to their eubacterial counterparts . The comparison of B stearothermophilus proteins with their E coli homologues showed that the proteins evolved at different rates . Some proteins are highly conserved with 64-76% identity, others are poorly conserved with only 25-34% identical amino acid residues.

Biochimie, 1991 Jun, 73(6), 689 - 97
Structure and evolution of ribonuclease P RNA; Brown JW et al.; Eubacterial RNase P contains a catalytic RNA that cleaves 5' leader sequences from precursor tRNAs . We review the current understanding of RNase P RNA structure and evolution, from the perspective of phylogenetic comparative analysis.

Appl Environ Microbiol, 1991 Jun, 57(6), 1707 - 13
Phylogenetic analysis of a natural marine bacterioplankton population by rRNA gene cloning and sequencing; Britschgi TB et al.; The identification of the prokaryotic species which constitute marine bacterioplankton communities has been a long-standing problem in marine microbiology . To address this question, we used the polymerase chain reaction to construct and analyze a library of 51 small-subunit (16S) rRNA genes cloned from Sargasso Sea bacterioplankton genomic DNA . Oligonucleotides complementary to conserved regions in the 16S rDNAs of eubacteria were used to direct the synthesis of polymerase chain reaction products, which were then cloned by blunt-end ligation into the phagemid vector pBluescript . Restriction fragment length polymorphisms and hybridizations to oligonucleotide probes for the SAR11 and marine Synechococcus phylogenetic groups indicated the presence of at least seven classes of genes . The sequences of five unique rDNAs were determined completely . In addition to 16S rRNA genes from the marine Synechococcus cluster and the previously identified but uncultivated microbial group, the SAR11 cluster {S . J . Giovannoni, T . B . Britschgi, C . L . Moyer, and K . G . Field . Nature (London) 345:60-63}, two new gene classes were observed . Phylogenetic comparisons indicated that these belonged to unknown species of alpha- and gamma-proteobacteria . The data confirm the earlier conclusion that a majority of planktonic bacteria are new species previously unrecognized by bacteriologists.

J Bacteriol, 1991 Jun, 173(12), 3855 - 63
Phylogenetic analysis and evolution of RNase P RNA in proteobacteria; Brown JW et al.; The secondary structures of the eubacterial RNase P RNAs are being elucidated by a phylogenetic comparative approach . Sequences of genes encoding RNase P RNA from each of the recognized subgroups (alpha, beta, gamma, and delta) of the proteobacteria have now been determined . These sequences allow the refinement, to nearly the base pair level, of the phylogenetic model for RNase P RNA secondary structure . Evolutionary change among the RNase P RNAs was found to occur primarily in four discrete structural domains that are peripheral to a highly conserved core structure . The new sequences were used to examine critically the proposed similarity (C . Guerrier-Takada, N . Lumelsky, and S . Altman, Science 246:1578-1584, 1989) between a portion of RNase P RNA and the "exit site" of the 23S rRNA of Escherichia coli . Phylogenetic comparisons indicate that these sequences are not homologous and that any similarity in the structures is, at best, tenuous.

J Biol Chem, 1991 May 25, 266(15), 9622 - 31
Biosynthesis of nucleotides, flavins, and deazaflavins in Methanobacterium thermoautotrophicum; Eisenreich W et al.; The biosynthesis of deazaflavins, flavins, ribonucleotides, and selected amino acids was studied in Methanobacterium thermoautotrophicum by incorporation of 13C-labeled acetate and pyruvate . 13C enrichments were monitored by 13C and 1H NMR spectroscopy . The biosynthesis of ribonucleotides follows the standard pathway . The xylene ring of riboflavin is formed from two pentose moieties in agreement with studies in yeasts and eubacteria . The pyrimidine ring and the ribityl side chain of the deazaflavin chromophore of coenzyme F420 are derived from the purine nucleotide pool . The phenolic ring and C-5 of the deazaflavin system are supplied by the shikimate pathway . A hypothetical mechanism for the assembly of the deazaflavin chromophore from 5-amino-6-ribitylamino-2,4-(1H,3H)-pyrimidinedione and 4-hydroxyphenyl-pyruvate is proposed.

J Endod, 1991 May, 17(5), 207 - 12
Reactivity of IgG from explant cultures of periapical lesions with implicated microorganisms; Baumgartner JC et al.; The presence of IgG in periapical inflammatory lesions suggests that immune responses participate in the disease process . The purpose of this investigation was to study the reactivity of IgG from the supernatant fluids of explant cultures of periapical lesions with microorganisms implicated in infections of endodontic origin . Ninety periapical lesions that had been contiguous with the apex of a root were removed and maintained in explant cultures . A dot-enzyme-linked immunosorbent assay (dot-ELISA) was used to demonstrate the presence of IgG in the supernatant fluids of the explant cultures reactive with a panel of microorganisms associated with infections of endodontic origin . The percentages of reactivity by dot-ELISA follow: Bacteroides intermedius (84%), B . buccae (12%), Porphyromonas (Bacteroides) gingivalis (50%), P . endodontalis (58%), P . asaccharolyticus (17%), Peptostreptococcus micros (44%), P . anaerobius (26%), Eubacterium alactolyticum (34%), Fusobacterium nucleatum (14%), and Actinomyces israelii (6%) . At least one of the three species of B . intermedius, P . gingivalis, or P . endodontalis tested gave a positive dot-ELISA with 89% of the supernatant fluids from explant cultures of periapical lesions . A lack of cross reactivity of IgG in supernatant fluids from explants of periapical lesions was demonstrated for the four strains of black-pigmented Bacteroides/Porphyromonas by dot-ELISA.

Biochim Biophys Acta, 1991 May 2, 1089(1), 113 - 9
Cloning and analysis of the gene (rpoDA) for the principal sigma factor of Pseudomonas aeruginosa; Tanaka K et al.; A gene (rpoDA) of Pseudomonas aeruginosa whose gene product has a homologous function and structure with the principal sigma factor of Escherichia coli was cloned and sequenced . The DNA region corresponding to one of the two hybridization signals found in P . aeruginosa DNA with a synthetic oligonucleotide probe (rpoD probe) was shown to be able to complement a temperature sensitive mutation of Escherichia coli rpoD gene . The amino acid sequence deduced from the nucleotide sequence of rpoDA showed an extensive homology with that of the principal sigma factor of E . coli throughout the entire region, which indicates that the two gene products have an essentially identical domain structure . A common basic structure observed among principal sigma factors of different eubacterial strains was proposed . RpoDA protein was identified in the extract of the cell carrying a plasmid clone with the rpoDA gene insert by Western blot analysis.

Trends Genet, 1991 May, 7(5), 145 - 8
Intron phylogeny: a new hypothesis; Cavalier-Smith T; The three major classes of intron are clearly of unequal antiquity . Structured (often self-splicing and sometimes mobile) introns are the most ancient, probably dating (at least for group I) from the ancestral (eubacterial) cell 3500 million years ago, and were originally restricted to tRNA . Protein-spliced introns (usually in tRNA) probably evolved from them by a radical change in splicing mechanism in the common ancestor of eukaryotes and archaebacteria, perhaps only about 1700 million years ago . Spliceosomal introns probably evolved from group-II-like self-splicing introns after the origin of the nucleus between 1700 and 1000 million years ago, and were probably mostly inserted into previously unsplit protein-coding genes after the origin of mitochondria 1000 million years ago.

Curr Genet, 1991 May, 19(5), 417 - 22
A small gene family in barley encodes ribosomal proteins homologous to yeast YL17 and L22 from archaebacteria, eubacteria, and chloroplasts; Madsen LH et al.; The amino acid sequences of two barley ribosomal proteins, termed HvL17-1 and HvL17-2, were decoded from green leaf cDNA clones . The N-terminal sequences of the derived barley proteins are 48% identical to the N-terminal amino acid sequence of protein YL17 from the large subunit of yeast cytoplasmic ribosomes . Via archaebacterial ribosomal proteins this homology extends to ribosomal protein L22 from eubacteria and chloroplast . Barley L17, and ribosomal proteins L22 and L23 from the archaebacteria Halobacterium halobium and H . marismortui, are 25-33% identical . Interestingly, the barley and archaebacterial proteins share a long, central stretch of amino acids, which is absent in the corresponding proteins from eubacteria and chloroplasts . Barley L17 proteins are encoded by a small gene family with probably only two members, represented by the cDNA clones encoding HvL17-1 and HvL17-2 . Both these genes are active in green leaf cells . The expression of the L17 genes in different parts of the 7-day old barley seedlings was analyzed by semiquantitative hybridization . The level of L17 mRNA is high in meristematic and young cells found in the leaf base and root tip . In the leaf, the L17 mRNA level rapidly decreases with increasing cell age, and in older root cells this mRNA is undetectable.

FEMS Microbiol Lett, 1991 May 1, 64(1), 29 - 33
Establishment of Eubacterium cellulosolvens in the digestive tract of axenic and meroxenic mice: influence of feed cellulose content; Boulahrouf A et al.; The cellulolytic bacterial species present in the caecum and colon contents of conventionally reared mature mice did not become established in the digestive tract when inoculated to axenic mice, whatever the size of the inoculum or the cellulose content of the diet . The cellulolytic bacterium Eubacterium cellulosolvens SC 10 isolated from mouse digestive flora was unable to become established in the digestive tract of axenic mice whatever the cellulose content of the diet; it requires a feed rich in cellulose and a highly diversified microflora.

J Bacteriol, 1991 May, 173(9), 2969 - 76
Transcription of the myxobacterial hemagglutinin gene is mediated by a sigma 54-like promoter and a cis-acting upstream regulatory region of DNA; Romeo JM et al.; Myxobacterial hemagglutinin (MBHA) is a major developmentally induced protein that accumulates during the period of cellular aggregation of the fruiting bacterium Myxococcus xanthus . In this study, DNA sequences mediating the transcriptional regulation of mbhA have been identified . Examination of nucleotide sequences upstream of the start site for mbhA transcription has indicated a region of DNA that bears strong homology to the consensus sequence for promoters recognized by the sigma 54 holoenzyme form of RNA polymerase of Escherichia coli and other eubacteria . Deletion of this sequence completely abolished mbhA transcription . Additionally, a cis-acting DNA element, affecting the efficiency of mbhA transcription, has been mapped within a region of DNA 89 to 276 nucleotides upstream of the sigma 54-like sequence . Transposon insertions, mapping within the cis element, drastically reduced mbhA transcriptional activity . These observations suggest that transcription of mbhA requires a productive interaction between a form of RNA polymerase that recognizes a sigma 54-like sequence and a transcriptional activator that binds to DNA sequences upstream of the mbhA promoter.

Mol Microbiol, 1991 May, 5(5), 1183 - 6
Selenocysteyl-tRNA occurs in the diatom Thalassiosira and in the ciliate Tetrahymena; Hatfield DL et al.; Selenocysteyl-tRNAs that decode UGA were identified previously in animal and bacterial cells and the genes for these tRNAs have been shown to be widespread in animals and eubacteria . In the present study, we identify a selenocysteyl-tRNA that codes for UGA in Thalassiosira pseudonana, which is a diatom, and in Tetrahymena borealis, which is a ciliate . The fact that these very diverse unicellular organisms also contain a selenocysteyl-tRNA suggests that selenocysteine-containing proteins and the use of UGA as a codon for selenocysteine are widespread, if not ubiquitous, in nature.

New Biol, 1991 May, 3(5), 430 - 8
4.5S RNA: does form predict function?
Brown S.
4.5S RNA is a stable RNA of Escherichia coli, and functional homologs of the molecule apparently exist in all prokaryotes: eubacteria, archebacteria, and mycoplasma . Genetic and physiological measurements of the function of 4.5S RNA in E . coli indicate a role for this RNA in protein synthesis . A conserved domain of 4.5S RNA displays structural similarity with the eukaryotic 7S RNA that functions in protein secretion . Although complementation by eukaryotic 7S RNAs remains to be demonstrated, a number of archaebacterial 7S RNAs are able to replace 4.5S RNA for growth of E . coli, and 4.5S RNA is able to mediate a number of 7S RNA functions in vitro . Surprisingly, no effects on protein secretion in E . coli have been directly attributed to 4.5S RNA . These observations raise the question of whether molecules of similar structure necessarily perform the same function.

Pathol Biol (Paris), 1991 May, 39(5), 379 - 83
{Strict anaerobic bacteria: comparative study of various beta-lactam antibiotics in combination with tazobactam or sulbactam}; Dubreuil L et al.; The minimal inhibitory concentrations of piperacillin (PIP) or cefotaxime (CTX) alone or in combination with tazobactam (TAZ) were determined against 168 anaerobes . All the strains were inhibited by PIP + TAZ, but certain strains resistant to CTX + TAZ were found among B . fragilis, Eubacterium and Peptostreptococcus . The second investigations included 30 strains of Bacteroides fragilis . Concentrations of 2, 4 and 8 mg/l of TAZ and sulbactam (SUL) were combined with piperacillin or cefotaxime . The two beta-lactamase-inhibitors had similar activities when used at 2 or 4 mg/l, but at 8 mg/l TAZ was more active than SUL . All B . fragilis strains were inhibited by PIP + TAZ or PIP + SUL, whereas resistance was observed with CTX + SUL or CTX + TAZ . On the same strains the activities of 6 beta-lactams (PIP, mezlocillin, ticarcillin (TIC), CTX, ceftriaxone and ceftazidime) were determined in combination with either SUL 4 mg/l or TAZ 8 mg/l . Only PIP or TIC + SUL or TAZ were able to inhibit at least 90% of tested strains . No resistance could be detected with PIP + TAZ combination . As conclusion, the two inhibitors when combined with PIP or TIC offered greater activity against both Gram positive or negative anaerobes and PIP + TAZ remained the more potent combination.

Nucleic Acids Res, 1991 Apr 25, 19 Suppl, 2189 - 91
Compilation of 5S rRNA and 5S rRNA gene sequences; Specht T et al.; This is an update for the 5S rRNA sequences of the BERLIN RNA DATABANK last published in 1990 (1) . The new entry consists of 25 eubacterial and 2 eukaryotic 5S rRNA sequences and 10 plant 5S rRNA pseudogenes (Table 1) . Thus the BERLIN RNA DATABANK contains as of February 1, 1991 the 5S rRNA sequences of 44 archaebacteria, 292 eubacteria, 20 plastids, 6 mitochondria, 321 eukaryotes and 21 eukaryotic pseudogenes . The BERLIN RNA DATABANK uses the format of the EMBL Nucleotide Sequence Data Library complemented by a Sequence Alignment (SA) field including secondary structure information.

FEBS Lett, 1991 Apr 9, 281(1-2), 114 - 8
Complete nucleotide sequence of the Mycobacterium leprae 23 S and 5 S rRNA genes plus flanking regions and their potential in designing diagnostic oligonucleotide probes; Liesack W et al.; The complete nucleotide sequences of the Mycobacterium leprae 23 S and 5 S rRNA genes and their flanking regions are presented . As compared to other eubacterial homologous molecules the 23 S rDNA exhibits two insertions . A 16 nucleotide long insertion is almost unique to members of the genus Mycobacterium, while the second represents an extended version of helix 54 . The potential of both insertions to serve as target for diagnostic oligonucleotide probes was proven by comparative sequence analysis of 23 S rRNA of several Mycobacterium species and by dot blot hybridization . In addition, a 19-mer oligonucleotide probe is described, which can be considered genus Mycobacterium-specific.

J Bacteriol, 1991 Apr, 173(7), 2244 - 9
Effect of paracrystalline protein surface layers on predation by Bdellovibrio bacteriovorus; Koval SF et al.; We determined that paracrystalline protein surface arrays (S layers) protected gram-negative eubacteria from predation by Bdellovibrio bacteriovorus . Aquaspirillum serpens VHA and MW5 and Aquaspirillum sinuosum were resistant to predation by B . bacteriovorus 6-5-S when fully covered by their S layers . The S layer of Aeromonas salmonicida A449 protected the cells from predication by B . bacteriovorus 109J . A predacious, plaque-forming vibrio that lysed an S-layer- variant of Caulobacter crescentus but was not predacious on the parental strain which possessed an S layer was isolated from raw sewage . Since S layers are stable components of many bacterial surfaces in nature, they can provide this protective function in both aquatic and terrestrial habitats where Bdellovibrio spp . are found.

J Lipid Res, 1991 Apr, 32(4), 659 - 66
Archaebacterial ether lipid diversity analyzed by supercritical fluid chromatography: integration with a bacterial lipid protocol; Hedrick DB et al.; A strategy has been developed for archaebacterial lipid analysis which provides three times the information to describe archaebacterial isolates and is compatible with simultaneous eubacterial/eukaryotic lipid analysis of environmental samples . Eubacterial and micro-eukaryotic biomass, community structure, and nutritional status have been routinely defined in environmental samples by lipid analysis . Lipid profiles are also useful in eubacterial identification and taxonomy . Polar lipid or whole cell ester-linked fatty acids are generally analyzed by gas chromatography-mass spectroscopy . Archaebacteria are characterized by their ether-linked membrane lipids . There is, however, less diversity in the side chains of archaebacterial membrane lipids as compared the eubacterial ester-linked membrane lipids . The information content of the archaebacterial lipid profile was increased by separately analyzing the polar lipid, glycolipid, and lipid-extracted residue fractions . Identification and quantification were performed by supercritical fluid chromatography . Results are presented for three species of methanogens and four thermoacidophile isolates, and compared with a literature review.

J Bacteriol, 1991 Apr, 173(8), 2442 - 50
Isolation and characterization of the gene encoding the principal sigma factor of the vegetative cell RNA polymerase from the cyanobacterium Anabaena sp . strain PCC 7120; Brahamsha B et al.; The filamentous cyanobacterium Anabaena sp . strain PCC 7120 responds to combined nitrogen deprivation by forming specialized nitrogen-fixing cells at regular intervals along the filament . Genetic and biochemical studies have indicated that regulation of gene expression during differentiation occurs at the transcriptional level . As part of a characterization of RNA polymerase during differentiation, the gene encoding the 52-kDa principal sigma factor of the Anabaena sp . strain PCC 7120 vegetative-cell RNA polymerase was isolated by using an oligonucleotide probe based on the sequence of the N-terminal seven amino acids of the purified protein . sigA codes for a 390-amino-acid polypeptide that has a predicted molecular weight of 45,641 . The amino acid sequence of the polypeptide encoded by sigA contains four regions corresponding to conserved domains of the principal RNA polymerase sigma factors of Escherichia coli (sigma 70) and Bacillus subtilis (sigma 43) . Thus, although the subunit composition of cyanobacterial RNA polymerase core differs from that of other eubacteria (G . J . Schneider and R . Haselkorn, J . Bacteriol . 170:4136-4140, 1988), the principal sigma factor of at least one cyanobacterium is typically eubacterial . In contrast to sigma 70 and sigma 43 operon organization, sigA is monocistronic and encodes two transcripts of 1.7 and 2.2 kb . The abundance of the 1.7-kb transcript remains constant under both nitrogen-replete and nitrogen-limiting conditions, whereas the 2.2-kb transcript is induced following the removal of combined nitrogen . Continued or enhanced transcription of sigA under nitrogen starvation conditions is consistent with the observation that the principal RNA polymerase in differentiating cells contains SigA.

Int J Syst Bacteriol, 1991 Apr, 41(2), 324 - 5
Phylogenetic analysis of Aquaspirillum magnetotacticum using polymerase chain reaction-amplified 16S rRNA-specific DNA; Eden PA et al.; The 16S rRNA gene of the magnetotactic magnetogen Aquaspirillum magnetotacticum MS1 was amplified by a polymerase chain reaction, using two eubacterial consensus oligodeoxynucleotide primers flanking the majority of the 16S rRNA gene, cloned, and sequenced . Phylogenetic analysis revealed that A . magnetotacticum MS1 belongs to the alpha-group of proteobacteria . This assignment offers perspective on the biochemical properties of A . magnetotacticum, since this organism is expected to have the general properties that are common to this phylogenetic group.

Protein Expr Purif, 1991 Apr-Jun, 2(2-3), 117 - 21
Rapid method for altering bacterial ribosome-binding sequences for overexpression of proteins in Escherichia coli; Roberts I et al.; In an Escherichia coli expression system, two genes, one from an anaerobic intestinal bacterium and one from E . coli, were overexpressed following the alteration of ribosome-binding (Shine-Dalgarno) sequences . For both genes, the polymerase chain reaction (PCR) was used to modify the ribosome-binding sequence and, at the same time, provide restriction endonuclease sequences at each end of the gene . These restriction endonuclease sequences were used for inserting the DNA into the E . coli plasmid vector pGEM2, which has the T7 promoter upstream from its multiple cloning sites . Each chimeric plasmid, made by ligating the PCR product into pGEM2, was transformed into E . coli strain HMS174(DE3) which, when induced, produces T7 RNA polymerase for regulated overexpression . The gene isolated from the anaerobic intestinal bacterium, a 27-kDa polypeptide gene from Eubacterium sp . strain 12708, when expressed using this system, produced about one-third of the total cell protein as measured in Coomassie-stained protein gels and confirmed by Western blots with rabbit antibody . The E . coli enzyme, a 28.4-kDa tRNA methylation enzyme, was increased fivefold in activity of cell extracts over that of the best previous strain.

J Biol Chem, 1991 Mar 25, 266(9), 5689 - 95
The RNA component of RNase P from the archaebacterium Haloferax volcanii; Nieuwlandt DT et al.; RNase P, an endoribonuclease responsible for generating the mature 5' termini of tRNA precursors, is composed of both RNA and protein . It has been demonstrated that the eubacterial RNase P RNA will, under the appropriate reaction conditions, exhibit catalytic activity in vitro . Evidence has not been obtained for catalytic activity by the RNAs of eukaryotic RNase P enzymes . Using a cDNA probe prepared from RNA copurifying with RNase P activity from the archaebacterium Haloferax volcanii, we have characterized the gene encoding the RNase P RNA . The proposed transcript from this gene can assume a structure resembling the eubacterial RNase P RNA and includes many of the highly conserved sequences of these RNAs . This RNA was incapable of cleaving pre-tRNA substrates in the absence of protein under a variety of in vitro conditions . Catalytic activity was observed when this RNA was combined with the protein subunit of the Bacillus subtilis RNase P complex . Similarities among the archaebacterial, eubacterial, and eukaryotic RNase P RNA sequences and structures are discussed.

Chem Pharm Bull (Tokyo), 1991 Mar, 39(3), 757 - 60
Barbaloin stimulates growth of Eubacterium sp . strain BAR, a barbaloin-metabolizing bacterium from human feces; Che QM et al.; Eubacterium sp . strain BAR, isolated from human feces, transformed barbaloin to aloe-emodin anthrone in a basal medium lacking carbohydrate . Barbaloin remarkably stimulated the growth of strain BAR in the basal medium, the stimulative extent of the growth depending on the amount of barbaloin added . The addition of D-glucose, D-galactose, maltose, cellobiose, sucrose or D-amygdalin to the basal medium containing barbaloin caused a decrease of the growth stimulated by barbaloin to the growth level with each sugar, resulting in a complete inhibition of the barbaloin transformation . On the other hand, the addition of D-fructose, which itself stimulated the growth of strain BAR, further increased the growth in the presence of barbaloin and little inhibited barbaloin transformation . Nojirimycin bisulfite, a specific inhibitor of glucosidases, potently inhibited the growth with barbaloin, but did not affect the growth with glucose or cellobiose . Also, nojirimycin bisulfite completely inhibited the transformation of barbaloin to aloe-emodin anthrone . These results indicate that a unique enzyme capable of cleaving the C-glycosyl bond is induced in strain BAR by barbaloin and, consequently, strain BAR grows by utilizing as a nutrient the carbohydrate liberated from barbaloin . It is further suggested that the barbaloin-cleaving enzyme is inhibited by nojirimycin bisulfite and that the induction of the enzyme is repressed with D-glucose and D-galactose.

Chem Pharm Bull (Tokyo), 1991 Mar, 39(3), 729 - 31
Induction and inhibition of novel sulfotransferase in a human intestinal bacterium, Eubacterium sp . A-44; Kim DH et al.; Induction and inhibition of a novel sulfotransferase produced by Eubacterium sp . A-44 isolated from human feces have been studied . Production of the enzyme was induced by phenylsulfate esters, sulfate donor substrates, but not by phenols, sulfate acceptor substrates, or inorganic sulfate . p-Nitrophenylsulfate (PNS), a good donor substrate, stimulated enzyme production more than 10-fold . Sulfotransferase production was strongly inhibited by phenylphosphate esters . Enzyme activity was competitively inhibited by phenylphosphate esters, but not by inorganic phosphate . High yields of sulfotransferase from sonicated cells were obtained when the bacteria were grown in a media containing 0.6% (w/v) or less of glycine.

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