Eubacterium

FEMS Microbiol Lett, 1991 Mar 1, 62(2-3), 139 - 43
A novel eubacterial phylum: comparative nucleotide sequence analysis of a tuf-gene of Flexistipes sinusarabici; Ludwig W et al.; A tuf-gene of Flexistipes sinusarabici has been cloned and sequenced . The primary structure of the predicted elongation factor protein was compared with available sequences of homologous genes and elongation factors Tu or 1 alpha of eubacteria, archaebacteria and eukaryotes . Based on elongation factor Tu data Flexistipes sinusarabici belongs to the eubacterial kingdom but no specific relationship to any phylum was detected . The amino acid of the elongation factor Tu of F . sinusarabici exhibits no striking pecularities . Among the highly conserved positions only two are different . Sites of known or postulated functions are conserved.

Microbiol Rev, 1991 Mar, 55(1), 143 - 90
Interaction of chlamydiae and host cells in vitro; Moulder JW; The obligately intracellular bacteria of the genus Chlamydia, which is only remotely related to other eubacterial genera, cause many diseases of humans, nonhuman mammals, and birds . Interaction of chlamydiae with host cells in vitro has been studied as a model of infection in natural hosts and as an example of the adaptation of an organism to an unusual environment, the inside of another living cell . Among the novel adaptations made by chlamydiae have been the substitution of disulfide-bond-cross-linked polypeptides for peptidoglycans and the use of host-generated nucleotide triphosphates as sources of metabolic energy . The effect of contact between chlamydiae and host cells in culture varies from no effect at all to rapid destruction of either chlamydiae or host cells . When successful infection occurs, it is usually followed by production of large numbers of progeny and destruction of host cells . However, host cells containing chlamydiae sometimes continue to divide, with or without overt signs of infection, and chlamydiae may persist indefinitely in cell cultures . Some of the many factors that influence the outcome of chlamydia-host cell interaction are kind of chlamydiae, kind of host cells, mode of chlamydial entry, nutritional adequacy of the culture medium, presence of antimicrobial agents, and presence of immune cells and soluble immune factors . General characteristics of chlamydial multiplication in cells of their natural hosts are reproduced in established cell lines, but reproduction in vitro of the subtle differences in chlamydial behavior responsible for the individuality of the different chlamydial diseases will require better in vitro models.

J Bacteriol, 1991 Mar, 173(6), 2086 - 92
Characterization of enzymes of the branched-chain amino acid biosynthetic pathway in Methanococcus spp; Xing RY et al.; Methanococcus aeolicus, Methanococcus maripaludis, and Methanococcus voltae contain similar levels of four enzymes of branched-chain amino acid biosynthesis: acetohydroxy acid synthase, acetohydroxy acid isomeroreductase, dihydroxy acid dehydratase, and transaminase B . Following growth at low partial pressures of H2-CO2, the levels of these enzymes in extracts of M . voltae are reduced three- to fivefold, which suggests that their synthesis is regulated . The enzymes from M . aeolicus were found to be similar to the eubacterial and eucaryotic enzymes with respect to molecular weights, pH optima, kinetic properties, and sensitivities to O2 . The acetohydroxy acid isomeroreductase has a specific requirement for Mg2+, and other divalent cations were inhibitory . It was stimulated threefold by K+ and NH4+ ions and was able to utilize NADH as well as NADPH . The partially purified enzyme was not sensitive to O2 . The dihydroxy acid dehydratase is extremely sensitive to O2, and it has a half-life under 5% O2 of 6 min at 25 degrees C . Divalent cations were required for activity, and Mg2+, Mn2+, Ni2+, Co2+, and Fe2+ were nearly equally effective . In conclusion, the archaebacterial enzymes are functionally homologous to the eubacterial and eucaryotic enzymes, which implies that this pathway is very ancient.

J Bacteriol, 1991 Mar, 173(5), 1835 - 7
Genes for 7S RNAs can replace the gene for 4.5S RNA in growth of Escherichia coli; Brown S; 4.5S RNAs of eubacteria and 7S RNAs of archaebacteria and eukaryotes exist in a hairpin conformation . The apex of this hairpin displays structural and sequence similarities among both 4.5S and 7S RNAs . Furthermore, a hyphenated sequence of 16 nucleotides is conserved in all eubacterial 4.5S RNAs examined . In this article I report that 7S RNAs that contain this 16-nucleotide sequence are able to replace 4.5S RNAs and permit growth of Escherichia coli.

J Bacteriol, 1991 Mar, 173(6), 2035 - 44
Sequence and TnphoA analysis of a Mycoplasma hyorhinis protein with membrane export function; Yogev D et al.; Proteins translocated across the single plasma membrane of mycoplasmas (class Mollicutes) represent important components likely to affect several interactions of these wall-less microbes with their respective hosts . However, identification and functional analysis of such proteins is hampered by the lack of mutational systems in mycoplasmas and by a perceived limitation in translating recombinant mycoplasma genes containing UGA (Trp) codons in other eubacteria . Here we directly analyze a gene encoding a Mycoplasma hyorhinis protein capable of promoting its membrane translocation . It was initially detected by screening a recombinant phage genomic library with antibody from a host with M . hyorhinis-induced arthritis and was localized by Tn5 and deletion mutations affecting expression of antigenic translational products . Sequence analysis of the isolated gene predicted a hydrophilic protein, P101, containing three UGA codons and a putative signal peptide with an uncharacteristic cluster of positively charged amino acids near its C terminus . Nevertheless, lambda::TnphoA transposon mutagenesis of an Escherichia coli plasmid bearing the p101 gene resulted in p101::TnphoA fusions expressing products that could translocate as much as 48 kDa of the P101 sequence (up to the first UGA codon) across the E . coli plasma membrane . Fusion proteins containing mature P101 sequences expressed mycoplasma epitopes and were found by cell fractionation and detergent phase partitioning to be integral membrane proteins in E . coli, suggesting a lack of signal peptide cleavage in this system . Importantly, identification of P101 by direct analysis of its export function relied neither on prior identification of the mycoplasmal product nor on complete expression of the product from the cloned mycoplasma gene.

Planta Med, 1991 Feb, 57(1), 15 - 9
Isolation of a human intestinal bacterium capable of transforming barbaloin to aloe-emodin anthrone; Che QM et al.; A strictly anaerobic bacterium, Eubacterium sp . BAR, was isolated from human feces as one of the intestinal bacteria capable of metabolizing barbaloin . The bacterium grew in PYF broth containing barbaloin and converted barbaloin to aloe-emodin anthrone . On the other hand, the bacterium had little metabolic activity in GAM broth.

J Steroid Biochem Mol Biol, 1991 Feb, 38(2), 257 - 63
Characteristics of 16-dehydroprogesterone reductase in cell extracts of the intestinal anaerobe, Eubacterium sp . strain 144; Watkins WE et al.; 16-Dehydroprogesterone reductase (16-DHPR) activity was present in cell extracts of Eubacterium sp . strain 144 only when the organism was grown in the presence of steroids containing a delta 16-17 double bond and C-20-ketone . Cells grown with 16-dehydropregnenolone contained 16-DHPR activity but lacked delta 4-5-3-keto steroid reductase activity . Pyruvate or sodium dithionite served as electron donors for 16-DHPR and both reactions required methyl viologen as an electron carrier . Neither NADH nor NADPH, with or without flavin nucleotides, were used by 16-DHPR . Enzyme activity was detected in the cytoplasmic fraction (40%) and membrane fraction (20%) of crude cell extracts, but 40% of the activity was unaccounted for following ultracentrifugation . 16-DHPR activity was unaffected by pH in potassium phosphate buffer over the range 5.0 to 8.5, but was inhibited by Tris-HCl above pH 7.0 . 16-DHPR activity was inhibited by sulfhydryl reagents, but inhibitors of electron transport reactions or metal chelators did not affect the enzyme.

Biochem J, 1991 Feb 1, 273 ( Pt 3), 627 - 34
Early steps of isoprenoid biosynthesis in Escherichia coli; Zhou D et al.; The incorporation of 2H- and 13C-labelled precursors into ubiquinone-8 (Uq-8) by strains of Escherichia coli was measured in order to define the pathway for the early steps in the biosynthesis of isoprenoids in these eubacteria . Cells grown with DL-{methyl-2H6}valine were found to label both the alpha-oxoisovaleric ('alpha-ketoisovaleric') acid alpha-oxoisohexanoic ('alpha-ketoisocaproic') acid, but not the Uq-8 . Since these acids are required for the biosynthesis of isoprenoids by the acetolactate pathway, the operation of this pathway in the biosynthesis of Uq-8 is excluded . Cells grown with {1,2-13C2}acetate and non-labelled glucose readily incorporated 13C2 units into fatty acids, but failed to incorporate any label into the Uq-8 . Cells grown with {U-13C6}glucose and non-labelled acetate, however, were found to label both the fatty acids and the Uq-8 . Oxidative cleavage with periodate/permanganate of the Uq-8 isolated from cells grown with U-13C6-labelled glucose produced laevulinic acid, which was shown to be derived from two C2 units and one C1 unit of the labelled glucose by mass-spectral analysis of the 4,5-dihydro-6-methyl-2-phenylpyridazin-3(2H)-one derivative . The results of this work indicate that the C-2 and C-3 carbon unit of pyruvate, not acetyl-CoA, is the precursor to isopentenyl pyrophosphate (IPP) in these cells; however, the labelling pattern observed is consistent with the established acetoacetate pathway of isoprenoid biosynthesis . These data, coupled with the observed lack of inhibition of the growth of E . coli by mevinolin, a specific inhibitor of 3-hydroxy-3-methylglutaryl-CoA, can be best rationalized by the biosynthesis of IPP occurring in E . coli through a series of bound intermediates.

Proc Natl Acad Sci U S A, 1991 Feb 1, 88(3), 921 - 5
CGG: an unassigned or nonsense codon in Mycoplasma capricolum; Oba T et al.; CGG is an arginine codon in the universal genetic code . We previously reported that in Mycoplasma capricolum, a relative of Gram-positive eubacteria, codon CGG did not appear in coding frames, including termination sites, and tRNA(ArgCCG) pairing with codon CGG, was not detected . These facts suggest that CGG is a nonsense (unassigned and untranslatable) codon--i.e., not assigned to arginine or to any other amino acid . We have investigated whether CGG is really an unassigned codon by using a cell-free translation system prepared from M . capricolum . Translation of synthetic mRNA containing in-frame CGG codons does not result in "read-through" to codons beyond the CGG codons--i.e., translation ceases just before CGG . Sucrose-gradient centrifugation profiles of the reaction mixture have shown that the bulk of peptide that has been synthesized is attached to 70S ribosomes and is released upon further incubation with puromycin . The result suggests that the peptide is in the P site of ribosome in the form of peptidyl-tRNA, leaving the A site empty . When in-frame CGG codons are replaced by UAA codons in mRNA, no read-through occurs beyond UAA, just as in the case of CGG . However, the synthesized peptide is released from 70S ribosomes, presumably by release factor 1 . These data suggest strongly that CGG is an unassigned codon and differs from UAA in that CGG is not used for termination.

FEMS Microbiol Lett, 1991 Feb, 62(1), 53 - 8
Propionigenium modestum: a separate line of descent within the eubacteria; Both B et al.; The complete nucleotide sequence of 16S rRNA from Propionigenium modestum was determined and compared with 380 16S rRNA sequences from representatives of all eu- and archaebacterial phyla known so far . The phylogenetic analysis of this data set indicated P . modestum to represent a new separated line of descent within the radiation of eubacterial phyla moderately related to cyanobacteria and Gram-positive bacteria with low DNA GC content.

Zhonghua Yi Xue Za Zhi, 1991 Feb, 71(2), 72 - 5, 6
{Bacteriological study on juvenile periodontitis}; Han N; The predominant cultivable microflora of 23 pockets in 15 juvenile periodontitis (JP) patients was studied for the first time in China using the current anaerobic methodology . Samples were taken with sterile paper points and dispersed on a vortex mixer . Then the diluted samples were plated on the non-selective blood agar plates and selective MGB medium which favors the growth of Actinobacillus actimycetemcomitans (Aa) and incubated in anaerobic chamber for 5 days . From each sample 15 or more isolated colonies were picked in sequence without selection and subcultured . The isolates were identified mainly by Schrechenberger's 4 hour rapid methods for biochemical and fermentative tests and the chromatographic analysis of acid end products using ion-chromatography . The results were as follows: 1 . The microflora of healthy sulci of 7 healthy young subjects was significantly different from that in the pocket of JP patients . The predominant species in healthy sulci were Streptococcus spp and Capnocytophaga gingivalis . 2 . The species increased significantly in JP patients in prevalence and proportions was Eubacterium . Other species in high proportions were Bacteroides oris, B . melaninogenicus, B . gingivalis, Capnocytophaga sputigena, and Actinomyces meyeri, etc . 3 . Actinobacillus actinomycetemcomitans was not detected in any of the samples.

Eur J Biochem, 1991 Jan 30, 195(2), 321 - 7
Gene for the ADP-ribosylatable elongation factor 2 from the extreme thermoacidophilic archaebacterium Sulfolobus acidocaldarius . Cloning, sequencing, comparative analysis; Schroder J et al.; The gene coding for ADP-ribosylatable elongation factor 2 (EF-2) from the extreme thermoacidophilic archaebacterium Sulfolobus acidocaldarius has been cloned and its sequence is reported . Amino acid sequence comparisons showed that EF-2 from S . acidocaldarius is more closely related to eukaryotic EF-2 than to eubacterial EF-G . Consensus sequences are derived from comparison of a region around the unique amino acid diphthamide, which is the target for ADP-ribosylation by diphtheria toxin in archaebacteria and eukaryotes . The conserved positions are likely to constitute a recognition site for the toxin and the histidine-modifying enzymes . A single transcript of approximately the size of the EF-2 gene was observed in Northern blot experiments . Transcription initiation and termination signals were identified in the immediate vicinity of the respective translation start and stop codons of the gene . These results indicate that, in contrast to all prokaryotic EF-2 genes studied previously, the gene of S . acidocaldarius is not located within the streptomycin operon but is transcribed separately.

FEMS Microbiol Lett, 1991 Jan 15, 61(2-3), 217 - 22
Expression in Caulobacter crescentus of the phosphate-starvation-inducible porin OprP of Pseudomonas aeruginosa; Walker SG et al.; The gene for the phosphate-starvation-inducible outer membrane protein OprP, of Pseudomonas aeruginosa was introduced into Caulobacter crescentus CB2A on a plasmid vector . As is the case in P . aeruginosa and Escherichia coli the oprP gene was inducible under conditions of limiting phosphate in C . crescentus . However, the maximal medium concentration of phosphate which still permitted induction of OprP was lower in C . crescentus (50 microM) than in P . aeruginosa (200 microM) . Induction of OprP was coincident with the process of stalk elongation, known to occur in C . crescentus under phosphate starvation conditions . When induced, OprP was localized to the cell envelope and became a major membrane protein, indicating that the Pseudomonas promoter was efficiently recognized in C . crescentus and that the gene product was targeted to the appropriate region of the cell . Our data provide support for the hypothesis that the mechanism for regulation of phosphate-starvation-inducible genes is highly conserved amongst the eubacteria.

FEMS Microbiol Lett, 1991 Jan 15, 61(2-3), 151 - 6
A phylogenetic comparison of the 16S rRNA sequence of the fish pathogen, Renibacterium salmoninarum, to gram-positive bacteria; Gutenberger SK et al.; The 16S rRNA of Renibacterium salmoninarum, the causative agent of bacterial kidney disease in salmonids, was sequenced by reverse transcriptase to produce a nearly complete sequence (97%) of 1475 nucleotides . Phylogenetic comparisons to seventeen genera and signature sequence analysis indicated that R . salmoninarum was a member of the high G + C Gram-positive eubacterial subdivision although the reported G + C value is only 53% . A phylogenetic tree details the relationship of R . salmoninarum to ten actinomycetes from diverse environments.

Free Radic Res Commun, 1991, 12-13 Pt 1, 443 - 9
Halobacterium halobium Mn-SOD gene: archaebacterial and eubacterial features; Salin ML et al.; A 1.8 kb PstI fragment from Halobacterium halobium DNA was found to hybridize to synthetic oligonucleotide probes constructed by using the sequence of the N-terminus of a Mn-containing superoxide dismutase purified from H . halobium . The entire insert containing a 600-bp coding sequence for Mn-SOD and its 5' and 3' flanking regions was sequenced . The derived amino acid sequence of the structural gene showed a similarity to other manganic and iron-containing superoxide dismutases in normally conserved regions . Primer extension analysis of the H . halobium Mn-SOD mRNA showed that gene transcription begins 14 bases upstream of the translational start . A Shine-Dalgarno sequence and archaebacterial consensus promoter sequences were observed . Several other promoter and terminator nucleotide sequences homologous to prokaryotic and eukaryotic organisms were found.

Biochem Cell Biol, 1991 Jan, 69(1), 72 - 8
Structure of the glycan chain from the surface layer glycoprotein of Bacillus alvei CCM 2051; Altman E et al.; The cell surface of the mesophilic eubacterium Bacillus alvei CCM 2051 is covered by an oblique arranged surface layer glycoprotein . The subunits revealed by sodium dodecyl sulfate - polyacrylamide gel electrophoresis were distinct bands of molecular masses 140,000, 128,000, and 127,000 . Proteolytic degradation of the purified S-layer glycoprotein yielded a single glycopeptide fraction with an apparent molecular mass of ca . 25,000 . Methylation analysis in conjunction with two-dimensional nuclear magnetic resonance experiments at 500 MHz established the branched trisaccharide (formula; see text) as the repeating unit for this glycan chain.

J Lipid Res, 1991 Jan, 32(1), 89 - 96
Mechanism of intestinal 7 alpha-dehydroxylation of cholic acid: evidence that allo-deoxycholic acid is an inducible side-product; Hylemon PB et al.; We previously reported that the 7 alpha-dehydroxylation of cholic acid appears to be carried out by a multi-step pathway in intestinal anaerobic bacteria both in vitro and in vivo . The pathway is hypothesized to involve an initial oxidation of the 3 alpha-hydroxy group and the introduction of a double bond at C4-C5 generating a 3-oxo-4-cholenoic bile acid intermediate . The loss of water generates a 3-oxo-4,6-choldienoic bile acid which is reduced (three steps) yielding deoxycholic acid . We synthesized, in radiolabel, the following putative bile acid intermediates of this pathway 7 alpha,12 alpha-dihydroxy-3-oxo-4-cholenoic acid, 7 alpha,12 alpha-dihydroxy-3-oxo-5 beta-cholanoic acid, 12 alpha-dihydroxy-3-oxo-4,6-choldienoic acid, and 12 alpha-hydroxy-3-oxo-4-cholenoic acid and showed that they could be converted to 3 alpha,12 alpha-dihydroxy-5 beta-cholanoic acid (deoxycholic acid) by whole cells or cell extracts of Eubacterium sp . VPI 12708 . During studies of this pathway, we discovered the accumulation of two unidentified bile acid intermediates formed from cholic acid . These bile acids were purified by thin-layer chromatography and identified by gas-liquid chromatography-mass spectrometry as 12 alpha-hydroxy-3-oxo-5 alpha-cholanoic acid and 3 alpha,12 alpha-dihydroxy-5 alpha-cholanoic (allo-deoxycholic acid) . Allo-deoxycholic acid was formed only in cell extracts prepared from bacteria induced by cholic acid, suggesting that their formation may be a branch of the cholic acid 7 alpha-dehydroxylation pathway in this bacterium.

J Periodontol, 1991 Jan, 62(1), 74 - 81
Microbiological study of HIV-related periodontitis; Rams TE et al.; The subgingival microbiota in 14 persons with HIV-periodontitis was examined . Subgingival plaque samples were collected with paper points, transported in VMGA III, and plated on anaerobic enriched brucella blood agar and various selective media . HIV-periodontitis sites revealed Actinobacillus actinomycetemcomitans, Wolinella recta, Peptostreptococcus micros, and Bacteroides intermedius, each averaging 7% to 16% of the cultivable subgingival flora in positive patients . High levels of spirochetes also were detected in diseased sites with phase-contrast microscopy . Low levels of Candida albicans or enteric Gram-negative rods were recovered in the subgingival flora in 7 HIV-periodontitis patients or Bacteroides fragilis, Fusobacterium necrophorum, Fusobacterium varium, and Eubacterium aerofaciens were recovered in 8 patients . These findings suggest that the major components of the subgingival microbial flora in HIV-periodontitis are similar to those associated with adult periodontitis in systemically healthy persons . However, HIV-periodontitis lesions also may contain organisms which are rarely found in common types of periodontitis . The etiological significance of specific periodontal organisms in HIV-periodontitis awaits further longitudinal study.

Biofactors, 1991 Jan, 3(1), 1 - 9
Organic osmolytes in methanogenic archaebacteria; Robertson DE et al.; Methanogenic archaebacteria have developed unique ways of dealing with osmotic stress . While several of them have transport systems capable of internalizing betaine, an osmolyte in many eubacteria, in general they have developed de novo synthesis of a novel series of beta-amino acids as compatible solutes . 13C-NMR spectroscopy has been the key tool in elucidating both the identity of these organic osmolytes and in investigating their dynamics.

J Biochem (Tokyo), 1991 Jan, 109(1), 45 - 8
Kinetic studies on a novel sulfotransferase from Eubacterium A-44, a human intestinal bacterium; Kim DH et al.; A novel sulfotransferase purified from a human intestinal bacterium stoichiometrically catalyzed the transfer of a sulfate group of phenylsulfate esters to phenolic compounds . Vmax values of the enzyme reaction were measured with various concentrations of a sulfate donor substrate, p-nitrophenylsulfate, and of a sulfate acceptor substrate, tyramine . Double reciprocal plots of the acceptor concentration and Vmax showed a linear correlation . One of the reaction products, tyramine O-sulfate, competitively inhibited the enzyme as to a donor substrate, p-nitrophenylsulfate (PNS), but the other reaction product, p-nitrophenol (PNP), noncompetitively inhibited it as to PNS . These kinetic data suggest that the sulfate transfer reaction proceeds according to a ping pong bi bi mechanism . The enzyme was activated by Mg2+ and inhibited by EDTA, which suggests that it is a metalloenzyme.

J Mol Evol, 1991 Jan, 32(1), 70 - 8
Evolution of RNA polymerases and branching patterns of the three major groups of Archaebacteria; Iwabe N et al.; The amino acid sequences of the largest subunits of the RNA polymerases I, II, and III from eukaryotes were compared with those of archaebacterial and eubacterial homologs, and their evolutionary relationships were analyzed in detail by a recently developed tree-making method, the likelihood method of protein phylogeny, as well as by the neighbor-joining method and the parsimony method, together with bootstrap analyses . It was shown that the best tree topologies predicted by the first two methods are identical, whereas the last one predicts a distinct tree . The maximum likelihood tree revealed that, after the separation from archaebacteria, the three eukaryotic RNA polymerases diverged from an ancestral precursor in the eukaryotic lineage . This result is contrasted with the published result showing multiple origins for the three eukaryotic polymerases . It was shown that eukaryotic RNA polymerase I evolved much more rapidly than RNA polymerases II and III: The N-terminal half of RNA polymerase I shows an extraordinarily high evolutionary rate, possibly due to relaxed functional constraints . In contrast the evolutionary rate of archaebacterial RNA polymerase is remarkably limited . In addition, including the second largest subunit of the RNA polymerase, a detailed analysis for the branching pattern of the three major groups of archaebacteria was carried out by the maximum likelihood method . It was shown that the three major groups of archaebacteria are likely to form a single cluster; that is, archaebacteria are likely to be monophyletic as originally proposed by Woese and his colleagues.

Arch Biochem Biophys, 1991 Jan, 284(1), 22 - 5
Gene cluster rpoBC1C2 in cyanobacteria does not constitute an operon; Xie WQ et al.; The core enzyme of the cyanobacterial DNA-dependent RNA polymerase contains a unique component, gamma, which is absent from the corresponding enzymes of other eubacteria . In the heterocystous cyanobacterium Nostoc commune the gene encoding gamma, rpoC1, is immediately adjacent to, and downstream of, rpoB . The rpoC1 gene, and a 3' adjacent gene, rpoC2, correspond to the single rpoC gene found in Escherichia coli with respect to those domains conserved within their translational products . Northern analysis and primer extension assay show that in N . commune, rpoC1 and rpoC2 are transcribed separately from rpoB . The promoter of rpoC1C2 can direct the expression of a promotorless lacZ gene in E . coli . As a consequence, cyanobacterial rpo gene expression is distinct from the mode of cotranscription described for the equivalent sequences found in other eubacteria, archaebacteria, and plant chloroplasts . Also in this paper, a simple protocol for RNA isolation, which should be applicable for RNA isolation from plant cells, is presented.

Virology, 1991 Jan, 180(1), 88 - 98
Identification, sequence, and expression of the gene encoding the second-largest subunit of the vaccinia virus DNA-dependent RNA polymerase; Amegadzie BY et al.; The gene, rpo 132, encoding the second-largest subunit of the vaccinia virus DNA-dependent RNA polymerase was identified and sequenced . Two complementary approaches, involving antiserum to purified vaccinia virus RNA polymerase, were used to locate the rpo 132 gene . One method involved the screening of a lambda gt11 library of vaccinia virus genome fragments and the other was based on the immunoprecipitation and polyacrylamide gel electrophoresis of the in vitro translation products of mRNA that hybridized to immobilized vaccinia virus DNA . The deduced open reading frame of the rpo 132 gene predicted a polypeptide of 1164 amino acid residues with sequence similarities to the second-largest RNA polymerase subunits of eubacteria, archaebacteria, and eukaryotes as well as to other poxviruses . Transcriptional analyses indicated that rpo 132 has both early and late RNA start sites and is expressed throughout infection.

Rheumatol Int, 1991, 11(4-5), 203 - 8
Histology of joint inflammation induced in rats by cell wall fragments of the anaerobic intestinal bacterium Eubacterium aerofaciens; Severijnen AJ et al.; To study the arthropathic properties of human intestinal bacteria, cell wall fragments (CWF) of the anaerobic bowel bacterium Eubacterium aerofaciens were injected intraperitoneally (i.p.) in arthritis-susceptible Lewis rats . Rat paw joints were subsequently studied for histopathological changes . A persisting synovitis accompanied by marginal erosions of cartilage and bone and a marked periosteal apposition of new bone tissue were the main features of the polyarthritis induced . These results are discussed in relation to streptococcal cell wall induced arthritis and compared with histopathological findings in rheumatoid arthritis (RA) in man.

Rev Latinoam Microbiol, 1991 Jan-Mar, 33(1), 87 - 101
{Functional and evolutionary aspects of the aminoacyl-tRNA synthetases}; Silva Gonzalez E et al.; A main event in protein bioshynthesis is the esterification of the correct aminoacid to cognate tRNA catalized by the aminoacyl-tRNA synthetases . The central role of this family of enzymes in metabolism is an evidence of their ancient origin . As it is the case in many others molecules involved in protein synthesis, the emergence of the aminoacyl-tRNA synthetases appears to be a problem that is not yet solved in order to understand the origin of the genetic translation . To obtain a comprehensive view of the evolution of the relationship between each one of the twenty aminoacyl-tRNA synthetases from one organism as well as from different sources (eubacteria, archaebacteria and eukaryotes) we review the information collected from the structural and catalytic properties of these enzyme . The results allow us to establish the following relationship between aminoacyl-tRNA synthetases . On one side there is a monofiletic origin for glutamyl, glutamynil and argynil-tRNA synthetases from Escherichia coli and for valyl, leucyl, metionyl, isoleucyl and phenylalanil-tRNA synthetases from eubacterias, archaebacterias and eukaryotes . On the other side there is an evolutionary relationship between aminoacyl-tRNA synthetases of eubacteria and organelles (plastids and mitochondria) and among eukaryotes and archaebacteria.

Comp Biochem Physiol A, 1991, 98(2), 351 - 4
Identification and properties of an alpha-amylase from a strain of Eubacterium sp . isolated from the rat intestinal tract; Delahaye EP et al.; 1 . A bacterial amylase was isolated from the intestinal content of monoxenic rats inoculated with Eubacterium sp . B86 . 2 . Affinity chromatography on cross-linked starch allowed its separation from rat endogenous amylases . 3 . The bacterial enzyme was characterized by its pI, molecular weight and action pattern . It behaves as a typical endo-amylase (alpha-amylase).

Science, 1990 Dec 14, 250(4987), 1570 - 3
An ancient group I intron shared by eubacteria and chloroplasts; Kuhsel MG et al.; Introns have been found in the genomes of all major groups of organisms except eubacteria . The presence of introns in chloroplasts and mitochondria, both of which are of eubacterial origin, has been interpreted as evidence either for the recent acquisition of introns by organelles or for the loss of introns from their eubacterial progenitors . The gene for the leucine transfer RNA with a UAA anticodon {tRNALeu (UAA)} from five diverse cyanobacteria and several major groups of chloroplasts contains a single group I intron . The intron is conserved in secondary structure and primary sequence, and occupies the same position, within the UAA anticodon . The homology of the intron across chloroplasts and cyanobacteria implies that it was present in their common ancestor and that it has been maintained in their genomes for at least 1 billion years.

Science, 1990 Dec 14, 250(4987), 1566 - 70
Bacterial origin of a chloroplast intron: conserved self-splicing group I introns in cyanobacteria; Xu MQ et al.; A self-splicing group I intron has been found in the gene for a leucine transfer RNA in two species of Anabaena, a filamentous nitrogen-fixing cyanobacterium . The intron is similar to one that is found at the identical position in the same transfer RNA gene of chloroplasts of land plants . Because cyanobacteria were the progenitors of chloroplasts, it is likely that group I introns predated the endosymbiotic association of these eubacteria with eukaryotic cells.

Eur J Biochem, 1990 Dec 12, 194(2), 367 - 72
The molybdoenzyme formylmethanofuran dehydrogenase from Methanosarcina barkeri contains a pterin cofactor; Karrasch M et al.; Recently formylmethanofuran dehydrogenase from the archaebacterium Methanosarcina barkeri has been shown to be a novel molybdo-iron-sulfur protein . We report here that the enzyme contains one mol of a bound pterin cofactor/mol molybdenum, similar but not identical to the molybdopterin of milk xanthine oxidase . The two pterins, after oxidation with I2 at pH 2.5, showed identical fluorescence spectra and, after oxidation with permanganate at pH 13, yielded pterin 6-carboxylic acid . They differed, however, in their apparent molecular mass: the pterin of formylmethanofuran dehydrogenase was 400 Da larger than that of milk xanthine oxidase, a property also exhibited by the pterin cofactor of eubacterial molybdoenzymes . A homogeneous formylmethanofuran dehydrogenase preparation was used for these investigations . The enzyme, with a molecular mass of 220 kDa, contained 0.5-0.8 mol molybdenum, 0.6-0.9 mol pterin, 28 +/- 2 mol non-heme iron and 28 +/- 2 mol acid-labile sulfur/mol based on a protein determination with bicinchoninic acid . The specific activity was 175 mumol.min-1.mg-1 (kcat = 640 s-1) assayed with methylviologen (app . Km = 0.02 mM) as artificial electron acceptor . The apparent Km for formylmethanofuran was 0.02 mM.

N Engl J Med, 1990 Dec 6, 323(23), 1573 - 80
The agent of bacillary angiomatosis . An approach to the identification of uncultured pathogens; Relman DA et al.; BACKGROUND . Bacillary angiomatosis is an infectious disease causing proliferation of small blood vessels in the skin and visceral organs of patients with human immunodeficiency virus infection and other immunocompromised hosts . The agent is often visualized in tissue sections of lesions with Warthin-Starry staining, but the bacillus has not been successfully cultured or identified . This bacillus may also cause cat scratch disease . METHODS . In attempting to identify this organism, we used the polymerase chain reaction . We used oligonucleotide primers complementary to the 16S ribosomal RNA genes of eubacteria to amplify 16S ribosomal gene fragments directly from tissue samples of bacillary angiomatosis . The DNA sequence of these fragments was determined and analyzed for phylogenetic relatedness to other known organisms . Normal tissues were studied in parallel . RESULTS . Tissue from three unrelated patients with bacillary angiomatosis yielded a unique 16S gene sequence . A sequence obtained from a fourth patient with bacillary angiomatosis differed from the sequence found in the other three patients at only 4 of 241 base positions . No related 16S gene fragment was detected in the normal tissues . These 16S sequences associated with bacillary angiomatosis belong to a previously uncharacterized microorganism, most closely related to Rochalimaea quintana . CONCLUSIONS . The cause of bacillary angiomatosis is a previously uncharacterized rickettsia-like organism, closely related to R . quintana . This method for the identification of an uncultured pathogen may be applicable to other infectious diseases of unknown cause.

J Med Microbiol, 1990 Dec, 33(4), 239 - 42
Comparison of identification methods for oral asaccharolytic Eubacterium species; Wade WG et al.; Thirty one strains of oral, asaccharolytic Eubacterium spp . and the type strains of E . brachy, E . nodatum and E . timidum were subjected to three identification techniques--protein-profile analysis, determination of metabolic end-products, and the API ATB32A identification kit . Five clusters were obtained from numerical analysis of protein profiles and excellent correlations were seen with the other two methods . Protein profiles alone allowed unequivocal identification.

J Bacteriol, 1990 Dec, 172(12), 7011 - 9
Cloning and sequencing of a bile acid-inducible operon from Eubacterium sp . strain VPI 12708; Mallonee DH et al.; Two bile acid-inducible polypeptides from Eubacterium sp . strain VPI 12708 with molecular weights of 27,000 and approximately 45,000 have previously been shown to be encoded by genes residing on a 2.9-kb EcoRI fragment . We now report the cloning and sequencing of three additional overlapping DNA fragments upstream from this EcoRI fragment . Together, these four fragments contain a large segment of a bile acid-inducible operon which encodes the 27,000- and 45,000-Mr (now shown to be 47,500-Mr) polypeptides and open reading frames potentially coding for four additional polypeptides with molecular weights of 59,500, 58,000, 19,500, and 9,000 to 11,500 . A bile acid-inducible polypeptide with an apparent Mr of 23,500, as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, was purified to homogeneity, and the N-terminal amino acid sequence that was obtained matched the sequence deduced from the open reading frame coding for the 19,500-Mr polypeptide . A short DNA segment containing the 3' downstream end of the gene coding for the 47,500-Mr polypeptide was not successfully cloned but was directly sequenced from DNA fragments synthesized by polymerase chain reaction . The mRNA initiation site for the bile acid-inducible operon was shown by primer extension to be immediately upstream from the gene encoding the 58,000-Mr polypeptide . A potential promoter region upstream from the mRNA initiation site displayed significant homology with the promoter regions of previously identified bile acid-inducible genes from Eubacterium sp . strain VPI 12708 . We hypothesize that this bile acid-inducible operon codes for most of the enzymes involved in the bile acid 7 alpha-dehydroxylation pathway in this bacterium.

J Bacteriol, 1990 Dec, 172(12), 6789 - 96
Nitrogenase in the archaebacterium Methanosarcina barkeri 227; Lobo AL et al.; The discovery of nitrogen fixation in the archaebacterium Methanosarcina barkeri 227 raises questions concerning the similarity of archaebacterial nitrogenases to Mo and alternative nitrogenases in eubacteria . A scheme for achieving a 20- to 40-fold partial purification of nitrogenase components from strain 227 was developed by using protamine sulfate precipitation, followed by using a fast protein liquid chromatography apparatus operated inside an anaerobic glove box . As in eubacteria, the nitrogenase activity was resolved into two components . The component 1 analog had a molecular size of approximately 250 kDa, as estimated by gel filtration, and sodium dodecyl sulfate-polyacrylamide gels revealed two predominant bands with molecular sizes near 57 and 62 kDa, consistent with an alpha 2 beta 2 tetramer as in eubacterial component 1 proteins . For the component 2 analog, a molecular size of approximately 120 kDa was estimated by gel filtration, with a subunit molecular size near 31 kDa, indicating that the component 2 protein is a tetramer, in contrast to eubacterial component 2 proteins, which are dimers . Rates of C2H2 reduction by the nearly pure subunits were 1,000 nmol h-1 mg of protein-1, considerably lower than those for conventional Mo nitrogenases but similar to that of the non-Mo non-V nitrogenase from Azotobacter vinelandii . Strain 227 nitrogenase reduced N2 at a higher rate per electron than it reduced C2H2, also resembling the non-Mo non-V nitrogenase of A . vinelandii . Ethane was not produced from C2H2 . NH4+ concentrations as low as 10 microM caused a transient inhibition of C2H2 reduction by strain 227 cells . Antiserum against component 2 Rhodospirillum rubrum nitrogenase was found to cross-react with component 2 from strain 227, and Western immunoblots using this antiserum showed no evidence for covalent modification of component 2 . Also, extracts of strain 227 cells prepared before and after switch-off had virtually the same level of nitrogenase activity . In conclusion, the nitrogenase from strain 227 is similar in overall structure to the eubacterial nitrogenases and shows greatest similarity to alternative nitrogenases.

J Bacteriol, 1990 Dec, 172(12), 6809 - 17
Characterization of the H(+)-pumping F1F0 ATPase of Vibrio alginolyticus; Krumholz LR et al.; The F1F0 ATPase of Vibrio alginolyticus was cloned from a chromosomal lambda library . The unc operon, which contains the structural genes for the ATPase, was sequenced and shown to have a gene organization of uncIBEFHAGDC . The sequence of each subunit was compared with those of other eubacterial ATPases . The V . alginolyticus unc genes exhibited greater similarity to the Escherichia coli unc genes than to any of the other bacterial unc genes for which the sequence is available . The ATPase was expressed in an E . coli unc deletion strain, and the ATP hydrolytic activity was characterized . It has a pH optimum of 7.6 and is stimulated by the addition of Triton X-100 or any of a variety of salts . The recombinant F1F0 was purified 30.4-fold and reconstituted into proteoliposomes . This enzyme catalyzed the pumping of protons coupled to ATP hydrolysis as measured in fluorescence quenching experiments but would not pump Na+ ions under similar conditions.

Proc Natl Acad Sci U S A, 1990 Dec, 87(24), 9509 - 13
Mutational analysis of an archaebacterial promoter: essential role of a TATA box for transcription efficiency and start-site selection in vitro; Reiter WD et al.; By using a recently developed in vitro transcription assay, the 16S/23S rRNA-encoding DNA promoter from the archaebacterium Sulfolobus sp . B12 was dissected by deletion and linker substitution mutagenesis . The analysis of 5' and 3' deletion mutants defined a core promoter region between positions -38 and -2 containing all information for efficient and specific transcription . Further characterization of this region by linker substitution mutagenesis indicated two sequence elements important for promoter function--one located between positions -38 and -25 (distal promoter element) and the other one located between positions -11 and -2 (proximal promoter element) . The distal promoter element encompassed the TATA-like "box A" element located approximately 26 nucleotides upstream of the majority of transcription start sites in archaebacteria (Archaeobacteria) . All mutations within this box A motif virtually abolished promoter function . Complete inactivation of the proximal promoter element was dependent on extensive mutagenesis; this element is not conserved between archaebacterial promoters except for a high A + T content in stable RNA gene promoters from Sulfolobus . Mutants containing insertions or deletions between the distal and proximal promoter elements were only slightly affected in their transcription efficiency but displayed a shift in their major initiation site, retaining an essentially fixed distance between the distal promoter element and the transcription start site . Thus, efficient transcription and start-site selection were dependent on a conserved TATA-like sequence centered approximately 26 nucleotides upstream of the initiation site, a situation unlike that of eubacterial promoters but resembling the core structure of most eukaryotic RNA polymerase II (and some RNA polymerase III) promoters . This finding suggests a common evolutionary origin of these promoters consistent with the known similarities between archaebacterial and eukaryotic RNA polymerases.

Oral Microbiol Immunol, 1990 Dec, 5(6), 340 - 5
Antimicrobial activities of thiolactomycin against gram-negative anaerobes associated with periodontal disease . f1; Hamada S et al.; Thiolactomycin (TLM), (4R)-(2E,5E)-2,4,6-trimethyl-3-hydroxy-2,5, 7-octatriene-4-thiolide, purified from a culture filtrate of a strain of the Nocardia species, was examined for antimicrobial activities against more than 100 strains of oral and periodontally associated bacteria . Nine other commonly used antibiotics were also included for the test . We found that TLM exhibited strong and selective antimicrobial activities against Bacteroides gingivalis and other oral black-pigmented Bacteroides species that may be etiologically associated with adult periodontitis . TLM also inhibited the growth of Actinobacillus actinomycetemcomitans, but did not affect the growth of oral streptococcal species and Eubacterium species . Strains of Eikenella corrodens were moderately susceptible to TLM, while Actinomyces viscosus strains were only slightly susceptible to it . Other antibiotics used for comparison showed a broad spectrum of antimicrobial activities in general . In conclusion, TLM exhibited highly selective antimicrobial activities to black-pigmented Bacteroides species and A . actinomycetemcomitans, both of which are implicated in the pathogenesis of human periodontal disease.

FEMS Microbiol Rev, 1990 Dec, 7(3-4), 377 - 82
Formate dehydrogenase; Ferry JG; Formate is a substrate, or product, of diverse reactions catalyzed by eukaryotic organisms, eubacteria, and archaebacteria . A survey of metabolic groups reveals that formate is a common growth substrate, especially among the anaerobic eubacteria and archaebacteria . Formate also functions as an accessory reductant for the utilization of more complex substrates, and an intermediate in energy-conserving pathways . The diversity of reactions involving formate dehydrogenases is apparent in the structures of electron acceptors which include pyridine nucleotides, 5-deazaflavin, quinones, and ferredoxin . This diversity of electron acceptors is reflected in the composition of formate dehydrogenase . Studies on these enzymes have contributed to the biochemical and genetic understanding of selenium, molybdenum, tungsten, and iron in biology . The regulation of formate dehydrogenase synthesis serves as a model for understanding general principles of regulation in anaerobic organisms.

Scand J Dent Res, 1990 Dec, 98(6), 472 - 81
Hydrophobicities of human polymorphonuclear leukocytes and oral Bacteroides and Porphyromonas spp., Wolinella recta, and Eubacterium yurii with special reference to bacterial surface structures; Haapasalo M et al.; The hydrophobicities of human polymorphonuclear leukocytes (PMNLs) and Bacteroides buccae, B . oris, B . oralis, B . veroralis, B . buccalis, B . heparinolyticus, B . intermedius, B . denticola, B . loescheii, B . melaninogenicus, Porphyromonas gingivalis, P . endodontalis, Wolinella recta, and Eubacterium yurii were studied by the hexadecane method . The majority of the strains were equally or less hydrophobic than the PMNLs . Only in the case of E . yurii and the only strain of B . buccalis were all strains more hydrophobic than the PMNLs . However, some strains of B . intermedius, B . oris, B . denticola, and P . gingivalis were also more hydrophobic than the PMNLs . With the exception of B . intermedius and species with a crystalline surface protein layer (S-layer), the strains of all other species with a thick capsule were more hydrophilic than the strains with little or no extracellular polymeric material . All strains of the S-layer species were either quite hydrophilic or hydrophobic depending on the species, totally irrespective of the presence of the capsule . The results suggest that the S-layers of oral anaerobic bacteria may be important determinants of cell surface hydrophobicity.

Curr Genet, 1990 Dec, 18(6), 547 - 51
Phylogenetic analysis of the RNA polymerases of Trypanosoma brucei, with special reference to class-specific transcription; Jess W et al.; We have sequenced the genes encoding to largest subunits of the three classes of DNA-dependent RNA polymerases of Trypanosoma brucei . The nucleotide and deduced amino acid sequences were compared and aligned with the corresponding sequences of other eukaryotes . Phylogenetic relationships were subsequently calculated with a distant matrix, a bootstrapped parsimony and a maximum-likelihood method . These independent calculations resulted in trees with very similar topologies . The analyses show that all the largest subunits of T . brucei are evolutionarily distant members within each of the three RNA polymerase classes . An early separation of the trypanosomal subunits from the eukaryotic lineage might form the fundamental basis for the unusual transcription process of this species . Finally, all dendrograms show a separate ramification for the largest subunit of RNA polymerase I, II and III . RNA polymerase II and/or III form a bifurcation with the archaebacterial lineage, RNA polymerase I, however, arises separately from the eubacterial beta' lineage . This suggests that the three eukaryotic RNA polymerase classes are not simply derived by two gene duplications of an ancestral gene with subsequent differentiation.

Proc Natl Acad Sci U S A, 1990 Nov, 87(22), 8918 - 22
Differential intron loss and endosymbiotic transfer of chloroplast glyceraldehyde-3-phosphate dehydrogenase genes to the nucleus; Liaud MF et al.; Chloroplast glyceraldehyde-3-phosphate dehydrogenase (GAPDH) is composed of two different subunits, GAPA and GAPB, which are encoded in the nucleus by two related genes of eubacterial origin . In the present work the genes encoding chloroplast GAPA and GAPB from pea have been cloned and sequenced . The gene for GAPB is split by eight introns . Two introns interrupt the region encoding the transit peptide and six are found within the region encoding the mature subunit, four of which are in identical or similar positions relative to genes for cytosolic GAPDH of eukaryotic organisms . As opposed to this, the gene encoding pea GAPA has only two introns in the region encoding the mature subunit . These findings strongly support the "intron early" hypothesis and suggest that the low number of introns in the gene for chloroplast GAPA is due to differential loss of introns during the streamlining period of the chloroplast genome following the GAPB/GAPA separation . We deduce from this that eubacteria and chloroplasts contained GT-AG introns until relatively recently and that the duplication event leading to the genes encoding GAPB and GAPA and their respective transit peptides occurred in the chloroplast progenitor prior to the successive transfer and functional reintegration of these genes into the nuclear environment . These conclusions imply that GAPA/GAPB transit peptides are of eubacterial origin.

J Bacteriol, 1990 Nov, 172(11), 6568 - 72
Mini-Tn5 transposon derivatives for insertion mutagenesis, promoter probing, and chromosomal insertion of cloned DNA in gram-negative eubacteria; de Lorenzo V et al.; A collection of Tn5-derived minitransposons has been constructed that simplifies substantially the generation of insertion mutants, in vivo fusions with reporter genes, and the introduction of foreign DNA fragments into the chromosome of a variety of gram-negative bacteria, including the enteric bacteria and typical soil bacteria like Pseudomonas species . The minitransposons consist of genes specifying resistance to kanamycin, chloramphenicol, streptomycin-spectinomycin, and tetracycline as selection markers and a unique NotI cloning site flanked by 19-base-pair terminal repeat sequences of Tn5 . Further derivatives also contain lacZ, phoA, luxAB, or xylE genes devoid of their native promoters located next to the terminal repeats in an orientation that affords the generation of gene-operon fusions . The transposons are located on a R6K-based suicide delivery plasmid that provides the IS50R transposase tnp gene in cis but external to the mobile element and whose conjugal transfer to recipients is mediated by RP4 mobilization functions in the donor.

J Bacteriol, 1990 Nov, 172(11), 6557 - 67
Transposon vectors containing non-antibiotic resistance selection markers for cloning and stable chromosomal insertion of foreign genes in gram-negative bacteria; Herrero M et al.; A simple procedure for cloning and stable insertion of foreign genes into the chromosomes of gram-negative eubacteria was developed by combining in two sets of plasmids (i) the transposition features of Tn10 and Tn5; (ii) the resistances to the herbicide bialaphos, to mercuric salts and organomercurial compounds, and to arsenite, and (iii) the suicide delivery properties of the R6K-based plasmid pGP704 . The resulting constructions contained unique NotI or SfiI sites internal to either the Tn10 or the Tn5 inverted repeats . These sites were readily used for cloning DNA fragments with the help of two additional specialized cloning plasmids, pUC18Not and pUC18Sfi . The newly derived constructions could be maintained only in donor host strains that produce the R6K-specified pi protein, which is an essential replication protein for R6K and plasmids derived therefrom . Donor plasmids containing hybrid transposons were transformed into a specialized lambda pir lysogenic Escherichia coli strain with a chromosomally integrated RP4 that provided broad-host-range conjugal transfer functions . Delivery of the donor plasmids into selected host bacteria was accomplished through mating with the target strain . Transposition of the hybrid transposon from the delivered suicide plasmid to a replicon in the target cell was mediated by the cognate transposase encoded on the plasmid at a site external to the transposon . Since the transposase function was not maintained in target cells, such cells were not immune to further transposition rounds . Multiple insertions in the same strain are therefore only limited by the availability of distinct selection markers . The utility of the system was demonstrated with a kanamycin resistance gene as a model foreign insert into Pseudomonas putida and a melanin gene from Streptomyces antibioticus into Klebsiella pneumoniae . Because of the modular nature of the functional parts of the cloning vectors, they can be easily modified and further selection markers can be incorporated . The cloning system described here will be particularly useful for the construction of hybrid bacteria that stably maintain inserted genes, perhaps in competitive situations (e.g., in open systems and natural environments), and that do not carry antibiotic resistance markers characteristic of most available cloning vectors (as is currently required of live bacterial vaccines).

Cell, 1990 Oct 19, 63(2), 417 - 24
A self-splicing group I intron in the DNA polymerase gene of Bacillus subtilis bacteriophage SPO1; Goodrich-Blair H et al.; We report a self-splicing intron in bacteriophage SPO1, whose host is the gram-positive Bacillus subtilis . The intron contains all the conserved features of primary sequence and secondary structure previously described for the group IA introns of eukaryotic organelles and the gram-negative bacteriophage T4 . The SPO1 intron contains an open reading frame of 522 nucleotides . As in the T4 introns, this open reading frame begins in a region that is looped out of the secondary structure, but ends in a highly conserved region of the intron core . The exons encode SPO1 DNA polymerase, which is highly similar to E . coli DNA polymerase I . The demonstration of self-splicing introns in viruses of both gram-positive and gram-negative eubacteria lends further evidence for their early origin in evolution.

Biochemistry, 1990 Oct 16, 29(41), 9733 - 6
Ribosomal protein L35: identification in spinach chloroplasts and isolation of a cDNA clone encoding its cytoplasmic precursor; Smooker PM et al.; We describe the isolation of spinach chloroplast ribosomal protein L35 and characterization of a cDNA clone encoding its cytoplasmic precursor . This protein was only recently identified in ribosomes, but the sequences of four L35 genes have now been reported and confirm its presence in eubacteria, chloroplasts, and cyanelles . Using N-terminal sequence data, oligonucleotides were designed and a cDNA library was screened . The nucleotide sequence of the cDNA clones shows that the spinach L35 protein is encoded as a precursor of 159 residues, comprising a mature protein of 73 residues and a transit peptide of 86 residues . The cleavage site for forming the mature protein is deduced to be Thr-Val-Phe-Ala decreases Ala-Lys-Gly-Tyr . The L35 protein in the photosynthetic organelle of the protozoan Cyanophora paradoxa is encoded in the organelle DNA {Bryant & Stirewalt (1990) FEBS Lett . 259, 273-280} . The corresponding gene has not been found in the chloroplast DNA of a lower plant (liverwort) and two higher plants . Our results demonstrate that the L35 protein in a higher plant (spinach) is encoded in the nucleus . This finding, in light of the endosymbiont hypothesis, suggests an organelle to nucleus transfer of the L35 gene at the evolutionary beginnings of land plants.

J Clin Microbiol, 1990 Oct, 28(10), 2375 - 6
Eubacterium lentum ATCC 43055, a new reference strain for quality control of anaerobic susceptibility tests; Barry AL et al.; A strain of Eubacterium lentum (ATCC 43055) was selected for quality control of anaerobic susceptibility tests . Multilaboratory collaborative studies with 13 different antimicrobial agents were reviewed, and MIC control limits were proposed for agar dilution tests with the new control strain.

J Bacteriol, 1990 Oct, 172(10), 6165 - 8
Nucleotide sequence analysis of the Leptospira biflexa serovar patoc rpsL and rpsG genes; Zuerner RL et al.; The Leptospira biflexa rpsL and rpsG genes were sequenced . Although similar in many respects, proteins encoded by these L . biflexa genes had several unusual features when compared with homologous proteins of other organisms . Unlike the rpsL genes of other eubacteria, the L . biflexa rpsL gene is adjacent to a rpoC-like gene.

Antimicrob Agents Chemother, 1990 Oct, 34(10), 1908 - 14
Synthesis of active nitroguaiacol ether derivatives of streptomycin; Abad JP et al.; The synthesis, purification, and biological properties of nitroguaiacol ether derivatives of streptomycin and their corresponding radioactive reduced products were examined . These derivatives are biologically active against gram-positive and gram-negative eubacteria and they are also photoreactive because of the presence of the nitroguaiacol group in the molecule . We demonstrated that these derivatives can be used as streptomycin analogs in photoaffinity labeling of the macromolecular structures related to the mode of action of the antibiotic.

Naturwissenschaften, 1990 Oct, 77(10), 472 - 5
Comparison of the biosynthesis of the methanobacterial pseudomurein and the eubacterial murein; Hartmann E et al.; From cell extracts of the archaebacterium Methanobacterium thermoautotrophicum 11 putative precursors of the pseudomurein were isolated and characterized . On the basis of the isolated intermediates, a biosynthetic pathway of the pseudomurein is proposed . Compared to the eubacterial murein the biosynthetic stages follow different pathways as indicated by the occurrence of a nucleotide-activated disaccharide and nucleotide-activated peptides.

Int J Syst Bacteriol, 1990 Oct, 40(4), 399 - 404
5S rRNA sequences of myxobacteria and radioresistant bacteria and implications for eubacterial evolution; Van den Eynde H et al.; 5S rRNA sequences were determined for the myxobacteria Cystobacter fuscus, Myxococcus coralloides, Sorangium cellulosum, and Nannocystis exedens and for the radioresistant bacteria Deinococcus radiodurans and Deinococcus radiophilus . A dendrogram was constructed by using weighted pairwise grouping based on these and all other previously known eubacterial 5S rRNA sequences, and this dendrogram showed differences as well as similarities compared with results derived from 16S rRNA analyses . In the dendrogram, Deinococcus 5S rRNA sequences clustered with 5S rRNA sequences of the genus Thermus, as suggested by the results of 16S rRNA analyses . However, in contrast to the 16S rRNA results, the Deinococcus-Thermus cluster divided the 5S rRNA sequences of the alpha subdivision of the class Proteobacteria from the 5S rRNA sequences of the beta and gamma subgroups of the Proteobacteria . The myxobacterial 5S rRNA sequence data failed to confirm the existence of a delta subgroup of the class Proteobacteria, which was suggested by the results of 16S rRNA analyses.

J Bacteriol, 1990 Oct, 172(10), 5624 - 30
Purification and partial characterization of the DNA-dependent RNA polymerase from Rickettsia prowazekii; Ding HF et al.; The DNA-dependent RNA polymerase was purified from Rickettsia prowazekii, an obligate intracellular bacterial parasite . Because of limitation of available rickettsiae, the classical methods for isolation of the enzyme from other procaryotes were modified to purify RNA polymerase from small quantities of cells (25 mg of protein) . The subunit composition of the rickettsial RNA polymerase was typical of a eubacterial RNA polymerase . R . prowazekii had beta' (148,000 daltons), beta (142,000 daltons), sigma (85,000 daltons), and alpha (34,500 daltons) subunits as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis . The appropriate subunits of the rickettsial RNA polymerase bound to polyclonal antisera against Escherichia coli core polymerase and E . coli sigma 70 subunit in Western blots (immunoblots) . The enzyme activity was dependent on all four ribonucleoside triphosphates, Mg2+, and a DNA template . Optimal activity occurred in the presence of 10 mM MgCl2 and 50 mM NaCl . Interestingly, in striking contrast to E . coli, approximately 74% of the rickettsial RNA polymerase activity was associated with the rickettsial cell membrane at a low salt concentration (50 mM NaCl) and dissociated from the membrane at a high salt concentration (600 mM NaCl).

J Vet Diagn Invest, 1990 Oct, 2(4), 318 - 22
Evaluation of four commercial anaerobic systems for identification of Eubacterium suis; Walker RL et al.; Four commercial anaerobic systems (CASs) were evaluated for usefulness in identification of Eubacterium suis . Twelve strains were evaluated in each system in triplicate, and results were interpreted independently by 5 individuals . Statistically significant differences (P less than 0.01) due to strain variation and reader interpretation accounted for discrepancies encountered . The reactivity, repeatability, and unique profiles generated made both CAS-1 and CAS-2 suitable adjuncts for identification of E . suis when colony morphology and Gram reaction were considered . Limited reactivity in CAS-3 limited its use as an aid in identification . Variability in test observations and the large number of numerical profiles generated precluded use of CAS-4.

FEMS Microbiol Lett, 1990 Sep 15, 59(3), 241 - 6
In vivo and in vitro methylation of the elongation factor EF-Tu from Euglena gracilis chloroplast; Toledo H et al.; Based on amino acid sequence similarities between the methylated elongation factor EF-Tu from Escherichia coli and the EF-Tu from Euglena gracilis chloroplast, we predicted that the latter could also be methylated in the presence of an appropriate methyltransferase . We found that, as reported for the eubacterial homologous protein, the organellar factor could be methylated in vivo and in vitro to yield monomethyllysine.

Nucleic Acids Res, 1990 Sep 11, 18(17), 5037 - 43
The organization and evolution of transfer RNA genes in Mycoplasma capricolum; Muto A et al.; The genes for presumably all the tRNA species in Mycoplasma capricolum, a derivative of Gram-positive eubacteria, have been cloned and sequenced . There are 30 genes encoding 29 tRNA species . This number is the smallest in all the known genetic systems except for mitochondria . The sequences of 9 tRNA genes of them have been previously reported (1-3) . Twenty-two genes are organized in 5 clusters consisting of nine, five, four and two genes (2 sets), respectively . The other eight genes exist as a single transcription unit . All the tRNAs are encoded each by a single gene, except for the occurrence of two tRNA(Lys)(TTT) genes . The arrangement of tRNA genes in the 9-gene cluster, the 5-gene cluster, the 4-gene cluster and one of the 2-gene clusters reveals extensive similarity with a part of the 21-tRNA gene cluster and/or the 16-tRNA gene cluster in Bacillus subtilis, respectively . The results suggest that the present M . capricolum tRNA genes have evolved from large tRNA gene clusters in the ancestral Gram-positive bacterial genome common to M . capricolum and B . subtilis, by discarding genes for redundant as well as non-obligate tRNAs, so that all the codons may be translated by as small a number of tRNAs as possible.

Gene, 1990 Sep 1, 93(1), 73 - 8
Structure of the dnaA region of Micrococcus luteus: conservation and variations among eubacteria; Fujita MQ et al.; A phylogenetic tree constructed by 5S rRNA analysis is composed of three major branches in eubacteria: high G + C Gram+, low G + C Gram+ and Gram- {Hori and Osawa, Mol . Biol . Evol . 4 (1987) 445-472} . We have shown that the characteristic dnaA region is common among Escherichia coli (Gram-), Pseudomonas putida (Gram-), and Bacillus subtilis (low G + C Gram+) . We have now determined the structure of the dnaA region of Micrococcus luteus, as a representative of the last branch, high G + C Gram+ . The dnaA gene and at least three other genes, rnpA, rpmH and dnaN were found to be conserved in M . luteus . Large nontranslatable regions were found flanking the dnaA gene . The upstream region is conserved in the four bacteria so far examined . On the other hand, the downstream region is conserved only in Gram+ bacteria, M . luteus and B . subtilis . The consensus sequence of the DnaA box in M . luteus seems to be TTGTCCACA, in contrast to TTATCCACA of other bacteria . These results confirm our hypothesis that the dnaA region is the replication origin of the ancestral bacteria and that the essential feature of the DnaA protein and DnaA-box combination is conserved in eubacteria.

J Mol Evol, 1990 Sep, 31(3), 205 - 10
Compartmentalized isozyme genes and the origin of introns; Iwabe N et al.; Both the mouse cytosolic malate dehydrogenase gene and its mitochondrial counterpart contain eight introns, of which two are present at identical positions between the isozyme genes . The probability that the two intron positions coincide by chance between the two genes has been shown to be significantly small (= 1.3 x 10(-3), suggesting that the conservation of the intron positions has a biological significance . On the basis of a rooted phylogenetic tree inferred from a comparison of these isozymes and lactate dehydrogenases, we have shown that the origins of the conserved introns are very old, possibly going back to a date before the divergence of eubacteria, archaebacteria, and eukaryotes . In the aspartate aminotransferase isozyme genes, five of the introns are at identical places . The origins of the five conserved introns, however, are not obvious at present . It remains possible that some or all of the conserved introns have evolved after the divergence of eubacteria and eukaryotes.

J Bacteriol, 1990 Sep, 172(9), 5343 - 51
Streptomyces hygroscopicus has two glutamine synthetase genes; Kumada Y et al.; Streptomyces hygroscopicus, which produces the glutamine synthetase inhibitor phosphinothricin, possesses at least two genes (glnA and glnB) encoding distinct glutamine synthetase isoforms (GSI and GSII) . The glnB gene was cloned from S . hygroscopicus DNA by complementation in an Escherichia coli glutamine auxotrophic mutant (glnA) . glnB was subcloned in Streptomyces plasmids by insertion into pIJ486 (pMSG3) and pIJ702 (pMSG5) . Both constructions conferred resistance to the tripeptide form of phosphinothricin (bialaphos) and were able to complement a glutamine auxotrophic marker in S . coelicolor . Sodium dodecyl sulfate-polyacrylamide gel electrophoretic analysis of S . lividans(pMSG5) revealed a highly overexpressed 40-kilodalton protein . When GS was purified from this strain, it was indistinguishable in apparent molecular mass from the 40-kilodalton protein . The nucleic acid sequence of the cloned region contained an open reading frame which encoded a protein whose size, amino acid composition, and N-terminal sequence corresponded to those of the purified GS . glnB had a high G + C content and codon usage typical of streptomycete genes . A comparison of its predicted amino acid sequence with the protein data bases revealed that it encoded a GSII-type enzyme which had previously been found only in various eucaryotes (47 to 50% identity) and nodulating bacteria such as Bradyrhizobium spp . (42% identity) . glnB had only 13 to 18% identity with eubacterial GSI enzymes . Southern blot hybridization experiments showed that sequences similar to glnB were present in all of the five other Streptomyces species tested, as well as Frankia species . These results do not support the previous suggestion that GSII-type enzymes found in members of the family Rhizobiaceae represent a unique example of interkingdom gene transfer associated with symbiosis in the nodule . Instead they imply that the presence of more than one gene encoding GS may be more common among soil microorganisms than previously appreciated.

Appl Environ Microbiol, 1990 Sep, 56(9), 2858 - 65
Use of oligodeoxynucleotide signature probes for identification of physiological groups of methylotrophic bacteria; Tsien HC et al.; Oligodeoxynucleotide sequences that uniquely complemented 16S rRNAs of each group of methylotrophs were synthesized and used as hybridization probes for the identification of methylotrophic bacteria possessing the serine and ribulose monophosphate (RuMP) pathways for formaldehyde fixation . The specificity of the probes was determined by hybridizing radiolabeled probes with slot-blotted RNAs of methylotrophs and other eubacteria followed by autoradiography . The washing temperature was determined experimentally to be 50 and 52 degrees C for 9-alpha (serine pathway) and 10-gamma (RuMP pathway) probes, respectively . RNAs isolated from serine pathway methylotrophs bound to probe 9-alpha, and RNAs from RuMP pathway methylotrophs bound to probe 10-gamma . Nonmethylotrophic eubacterial RNAs did not bind to either probe . The probes were also labeled with fluorescent dyes . Cells fixed to microscope slides were hybridized with these probes, washed, and examined in a fluorescence microscope equipped with appropriate filter sets . Cells of methylotrophic bacteria possessing the serine or RuMP pathway specifically bind probes designed for each group . Samples with a mixture of cells of type I and II methanotrophs were detected and differentiated with single probes or mixed probes labeled with different fluorescent dyes, which enabled the detection of both types of cells in the same microscopic field.

FEMS Microbiol Lett, 1990 Sep 1, 59(1-2), 139 - 43
The phylogenetic position of Peptococcus niger based on 16S rRNA sequence studies; Ludwig W et al.; A 1330 base-pair fragment of a 16S rRNA gene has been amplified, cloned and sequenced . Comparison to other 16S rRNA sequences of eubacteria showed that P . niger represents a deep branch within the subdivision "Gram-positive with Gram-negative cell walls" . It is not related to peptostreptococci, representatives of this genus studied so far are more closely related to clostridia.

Biochim Biophys Acta, 1990 Aug 27, 1050(1-3), 56 - 60
Evidence for an additional archaebacterial gene cluster in Halobacterium marismortui encoding ribosomal proteins HL46e and HL30; Bergmann U et al.; A small and extremely basic ribosomal protein (HL46e) has been purified from Halobacterium marismortui using reversed-phase high-performance liquid chromatography (HPLC) . The amino acid sequence of the protein was determined by automated N-terminal and internal sequence analysis . Comparison of this sequence with other ribosomal protein sequences from eubacteria, archaebacteria and eukaryotes revealed a strong homology to SL46e from Sulfolobus solfataricus, YeaL46 from yeast and RL39 from rat . No significant sequence similarly was found to any eubacterial ribosomal protein so far known . Using a specific oligonucleotide probe the HL46e gene was identified, cloned and the nucleotide sequence including the 5'- and 3'-flanking regions were analysed . The HL46e gene is followed by the gene coding for HL30 . A putative halobacterial promoter sequence with the motive 'TTTAAA' has been localized 32 bp upstream of the HL46e gene and a putative terminator sequence localized downstream from the HL30 gene . An equivalent to this HL46e/HL30 operon is apparently not present in Escherichia coli.

Biochim Biophys Acta, 1990 Aug 27, 1050(1-3), 61 - 8
Cloning and characterization of the genes for ribosomal proteins L10 and L12 from Synechocystis Sp . PCC 6803: comparison of gene clustering pattern and protein sequence homology between cyanobacteria and chloroplasts; Sibold C et al.; The endosymbiont theory proposes that chloroplasts have originated from ancestral cyanobacteria through a process of engulfment and subsequent symbiotic adaptation . The molecular data for testing this theory have mainly been the nucleotide sequence of rRNAs and of photosystem component genes . In order to provide additional data in this area, we have isolated genomic clones of Synechocystis DNA containing the ribosomal protein gene cluster rplJL . The nucleotide sequence of this cluster and flanking regions was determined and the derived amino acid sequences were compared to the available homologous sequences from other eubacteria and chloroplasts . In Escherichia coli these two genes are part of a larger cluster, i.e., rplKAJL-rpoBC . In Synechocystis, the genes for the RNA polymerase subunit (rpoBC) are shown to be widely separated from the r-protein genes . The Synechocystis gene arrangement is similar to that in the chloroplast system, where the rpoBC1C2 and rplKAJL clusters are separated and located in two cell compartments, the chloroplast and the nucleus, respectively.

J Mol Biol, 1990 Aug 5, 214(3), 737 - 49
Three-dimensional structure of the mammalian cytoplasmic ribosome; Verschoor A et al.; A three-dimensional reconstruction of the 80 S ribosome from rabbit reticulocytes has been calculated from low-dose electron micrographs of a negatively stained single-particle specimen . At 37 A resolution, the precise orientations of the 40 S and 60 S subunits within the monosome can be discerned . The translational domain centered on the upper portion of the subunit/subunit interface is quite open, allowing considerable space between the subunits for interactions with the non-ribosomal macromolecules involved in protein synthesis . Further, the cytosolic side of the monosome is strikingly more open than the membrane-attachment side, suggesting a greater ease of communication with the cytoplasm, which would facilitate the inwards and outwards diffusion of a number of ligands . Although the 60 S subunit portion of the 80 S structure shows essentially all of the major morphological features identified for the eubacterial 50 S large subunit, it appears to possess a region of additional mass that evidently accounts for the more ellipsoidal form of the eukaryotic subunit.

Appl Environ Microbiol, 1990 Aug, 56(8), 2572 - 5
Tangential flow filtration and preliminary phylogenetic analysis of marine picoplankton; Giovannoni SJ et al.; A procedure was developed for harvesting gram quantities of microbial biomass from oligotrophic waters, when mixed populations are present in low abundance . Picoplankton from Atlantic Ocean (Hydrostation S, Sargasso Sea) and Pacific Ocean (Aloha Station) sites were collected in a three-stage process: (i) collection of seawater through an intake covered with 10-microns-pore Nytex; (ii) concentration by a tangential flow filtration device equipped with 10 ft2 (0.929 m2) of 0.1-micron-pore fluorocarbon membrane; (iii) collection of cells from concentrate by centrifugation . The overall efficiency of picoplankton recovery was at least 37% . The cellular morphotypes recovered matched those of the original population . DNA was prepared from frozen cell pellets by enzymatic digestion, solvent extraction, and isopycnic centrifugation . As indicated by the binding of kingdom-specific hybridization probes to the purified DNA, the Sargasso Sea picoplankton in this collection were largely eubacteria.

Carcinogenesis, 1990 Aug, 11(8), 1381 - 7
Biliary excretion of glutathione conjugates of 4,5-epoxy-4,5-dihydro-1- nitropyrene and 9,10-epoxy-9,10-dihydro-1-nitropyrene in rats administered 1-nitropyrene orally and their further metabolism in the intestinal tract; Kinouchi T et al.; 1-Nitropyrene (1-NP), a mutagenic and carcinogenic substance that occurs in the environment, is metabolized by reductive and oxidative pathways . 4,5-Epoxy,4,5-dihydro-1-NP (1-NP 4,5-oxide) and 9,10-epoxy-9,10-dihydro-1-NP (1-NP 9,10-oxide) are oxidatively activated metabolites of 1-NP, and react with glutathione . HPLC analysis of biliary metabolites from rats administered {3H}1-NP orally showed the presence of glutathione conjugates of 1-NP 4,5-oxide and 1-NP 9,10-oxide and their metabolites, cysteinylglycine and cysteine conjugates . During the 48 h following {3H}1-NP administration, 21.4% of the biliary metabolites were excreted as glutathione, cysteinylglycine and cysteine conjugates of 1-NP oxides . The proportions of glutathione conjugates of 1-NP 4,5-oxide and 1-NP 9,10-oxide were 2.6 and 10.4% respectively . Bile and pancreatic juice collected from normal rats metabolized glutathione conjugates of 1-NP oxides and produced the corresponding cysteinylglycine and cysteine conjugates . In particular, the gamma-glutamyltransferase activity of pancreatic juice was markedly high . When glutathione conjugates of 1-NP oxides were incubated with a cell-free extract of small intestinal contents, they decreased rapidly and cysteine conjugates increased via cysteinylglycine conjugates, indicating that pancreatic juice plays an important role in the small intestine for the metabolism of glutathione conjugates to corresponding cysteine conjugates . Although degradation activity of small intestinal contents for cysteine conjugates was very low, degradation activity of the contents of the cecum and large intestine was high and was inhibited by an inhibitor of cysteine conjugate beta-lyase (beta-lyase) activity, aminooxyacetic acid . Furthermore, the intestinal anaerobic bacteria Peptostreptococcus magnus and Eubacterium limosum showed high beta-lyase activity, suggesting that the cysteine conjugates might be further metabolized by beta-lyase of the normal bacterial flora in the lower intestine to the reactivated metabolites and reabsorbed.

J Bacteriol, 1990 Aug, 172(8), 4420 - 6
Multiple copies of a bile acid-inducible gene in Eubacterium sp . strain VPI 12708; Gopal-Srivastava R et al.; Eubacterium sp . strain VPI 12708 is an anaerobic intestinal bacterium which possesses inducible bile acid 7-dehydroxylation activity . Several new polypeptides are produced in this strain following induction with cholic acid . Genes coding for two copies of a bile acid-inducible 27,000-dalton polypeptide (baiA1 and baiA2) have been previously cloned and sequenced . We now report on a gene coding for a third copy of this 27,000-dalton polypeptide (baiA3) . The baiA3 gene has been cloned in lambda DASH on an 11.2-kilobase DNA fragment from a partial Sau3A digest of the Eubacterium DNA . DNA sequence analysis of the baiA3 gene revealed 100% homology with the baiA1 gene within the coding region of the 27,000-dalton polypeptides . The baiA2 gene shares 81% sequence identity with the other two genes at the nucleotide level . The flanking nucleotide sequences associated with the baiA1 and baiA3 genes are identical for 930 bases in the 5' direction from the initiation codon and for at least 325 bases in the 3' direction from the stop codon, including the putative promoter regions for the genes . An additional open reading frame (occupying from 621 to 648 bases, depending on the correct start codon) was found in the identical 5' regions associated with the baiA1 and baiA3 clones . The 5' sequence 930 bases upstream from the baiA1 and baiA3 genes was totally divergent . The baiA2 gene, which is part of a large bile acid-inducible operon, showed no homology with the other two genes either in the 5' or 3' direction from the polypeptide coding region, except for a 15-base-pair presumed ribosome-binding site in the 5' region . These studies strongly suggest that a gene duplication (baiA1 and baiA3) has occurred and is stably maintained in this bacterium.

FEMS Microbiol Rev, 1990 Aug, 6(4), 351 - 81
Thiosulfate, polythionates and elemental sulfur assimilation and reduction in the bacterial world; Le Faou A et al.; Among sulfur compounds, thiosulfate and polythionates are present at least transiently in many environments . These compounds have a similar chemical structure and their metabolism appears closely related . They are commonly used as energy sources for photoautotrophic or chemolithotrophic microorganisms, but their assimilation has been seldom studied and their importance in bacterial physiology is not well understood . Almost all bacterial strains are able to cleave these compounds since they possess thiosulfate sulfur transferase, thiosulfate reductase or S-sulfocysteine synthase activities . However, the role of these enzymes in the assimilation of thiosulfate or polythionates has not always been clearly established . Elemental sulfur is, on the contrary, very common in the environment . It is an energy source for sulfur-reducing eubacteria and archaebacteria and many sulfur-oxidizing archaebacteria . A phenomenon still not well understood is the 'excessive assimilatory sulfur metabolism' as observed in methanogens which perform a sulfur reduction which exceeds their anabolic needs without any apparent benefit . In heterotrophs, assimilation of elemental sulfur is seldom described and it is uncertain whether this process actually has a physiological significance . Thus, reduction of thiosulfate and elemental sulfur is a common but incompletely understood feature among bacteria . These activities could give bacteria a selective advantage, but further investigations are needed to clarify this possibility . Presence of thiosulfate, polythionates and sulfur reductase activities does not imply obligatorily that these activities play a role in thiosulfate, polythionates or sulfur assimilation as these compounds could be merely intermediates in bacterial metabolism . The possibility also exists that the assimilation of these sulfur compounds is just a side effect of an enzymatic activity with a completely different function . As long as these questions remain unanswered, our understanding of sulfur and thiosulfate metabolism will remain incomplete.

Plant Mol Biol, 1990 Aug, 15(2), 307 - 15
Evidence for a composite phylogenetic origin of the plastid genome of the brown alga Pylaiella littoralis (L.) Kjellm; Assali NE et al.; The nucleotide sequence and the 5' flanking region of the rbcL gene coding for the large subunit of ribulose bisphosphate-1,5-carboxylase/oxygenase of Pylaiella littoralis, a brown alga, has been determined and the deduced amino-acid sequence has been compared to those of various photosynthetic and chemoautotrophic Eubacteria, of a red alga and of green plastids (Euglena gracilis, green algae and higher plants) . Unlike the rbcL genes of green plastids which are more closely related to those of cyanobacteria, the P . littoralis rbcL gene is more closely related to that of a beta-purple bacterium, as was found for the rbcS gene of another chromophytic alga {Boczar et al., Proc Natl Acad Sci USA 86: 4996-4999, 1989} . Matrix data of homology between the rbcL gene of P . littoralis and the same gene of other organisms are presented . Based on our previous report, the gene coding for the 16S rRNA from P . littoralis is closely related to that of E . gracilis (Markowicz et al., Curr Genet 14: 599-608, 1988) . We suggest that the large plastid DNA molecule of P . littoralis is a phylogenetically composite genome which probably resulted from mixed endosymbiosis events, or from a horizontal transfer of DNA.

Oral Microbiol Immunol, 1990 Aug, 5(4), 195 - 201
The formation of hydrogen sulfide and methyl mercaptan by oral bacteria; Persson S et al.; The capacity to form volatile sulfur compounds was tested in bacteria isolated from subgingival microbiotas and in a representative number of reference strains . A majority of the 75 tested oral bacterial species and 7 unnamed bacterial taxa formed significant amounts of hydrogen sulfide from L-cysteine . The most active bacteria were found in the genera Peptostreptococcus, Eubacterium, Selenomonas, Centipeda, Bacteroides and Fusobacterium . Methyl mercaptan from L-methionine was formed by some members of the genera Fusobacterium, Bacteroides, Porphyromonas and Eubacterium . When incubated in serum for 7 d, the most potent producers of hydrogen sulfide were Treponema denticola and the black-pigmented species, Bacteroides intermedius, Bacteroides loescheii, Porphyromonas endodontalis and Porphyromonas gingivalis . P . endodontalis and P . gingivalis also produced significant amounts of methyl mercaptan in serum . No other volatile sulfur compound was detected in serum or in the presence of L-cysteine and L-methionine . These findings significantly increase the list of oral bacteria known to produce volatile sulfur compounds.

Biochim Biophys Acta, 1990 Jul 6, 1039(3), 343 - 7
Amino acid sequences of the ribosomal proteins HL30 and HmaL5 from the archaebacterium Halobacterium marismortui; Hatakeyama T et al.; The complete amino acid sequences of the ribosomal proteins HL30 and HmaL5 from the archaebacterium Halobacterium marismortui were determined . Protein HL30 was found to be acetylated at its N-terminal amino acid and shows homology to the eukaryotic ribosomal proteins YL34 from yeast and RL31 from rat . Protein HmaL5 was homologous to the protein L5 from Escherichia coli and Bacillus stearothermophilus as well as to YL16 from yeast . HmaL5 shows more similarities to its eukaryotic counterpart than to eubacterial ones.

J Biol Chem, 1990 Jul 5, 265(19), 10925 - 8
Side chain conjugation prevents bacterial 7-dehydroxylation of bile acids; Batta AK et al.; The effect of side chain conjugation on 7-dehydroxylation of bile acids has been investigated . C24-bile acids and their glycine and taurine conjugates and keto bile acids were incubated with pure strains of Eubacterium sp . VPI 12708 . Bile acids of the 5 alpha- or 5 beta-series with a free terminal carboxyl group and a 3 alpha, 7 alpha-dihydroxy system were very effectively 7 alpha-dehydroxylated, whereas 7 beta-hydroxy bile acids resisted 7-dehydroxylation . Oxo bile acids were metabolized at the oxygen function also . Glycine- and taurine-conjugated bile acids were neither deamidated nor 7-dehydroxylated by the bacteria . Thus, side chain conjugation prevents 7-dehydroxylation of bile acids by Eubacterium sp . VPI 12708.

Appl Environ Microbiol, 1990 Jul, 56(7), 2012 - 20
Stable carbon isotope analysis of nucleic acids to trace sources of dissolved substrates used by estuarine bacteria; Coffin RB et al.; The natural abundance of stable carbon isotopes measured in bacterial nucleic acids extracted from estuarine bacterial concentrates was used to trace sources of organic matter for bacteria in aquatic environments . The stable carbon isotope ratios of Pseudomonas aeruginosa and nucleic acids extracted from cultures resembled those of the carbon source on which bacteria were grown . The carbon isotope discrimination between the substrate and total cell carbon from bacterial cultures averaged 2.3% +/- 0.6% (n = 13) . Furthermore, the isotope discrimination between the substrate and nucleic acids extracted from bacterial cultures was 2.4% +/- 0.4% (n = 10), not significantly different from the discrimination between bacteria and the substrate . Estuarine water samples were prefiltered through 1-micron-pore-size cartridge filters . Bacterium-sized particles in the filtrates were concentrated with tangential-flow filtration and centrifugation, and nucleic acids were then extracted from these concentrates . Hybridization with 16S rRNA probes showed that approximately 90% of the nucleic acids extracted on two sample dates were of eubacterial origin . Bacteria and nucleic acids from incubation experiments using estuarine water samples enriched with dissolved organic matter from Spartina alterniflora and Cyclotella caspia had stable carbon isotope values similar to those of the substrate sources . In a survey that compared diverse estuarine environments, stable carbon isotopes of bacteria grown in incubation experiments ranged from -31.9 to -20.5% . The range in isotope values of nucleic acids extracted from indigenous bacteria from the same waters was similar, -27.9 to -20.2% . Generally, the lack of isotope discrimination between bacteria and nucleic acids that was noted in the laboratory was observed in the field.(ABSTRACT TRUNCATED AT 250 WORDS)

J Biochem Biophys Methods, 1990 Jul-Aug, 21(2), 155 - 64
Identification of bacterial periplasmic glycine betaine-binding protein after electrophoresis and affinity labeling; Talibart R et al.; Antibodies were elicited in rabbits against periplasmic proteins obtained by cold osmotic shock from the Gram-negative eubacterium Rhizobium meliloti . When analyzed by crossed immunoelectrophoresis (CIE), the periplasmic proteins gave rise to 20 distinct immunoprecipitates corresponding to the same number of bands in polyacrylamide gel electrophoresis (PAGE) under non-denaturing conditions and in SDS-PAGE . The periplasmic glycine betaine-binding protein (GB-BP) was identified by autoradiography after affinity labeling with {14C}glycine betaine in PAGE and in CIE gels . The binding proved to be quite specific to glycine betaine, since the GB-BP was not labeled by choline (a metabolic precursor of glycine betaine in Escherichia coli and Rhizobium meliloti) and 15 distinct L-amino acids, including L-proline which, like glycine betaine is also an osmoprotectant . Affinity labeling of the GB-BP with {14C}glycine betaine after protein separation by PAGE or CIE is a simple and sensitive technique permitting the GB-BP to the unambiguously detected and identified in samples of complex protein mixtures containing down to 2 micrograms of GB-BP in PAGE and only 0.2 micrograms in CIE.

J Periodontol, 1990 Jul, 61(7), 405 - 11
Effects of subgingival irrigation on bacteremia following scaling and root planing; Waki MY et al.; The purpose of this study was to determine the effects of professional subgingival irrigation, together with subsequent patient administered home marginal irrigation, on the incidence of bacteremia after scaling and root planing (Sc/RP) . A total of 60 periodontal maintenance patients were assigned to either Group 1: subgingival irrigation, with 0.12% CHX and daily marginal irrigation with 0.04% CHX; Group 2: subgingival irrigation with 0.12% CHX and daily marginal irrigation with water; Group 3: subgingival and daily marginal irrigation with water; Group 4: Non-irrigation (control) . Patients entered the study after receiving a thorough periodontal maintenance appointment including a complete examination, Sc/RP, and standard oral hygiene instruction . Blood samples were taken at the 3-month visit before and after Sc/RP . Microbiological culturing was done using the Septi-Chek system, selective and non-selective media . Results from 54 patients showed that bacteremia was detected prior to Sc/RP in 2 patients and after Sc/RP in 10 patients . No significant effect by treatment regimens on post Sc/RP bacteremia could be detected . The organisms isolated included Eubacterium lentum, Propionibacterium acnes, Streptococcus species, Neisseria species, Candida albicans, Staphylococcus species, and un-identified Gram-negative rods.

J Periodontol, 1990 Jul, 61(7), 412 - 9
Serum antibodies to periodontal bacteria; Gunsolley JC et al.; The purpose of this study was to determine how serum antibodies reactive with periodontitis-associated bacteria with relates to the diagnosis of periodontitis subjects . Study groups included localized juvenile periodontitis (LJP) subjects, severe periodontitis (SP) subjects, chronic adult periodontitis (AP) subjects, and age matched controls . Twenty-two bacterial strains, representing 18 different species most commonly found in early onset periodontitis were evaluated using serum from LJP, SP, and age matched controls . Serum IgG reactive with these organisms was determined using a radioimmunoassay (RIA) . Serum antibody reactive with 13 bacterial strains differed significantly (P less than 0.01) between the three clinical groups . Discriminate analysis revealed that antibodies reactive with 5 bacterial strains of the 13 were able to identify the clinical group to which subjects belonged 79% of the time with control subjects being correctly identified 100% of the time, LJP subjects 78% of the time, and SP subjects 60% of the time . These strains included two strains of Actinobacillus actinomycetemcomitans (Y4 and N27), Fusobacterium nucleatum (E1D1), Eubacterium brachy, and Bacteroides gingivalis . The low classification rate of SP subjects suggested heterogeneity . The SP group could be divided into three subgroups using the serological data . One subgroup, with "super" severe attachment loss, generally lacked antibody reactive with these five organisms, another subgroup was serologically similar to LJP subjects, while the third subgroup had antibodies to additional organisms . This suggests that some SP subjects may represent a more advanced form of LJP . Comparison of antibody reactivity of AP subjects with age matched controls to 23 bacterial types revealed that mean serum antibody reactivity to only Bacteroides gingivalis was higher in AP subjects.(ABSTRACT TRUNCATED AT 250 WORDS)

J Mol Evol, 1990 Jul, 31(1), 51 - 68
Compositional statistics: an improvement of evolutionary parsimony and its application to deep branches in the tree of life; Sidow A et al.; We present compositional statistics, a new method of phylogenetic inference, which is an extension of evolutionary parsimony . Compositional statistics takes account of the base composition of the compared sequences by using nucleotide positions that evolutionary parsimony ignores . It shares with evolutionary parsimony the features of rate invariance and the fundamental distinction between transitions and transversions . Of the presently available methods of phylogenetic inference, compositional statistics is based on the fewest and mildest assumptions about the mode of DNA sequence evolution . It is therefore applicable to phylogenetic studies of the most distantly related organisms or molecules . This was illustrated by analyzing conservative positions in the DNA sequences of the large subunit of RNA polymerase from three archaebacterial groups, a eubacterium, a chloroplast, and the three eukaryotic polymerases . Internally consistent results, which are in accord with our knowledge of organelle origin and archaebacterial physiology, were achieved.

J Biol Chem, 1990 Jun 25, 265(18), 10403 - 9
Site-directed mutagenesis of the conserved beta subunit tyrosine 331 of Escherichia coli ATP synthase yields catalytically active enzymes; Wise JG; The ATP synthases of eubacteria and eukaryotes possess a conserved tyrosine (beta 331) that is labeled by ATP analogs and is believed to be at the catalytic site . In this report, this tyrosine was replaced by Phe, Ser, Cys, Gly, and Ala in an attempt to determine its role in catalysis . Each of the beta 331 mutant strains assembled an ATP synthase . Membranes from the beta 331-Ser, -Cys, -Ala, or -Gly strains showed strongly attenuated ATP hydrolysis and ATP-driven proton-pumping activities . The beta 331-Phe membranes showed nearly normal ATPase and functional proton pumping . A new purification procedure yielding highly active unc+ F1 (ATPase rates greater than 1000 s-1) allowed rapid isolation of soluble F1-ATPases . Kinetic analyses of purified enzymes confirmed that the structural and functional properties of beta 331-Tyr can be substituted by Phe but not effectively by Ser, Cys, Ala, or Gly . Since all of the beta 331 mutant enzymes catalyzed measurable ATP hydrolysis, it is clear that beta 331-Tyr is not directly involved in the bond making-breaking steps of catalysis . The ability of the beta 331-Phe enzyme to rapidly bind and hydrolyze ATP, and the results with other beta 331 mutant enzymes, suggests that a residue with an aromatic character is required at this position.

J Mol Biol, 1990 Jun 20, 213(4), 811 - 8
Scanning model for translational reinitiation in eubacteria; Adhin MR et al.; Premature termination of translation in eubacteria, like Escherichia coli, often leads to reinitiation at nearby start codons . Restarts also occur in response to termination at the end of natural coding regions, where they serve to enforce translational coupling between adjacent cistrons . Here, we present a model in which the terminated but not released ribosome reaches neighboring initiation codons by lateral diffusion along the mRNA . The model is based on the finding that introduction of an additional start codon between the termination and the reinitiation site consistently obstructs ribosomes to reach the authentic restart site . Instead, the ribosome now begins protein synthesis at this newly introduced AUG codon . This ribosomal scanning-like movement is bidirectional, has a radius of action of more than 40 nucleotides in the model system used, and activates the first encountered restart site . The ribosomal reach in the upstream direction is less than in the downstream one, probably due to dislodging by elongating ribosomes . The proposed model has parallels with the scanning mechanism postulated for eukaryotic translational initiation and reinitiation.

J Bacteriol, 1990 Jun, 172(6), 3264 - 8
Isolation and characterization of the 5S rRNA gene of Leptospira interrogans; Fukunaga M et al.; The gene encoding the 5S rRNA for Leptospira interrogans serovar canicola strain Moulton was isolated and sequenced . The 5S rRNA gene occurs as a single copy within the genome and encodes a 117-nucleotide-long RNA molecule . The 5S rRNA gene is flanked at both the 5' and 3' ends by regions of A + T-rich sequences, and the 5'-flanking region contains a promoter sequence . L . interrogans has a unique and remarkable organization of the 5S rRNA gene . The 5S rRNA molecule exhibits a strong similarity to typical eubacterial 5S rRNA in terms of overall secondary structure, while the primary sequence is conserved to a lesser degree . Restriction analysis of the 5S rRNA gene indicated that the DNA sequence including the 5S rRNA gene is highly conserved in the genomes of parasitic leptospires.

J Clin Microbiol, 1990 Jun, 28(6), 1089 - 93
Detection of Borrelia burgdorferi using the polymerase chain reaction; Malloy DC et al.; Segments of the ospA gene of Borrelia burgdorferi were amplified by the polymerase chain reaction (PCR) . Oligonucleotide primers used in the reaction flank a 309-base-pair segment within the ospA gene . After 35 cycles of amplification, the product could be detected by agarose gel electrophoresis or dot hybridization with a 32P-labeled probe . This segment was amplified in all strains of B . burgdorferi tested, but it was not detected in other bacterial species . An additional primer pair which has a broad specificity for conserved 16S rRNA sequences that are present in eubacteria amplified a 215-base-pair fragment in all organisms tested . The sensitivity of PCR for the detection of B . burgdorferi in clinical samples was evaluated by seeding blood and urine specimens with B . burgdorferi and subjecting them to amplification . We were able to detect 10 organisms per ml of blood or urine, using PCR with dot hybridization detection . DNA extraction is not required for sample preparation . Blood and urine specimens were obtained from canines with clinical and serologic evidence of Lyme disease and subjected to PCR analysis . Of 17 clinical specimens from 15 animals, one blood specimen showed reactivity in the PCR.

J Bacteriol, 1990 Jun, 172(6), 2986 - 95
Multiple translational products from a Mycoplasma hyorhinis gene expressed in Escherichia coli; Notarnicola SM et al.; We analyzed protein expression from a cloned Mycoplasma hyorhinis genomic fragment that produces in Escherichia coli a set of related polypeptides of 110, 100, 65, and 55 kilodaltons from a coding region of just over 3.0 kilobases . Expression of these multiple products resulted from a mechanism operating at the translational level but not from truncation at UGA termination codons, which are known to encode tryptophan in several mycoplasma species . The structural relatedness of the proteins was demonstrated by two-dimensional tryptic peptic mapping, but their generation by posttranslational processing was ruled out by pulse-chase labeling analysis . Examination of proteins expressed from plasmid constructs and tryptic peptide analysis of these polypeptides and the original set of proteins revealed that they share carboxy-terminal regions, an observation inconsistent with truncation at UGA codons . Expression of proteins from this cloned fragment was not dependent on vector sequences and was observed when the coding region was placed under control of a T7 promoter, suggesting that all products were translated from a single message . Expression of related products in mycoplasmas was examined by immunoblot analysis of M . hyorhinis proteins with antiserum against overexpressed recombinant proteins . A single 115-kilodalton mycoplasma protein was detected, which is larger than any of the related proteins expressed in E . coli . Our analysis indicated that translation initiation sites are used in E . coli that are not active in mycoplasmas, thereby defining differences between the translational regulatory signals of mycoplasmas and eubacteria.

J Bacteriol, 1990 Jun, 172(6), 2955 - 61
Gene fusions with lacZ by duplication insertion in the radioresistant bacterium Deinococcus radiodurans; Lennon E et al.; Deinococcus radiodurans is the most-studied species of a eubacterial family characterized by extreme resistance to DNA damage . We have focused on developing molecular biological techniques to investigate the genetics of this organism . We report construction of lacZ gene fusions by a method involving both in vitro splicing and the natural transformation of D . radiodurans . Numerous fusion strains were identified by expression of beta-galactosidase . Among these fusion strains, several were inducible by exposure to the DNA-damaging agent mitomycin C, and four of the inducible fusion constructs were cloned in Escherichia coli . Hybridization studies indicate that one of the damage-inducible genes contains a sequence reiterated throughout the D . radiodurans chromosome . Survival measurements show that two of the fusion strains have increased sensitivity to mitomycin C, suggesting that the fusions within these strains inactivate repair functions.

Mol Gen Genet, 1990 Jun, 222(1), 129 - 34
Five transfer RNA genes lacking CCA termini are clustered in the chromosome of Streptomyces rimosus; Plohl M et al.; The nucleotide sequence of a 1105 bp Streptomyces rimosus DNA fragment containing five transfer RNA genes was determined . Two tRNA(Gln) (CUG) genes, differing by 1 bp in the aminoacyl stem, and three identical tRNA(Glu) (CUC) genes were identified . The five tRNA genes, arranged in the order: Gln1-Glu1-Glu2-Gln2-Glu3, were separated by short, nonhomologous intergenic regions . Surprisingly, none of these tRNA genes encoded the CCA 3' terminus of mature tRNAs . All five encoded tRNAs for the translation of GC rich codons, which are preferentially used in Streptomyces genes (CAG and GAG, respectively) . We recently reported nucleotide sequences of two initiator tRNA genes from S . rimosus, which also do not encode the CCA end of mature tRNAs . It is therefore very likely that S . rimosus represents an example of those eubacteria in which the majority of tRNA genes do not encode the 3' terminal CCA end of mature tRNAs . Evolutionary implications of this finding remain to be elucidated.

FEBS Lett, 1990 May 21, 264(2), 218 - 22
McrI: a novel class-II restriction endonuclease from Micrococcus cryophilus recognizing 5'-CGRY/CG-3'; Brensing-Kuppers J et al.; A new class-II restriction endonuclease, McrI, with a novel sequence specificity as isolated from the Gram-positive eubacterium Micrococcus cryophilus . McrI recognizes the palindromic hexanucleotide sequence . {sequence: see text} The novel enzyme in the presence of Mg2(+)-ions cleaves specifically both strands as indicated by the arrows . The staggered cuts generate 3'-protruding ends with single-stranded 5'-RY-3' dinucleotide extensions . The McrI recognition sequence was deduced from mapping data on DNAs of bacteriophages theta X174RF and M13mp18RF characterized by one and four cleavage sites, respectively . The cut positions within both strands of the recognition sequence were determined in sequencing experiments by analyzing hydrolysis of phosphodiester bonds within a polylinker region of M13mp18RF DNA containing an additional McrI recognition site including treatment with T4 DNA polymerase . The novel enzyme may be a useful tool for cloning experiments by completion of the enzymes EclXI (5'-C/GGCCG-3'), NotI (5'-GC/GGCCGC-3'), PvuI (5'-CGAT/CG-3') as well as EaeI (5'-Y/GGCCR-3') and XhoII (5'-Y/GATCR-3') characterized by partly identical sequence specificities.

Nature, 1990 May 17, 345(6272), 268 - 70
The gain of two chloroplast tRNA introns marks the green algal ancestors of land plants; Manhart JR et al.; The relationship of green algae to land plants has greatly interested botanists for more than a century . In recent years, several characters, particularly ultrastructural ones, have been used to define a green algal group (Charophyceae) from which land plants are thought to have arisen . Here we provide the first molecular genetic evidence in support of the charophycean origin of land plants . Group II introns have previously been found in both the tRNAAla and tRNAIle genes of all land plant chloroplast DNAs examined, whereas all algae and eubacteria examined have uninterrupted genes . The distribution of these introns in Coleochaete, Nitella and Spirogyra, members of the Charophyceae, confirms that these taxa are part of the lineage that gave rise to land plants . Furthermore, the intron data place Coleochaete and Nitella closer to land plants than Spirogyra . These introns were most probably acquired by the chloroplast genome more than 400-500 million years ago, the time of land plant origin.

Biochim Biophys Acta, 1990 May 8, 1038(3), 346 - 54
Conformational study of the chromosomal protein MC1 from the archaebacterium Methanosarcina barkeri; Imbert M et al.; Methanogen chromosomal protein MC1 is a polypeptide of 93 amino acid residues (Mr 10,757) which represents the major protein associated with the DNA of the archaebacterium Methanosarcina barkeri and can protect DNA against thermal denaturation . The conformation of protein MC1 has been investigated by means of predictive methods, infrared spectroscopy, circular dichroism and tryptophan fluorescence studies . Protein MC1 has a low amount of alpha-helix but contains antiparallel beta-sheet strands . The larger hydrophobic cluster which contains tryptophan at position 61 appears buried in the protein . Addition of salts induces the unfolding of the protein and makes the tryptophan indole ring more rigid . With respect to its primary structure and its conformation, protein MC1 appears radically different from the chromosomal DNA-binding protein II (also called HU-type protein) in eubacteria.

Nature, 1990 May 3, 345(6270), 60 - 3
Genetic diversity in Sargasso Sea bacterioplankton; Giovannoni SJ et al.; Bacterioplankton are recognized as important agents of biogeochemical change in marine ecosystems, yet relatively little is known about the species that make up these communities . Uncertainties about the genetic structure and diversity of natural bacterioplankton populations stem from the traditional difficulties associated with microbial cultivation techniques . Discrepancies between direct counts and plate counts are typically several orders of magnitude, raising doubts as to whether cultivated marine bacteria are actually representative of dominant planktonic species . We have phylogenetically analysed clone libraries of eubacterial 16S ribosomal RNA genes amplified from natural populations of Sargasso Sea picoplankton by the polymerase chain reaction . The analysis indicates the presence of a novel microbial group, the SAR11 cluster, which appears to be a significant component of this oligotrophic bacterioplankton community . A second cluster of lineages related to the oxygenic phototrophs--cyanobacteria, prochlorophytes and chloroplasts--was also observed . However, none of the genes matched the small subunit rRNA sequences of cultivated marine cyanobacteria from similar habitats . The diversity of 16S rRNA genes observed within the clusters suggests that these bacterioplankton may be consortia of independent lineages sharing surprisingly distant common ancestors.

J Clin Periodontol, 1990 May, 17(5), 288 - 91
Serum antibodies and loss of periodontal bone in patients with rheumatoid arthritis; Tolo K et al.; The number of teeth, % of alveolar bone loss, serum IgG, and serum antibodies to Bacteroides gingivalis, Capnocytophaga ochracea and Eubacterium saburreum were recorded in 37 patients diagnosed with rheumatoid arthritis (RA) and in an age- and sex-matched control group of 37 individuals free from RA . The RA group had a significantly increased loss of teeth and loss of alveolar bone compared to the control group . The RA patients also had a significantly increased level of serum IgG . In the total material, 26% of the variation in loss of alveolar bone was accounted for by age, diagnosis of rheumatoid arthritis, and levels of antibodies against B . gingivalis and E . saburreum . In the RA group, 48% of this variation was accounted for by age, total serum IgG and IgG antibodies to B . gingivalis and E . saburreum.

Mol Gen Genet, 1990 May, 221(3), 315 - 21
Structure of the archaebacterial 7S RNA molecule; Kaine BP; The genes encoding the 7S RNAs of the archaebacteria Archaeoglobus fulgidus, Methanosarcina acetivorans, Sulfolobus, solfataricus, and Thermococcus celer have been isolated . All four genes occur as single genomic copies and are flanked by sequences containing potential signals for transcriptional promotion and termination . The genes encode RNA molecules approximately 300 nucleotides in length which conform strictly to a model of secondary structure common to all described archaebacterial 7S RNAs . Archaebacterial 7S RNAs exhibit a strong similarity to eukaryotic 7S RNAs in terms of overall secondary structure, while primary sequence conservation is limited to a specific structural domain of the molecule . This domain displays strong primary and secondary structural similarity to features of small eubacterial RNAs, including the small cytoplasmic (sc) RNA of Bacillus subtilis and the 4.5S RNA of Escherichia coli . Conservation of this structural domain among divergent RNA molecules across three kingdoms suggests that these RNAs are the descendants of a unique subcellular structure present before the divergence of the archaebacterial, eubacterial and eukaryotic kingdoms.

J Mol Evol, 1990 May, 30(5), 463 - 76
Small ribosomal subunit RNA sequences, evolutionary relationships among different life forms, and mitochondrial origins; Van de Peer Y et al.; A tree was constructed from a structurally conserved area in an alignment of 83 small ribosomal subunit sequences of eukaryotic, archaebacterial, eubacterial, plastidial, and mitochondrial origin . The algorithm involved computation and optimization of a dissimilarity matrix . According to the tree, only plant mitochondria belong to the eubacterial primary kingdom, whereas animal, fungal, algal, and ciliate mitochondria branch off from an internal node situated between the tree primary kingdoms . This result is at variance with a parsimony tree of similar size published by Cedergren et al . (J Mol Evol 28:98-112, 1988), which postulates the mitochondria to be monophyletic and to belong to the eubacterial primary kingdom . The discrepancy does not follow from the use of conflicting sequence alignments, hence it must be due to the use of different treeing algorithms . We tested our algorithm on a set of sequences resulting from a simulated evolution and found it capable of faithfully reconstructing a branching topology that involved very unequal evolutionary rates . The use of more limited or more extended areas of the complete sequence alignment, comprising only very conserved or also more variable portions of the small ribosomal subunit structure, does have some influence on the tree topology . In all cases, however, the nonplant mitochondria seem to branch off before the emergence of eubacteria, and the differences are limited to the branching pattern among different types of mitochondria.

J Bacteriol, 1990 May, 172(5), 2447 - 55
A polymerase chain reaction-based approach to cloning sigma factors from eubacteria and its application to the isolation of a sigma-70 homolog from Chlamydia trachomatis; Engel JN et al.; Taking advantage of the known sequence conservation of portions of bacterial sigma factor proteins, we have designed degenerate oligonucleotides corresponding to these domains and used these synthetic DNA sequences as primers in a polymerase chain reaction (PCR) to amplify DNA sequences from the chlamydial genome . The PCR products were used as a probe to recover the genomic fragments from a library of cloned murine Chlamydia trachomatis DNA . Sequence analysis of one of these clones revealed striking homology to the sigma-70 protein of Escherichia coli and the sigma-43 protein of Bacillus subtilis, strongly implying that this locus (sigA) encodes the major vegetative sigma factor of murine C . trachomatis . This PCR-based approach will be broadly applicable to the cloning of major sigma factors from other eubacteria.

J Struct Biol, 1990 May, 103(3), 197 - 203
Electron microscopy and image analysis reveal common principles of organization in two large protein complexes: groEL-type proteins and proteasomes; Zwickl P et al.; In an attempt to settle the question of whether the multicatalytic proteinase or proteasome exist in all three kingdoms of life--eukaryotes, archaebacteria, and eubacteria--we have undertaken a search for them in the eubacterium Comamonas acidovorans . We have, in fact, isolated and purified a cylinder-shaped particle . However, according to various structural and biochemical criteria this turned out to be more reminiscent of the groEL protein from Escherichia coli and its homologs than to proteasomes of eukaryotic or archaebacterial origin . N-terminal sequencing provided definite proof for its belonging to this family of molecular chaperonins . Image analysis of electron micrographs revealed that the C . acidovorans groEL-like protein and proteasomes in spite of their significantly different dimensions have certain principles of organization in common.

Mol Biol Evol, 1990 May, 7(3), 255 - 69
Designer invariants for large phylogenies; Sankoff D; The Cavender-Felsenstein edge-length invariants for binary characters on 4-trees provide the starting point for the development of "customized" invariants for evaluating and comparing phylogenetic hypotheses . The binary character invariants may be generalized to k-valued characters without losing the quadratic nature of the invariants as functions of the theoretical frequencies f(UVXY) of observable character configurations (U at organism 1, V at 2, etc.) . The key to the approach is that certain sets of these configurations constitute events which are probabilistically independent from other such sets, under the symmetric Markov change models studied . By introducing more complex sets of configurations, we find the quadratic invariants for 5-trees in the binary model and for individual edges in 6-trees or, indeed, in any size tree . The same technique allows us to formulate invariants for entire trees, but these are cubic functions for 6-trees and are higher-degree polynomials for larger trees . With k-valued characters and, especially, with large trees, the types of configuration sets (events) used in the simpler examples are too rare (i.e., their predicted frequencies are too low) to be useful, and the construction of meaningful pairs of independent events becomes an important and nontrivial task in designing invariants suited to testing specific hypotheses . In a very natural way, this approach fits in with well-known statistical methodology for contingency tables . We explore use of events such as "only transitions occur for character i (i.e., position i in a nucleic acid sequence) in subtree a" in analyzing a set of data on ribosomal RNA in the context of the controversy over the origins of archaebacteria, eubacteria, and eukaryotes.

Biochim Biophys Acta, 1990 Apr 26, 1016(3), 371 - 7
The F1-type ATPase in anaerobic Lactobacillus casei; Muntyan MS et al.; An ATPase from anaerobic Lactobacillus casei has been isolated and 100-times purified . The 400 kDa enzyme molecule was found to have a hexagonal structure 10 nm in diameter composed of at least six protein masses . SDS-electrophoresis reveals four or, under certain conditions, five types of subunit, of apparent molecular masses 57 (alpha), 55 (beta), 40 (gamma), 22 (delta) and 14 (epsilon) kDa with stoichiometry of 3 alpha, 3 beta, gamma, delta, epsilon . The following features resembling F1-ATPases from other sources were found to be inherent in the solubilized L . casei ATPase . (i) Detachment from the membrane desensitizes ATPase to low DCCD concentrations and sensitizes it to water-soluble carbodiimide . (ii) Soluble ATPase is inhibited by Nbf chloride and azide, is resistant to SH-modifiers and is activated by sulfite and octyl glucoside, the activating effect being much stronger than in the case of the membrane-bound ATPase . Substrate specificity of the enzyme is also similar to that of other factors F1 . Divalent cations strongly activate the soluble enzyme when added at a concentration equal to that of ATP . An excess of Mn2+, Mg2+ or Co2+ inhibits ATPase activity of F1, whereas that of Ca2+ induces its further activation . No other F1-like ATPases are found in L . casei . It is concluded that this anaerobic bacterium possesses a typical F1-ATPase similar to those in mitochondria, chloroplasts, aerobic and photosynthetic eubacteria.

Nucleic Acids Res, 1990 Apr 25, 18 Suppl, 2215 - 30
Compilation of 5S rRNA and 5S rRNA gene sequences; Specht T et al.; The BERLIN RNA DATABANK as of December 31, 1989, contains a total of 667 sequences of 5S rRNAs or their genes, which is an increase of 114 new sequence entries over the last compilation (1) . It covers sequences from 44 archaebacteria, 267 eubacteria, 20 plastids, 6 mitochondria, 319 eukaryotes and 11 eukaryotic pseudogenes . The hardcopy shows only the list of those organisms whose sequences have been determined . 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.

J Biol Chem, 1990 Apr 15, 265(11), 6436 - 40
Overexpression of the methanococcal ribosomal protein L12 in Escherichia coli and its incorporation into halobacterial 50 S subunits yielding active ribosomes; Kopke AK et al.; The gene for the ribosomal L12 protein from the archaebacterium Methanococcus vannielii was cloned into the expression vector pKK223-3 . The protein was overexpressed and remained stable in Escherichia coli XL1 cells . Purification yielded a protein with the same amino acid composition and sequence as in Methanococcus but it was acetylated at the N terminus as in the case with the homologous protein of E . coli . The in vivo incorporation of the overexpressed protein into the E . coli ribosomes was not observed . The overexpressed M . vannielii protein MvaL12e was incorporated into halobacterial ribosomes, thereby displacing the corresponding halobacterial L12 protein . Intact 70 S ribosomes were reconstituted from halobacterial 50 S subunits carrying the MvaL12e protein . These ribosomes were as active as native halobacterial ribosomes in a poly(U) assay . On the other hand, our attempts to incorporate L12 proteins from Bacillus stearothermophilus and E . coli into halobacterial ribosomes were not successful . These results support the conclusion which is based on primary sequence and predicted secondary structure comparisons that there exist two distinct L12 protein families, namely the eubacterial L12 protein family and the eukaryotic/archaebacterial L12 protein family.

J Bacteriol, 1990 Apr, 172(4), 1823 - 7
Heat shock response in mycoplasmas, genome-limited organisms; Dascher CC et al.; We have measured the effect of heat shock on three mycoplasmas (Acholeplasma laidlawii K2 and JA1 and Mycoplasma capricolum Kid) and demonstrated the induction of mycoplasma heat shock proteins under these conditions . Increased synthesis of at least 5 heat shock proteins in A . laidlawii K2, 11 heat shock proteins in A . laidlawii JA1, and 7 heat shock proteins in M . capricolum was observed by electrophoretic analysis of proteins from heat-shocked cells in sodium dodecyl sulfate-polyacrylamide gels . In all three strains, major heat shock proteins (66 to 68 and 26 to 29 kilodaltons {kDa}) were found . The 66- to 68-kDa protein cross-reacted with antibody to Escherichia coli DnaK protein, suggesting that this heat shock protein has been conserved in spite of major reductions in genetic complexity during mycoplasma evolution . A . laidlawii also contained a 60-kDa protein that cross-reacted with eubacterial GroEL protein and a 40-kDa protein that cross-reacted with E . coli RecA protein . Unlike with coliphages, the mycoplasma virus L2 progeny yield was not increased when virus was plated on heat-shocked A . laidlawii host cells . However, UV-irradiated L2 virus could be host cell reactivated by both A . laidlawii SOS repair and heat shock systems.

J Gen Microbiol, 1990 Apr, 136 ( Pt 4), 651 - 6
Cloning of genes required for amino acid biosynthesis from Leptospira interrogans serovar icterohaemorrhagiae; Richaud C et al.; Leptospira interrogans belongs to a large family of important pathogens, which is part of the order Spirochaetales, a distinct group of eubacteria . In order to obtain a better understanding of the genetic organization of this species, we have constructed a DNA library of the serovar icterohaemorrhagiae, using the Escherichia coli vector pUC13 . We have isolated Leptospira DNA fragments containing the genetic information required to complement strains of E . coli with defects in proline and leucine biosynthesis . While a 3.9 kb fragment which complemented proA also complemented proB, a 15 kb fragment complementing leuB could not complement other leu mutations . The L . interrogans origin of the cloned DNA fragments was confirmed by DNA-DNA hybridization . The hydridization was specific to the pathogenic species and was not seen with the saprophytic species L . biflexa.

Mol Gen Genet, 1990 Apr, 221(2), 273 - 9
Construction of an integration vector for use in the archaebacterium Methanococcus voltae and expression of a eubacterial resistance gene; Gernhardt P et al.; An integration vector for use in Methanococcus voltae was constructed, based on the Escherichia coli vector pUC18 . It carries the structural gene for puromycin transacetylase from Streptomyces alboniger, which is flanked by expression signals of M . voltae structural genes and hisA gene sequences of this bacterium . Transformed M . voltae cells are puromycin resistant . Several types of integration of the vector into the chromosome were found . Only one case was due to nonhomologous recombination . The integrated sequences were stable under selective pressure but were slowly lost in some cases in the absence of the selective drug . The vector could be excised from M . voltae chromosomal DNA, recircularized and transformed back into E . coli.

J Bacteriol, 1990 Apr, 172(4), 2150 - 9
Effect of chemical fixatives on accurate preservation of Escherichia coli and Bacillus subtilis structure in cells prepared by freeze-substitution; Graham LL et al.; Five chemical fixatives were evaluated for their ability to accurately preserve bacterial ultrastructure during freeze-substitution of select Escherichia coli and Bacillus subtilis strains . Radioisotopes were specifically incorporated into the peptidoglycan, lipopolysaccharide, and nucleic acids of E . coli SFK11 and W7 and into the peptidoglycan and RNA of B . subtilis 168 and W23 . The ease of extraction of radiolabels, as assessed by liquid scintillation counting during all stages of processing for freeze-substitution, was used as an indicator of cell structural integrity and retention of cellular chemical composition . Subsequent visual examination by electron microscopy was used to confirm ultrastructural conformation . The fixatives used were: 2% (wt/vol) osmium tetroxide and 2% (wt/vol) uranyl acetate; 2% (vol/vol) glutaraldehyde and 2% (wt/vol) uranyl acetate; 2% (vol/vol) acrolein and 2% (wt/vol) uranyl acetate; 2% (wt/vol) gallic acid; and 2% (wt/vol) uranyl acetate . All fixatives were prepared in a substitution solvent of anhydrous acetone . Extraction of cellular constituents depended on the chemical fixative used . A combination of 2% osmium tetroxide-2% uranyl acetate or 2% gallic acid alone resulted in optimum fixation as ascertained by least extraction of radiolabels . In both gram-positive and gram-negative organisms, high levels of radiolabel were detected in the processing fluids in which 2% acrolein-2% uranyl acetate, 2% glutaraldehyde-2% uranyl acetate, or 2% uranyl acetate alone were used as fixatives . Ultrastructural variations were observed in cells freeze-substituted in the presence of different chemical fixatives . We recommend the use of osmium tetroxide and uranyl acetate in acetone for routine freeze-substitution of eubacteria, while gallic acid is recommended for use when microanalytical processing necessitates the omission of osmium.

J Bacteriol, 1990 Apr, 172(4), 2141 - 9
Evaluation of freeze-substitution and conventional embedding protocols for routine electron microscopic processing of eubacteria; Graham LL et al.; Freeze-substitution and more conventional embedding protocols were evaluated for their accurate preservation of eubacterial ultrastructure . Radioisotopes were specifically incorporated into the RNA, DNA, peptidoglycan, and lipopolysaccharide of two isogenic derivatives of Escherichia coli K-12 as representative gram-negative eubacteria and into the RNA and peptidoglycan of Bacillus subtilis strains 168 and W23 as representative gram-positive eubacteria . Radiolabeled bacteria were processed for electron microscopy by conventional methods with glutaraldehyde fixation, osmium tetroxide postfixation, dehydration in either a graded acetone or ethanol series, and infiltration in either Spurr or Epon 812 resin . A second set of cells were simultaneously freeze-substituted by plunge-freezing in liquid propane, substituting in anhydrous acetone containing 2% (wt/vol) osmium tetroxide, and 2% (wt/vol) uranyl acetate, and infiltrating in Epon 812 . Extraction of radiolabeled cell components was monitored by liquid scintillation counting at all stages of processing to indicate retention of cell labels . Electron microscopy was also used to visually confirm ultrastructural integrity . Radiolabeled nucleic acid and wall components were extracted by both methods . In conventionally embedded specimens, dehydration was particularly damaging, with ethanol-dehydrated cells losing significantly more radiolabeled material during dehydration and subsequent infiltration than acetone-treated cells . For freeze-substituted specimens, postsubstitution washes in acetone were the most deleterious step for gram-negative cells, while infiltration was more damaging for gram-positive cells . Autoradiographs of specimens collected during freeze-substitution were scanned with an optical densitometer to provide an indication of freezing damage; the majority of label lost from freeze-substituted cells was a result of poor freezing to approximately one-half of the cell population, thus accounting for the relatively high levels of radiolabel detected in the processing fluids . These experiments revealed that gram-positive and gram-negative cells respond differently to freezing; these differences are discussed with reference to wall structure . It was apparent that the cells frozen first (ie., the first to contact the cryogen) retained the highest percentage of all radioisotopes, and the highest level of cellular infrastructure, indicative of better preservation . The preservation of these select cells was far superior to that obtained by more conventional techniques.

Nucleic Acids Res, 1990 Mar 25, 18(6), 1565 - 9
Role of GC-biased mutation pressure on synonymous codon choice in Micrococcus luteus, a bacterium with a high genomic GC-content; Ohama T et al.; The GC (G + C, or G or C)-contents of codon silent positions in all two-codon sets and three codons AUY/A (IIe), and in most of the family boxes of Micrococcus luteus (genomic GC-content: 74%) are 95% to 100% in both the highly and weakly expressed genes . In some family boxes, there is a decrease in NNC codons and an increase in NNG codons from the highly expressed to weakly expressed genes without apparent involvement of NNU and NNA codons . From these observations, we conclude that the selective use of synonymous codons in M . luteus may be largely determined by GC-biased mutation pressure and that in the highly expressed genes tRNAs would act as a weak selection pressure in some family boxes . Available data suggest that the effect of selection pressure by tRNAs on the synonymous codon choice becomes more apparent in the highly expressed genes in eubacteria with intermediate GC-contents such as Escherichia coli and Bacillus subtilis, and that the U/C ratio of the codon third positions in NNU/C-type two-codon sets in the weakly expressed genes would represent the approximate magnitude of directional mutation pressure throughout eubacteria.

Nature, 1990 Mar 15, 344(6263), 262 - 5
Evolutionary transfer of the chloroplast tufA gene to the nucleus; Baldauf SL et al.; Evolutionary gene transfer is a basic corollary of the now widely accepted endosymbiotic theory, which proposes that mitochondria and chloroplasts originated from once free-living eubacteria . The small organellar chromosomes are remnants of larger bacterial genomes, with most endosymbiont genes having been either transferred to the nucleus soon after endosymbiosis or lost entirely, with some being functionally replaced by pre-existing nuclear genes . Several lines of evidence indicate that relocation of some organelle genes could have been more recent . These include the abundance of non-functional organelle sequences of recent origin in nuclear DNA, successful artificial transfer of functional organelle genes to the nucleus, and several examples of recently lost organelle genes, although none of these is known to have been replaced by a nuclear homologue that is clearly of organellar ancestry . We present gene sequence and molecular phylogenetic evidence for the transfer of the chloroplast tufA gene to the nucleus in the green algal ancestor of land plants.

Experientia, 1990 Mar 15, 46(3), 274 - 6
Widespread occurrence of 2-acetylthiazole-4-carboxylic acid in biological material; White RH; 2-Acetylthiazole-4-carboxylic acid was shown to be widely distributed in all organisms tested, which included members of the eukaryotes, archaebacteria, and eubacteria . This thiazole, which was identified and quantitated as the methyl ester methoxyamine derivative, was found in these organisms at levels of from 27 to 1100 nmol/g dry weight (d.wt) of tissue . On the basis of its widespread occurrence, the levels at which it occurs in these organisms, and its chemical structure, which contains a reactive carbonyl group, it is proposed that this compound is a previously undescribed coenzyme.

Eur J Biochem, 1990 Mar 10, 188(2), 219 - 29
Evolution of large-subunit rRNA structure . The diversification of divergent D3 domain among major phylogenetic groups; Michot B et al.; During evolution, the potential for sequence (and length) variation of large-subunit rRNA has been mostly restricted over 12 divergent domains (termed D1-D12) interspersed along the molecule . Here, we have focused our attention onto the D3 divergent domain, through a detailed analysis of its pattern of variation in the phylogeny, both in terms of primary and secondary structures . We have systematically compared all the procaryotic and eucaryotic sequences published so far (i.e . 36 species), together with a series of 10 additional eucaryotic specimens, which were determined by direct RNA sequencing . Secondary structures supported by comparative evidence have been derived for archaebacteria, eubacteria and eucaryotes respectively, which shows that the D3 domain contains a subset of universally conserved structural features interspersed with four variable subdomains . Within the four portions where a structural diversification has taken place, elementary structures specific of large phylogenetic groups can be identified . Remarkably such diversified structures appear to be preserved despite sequence divergence, suggesting they correspond to functionally important structures . Accordingly, the mode of sequence variation of the D3 domain suggests this region of the molecule may encode elementary functions of rRNA which could have significantly diversified during the evolution of the major groups of organisms.

Int Endod J, 1990 Mar, 23(2), 100 - 6
The microbial flora from root canals and periodontal pockets of non-vital teeth associated with advanced periodontitis; Kobayashi T et al.; Microflora from root canals and periodontal pockets of periodontally affected teeth were compared in order to elucidate the as yet unknown relationship between pulpal and periodontal disease . Caries-free teeth affected with advanced periodontitis and diagnosed as clinically dead by electric pulp testing were selected . The root canals and periodontal pockets were sampled, and the bacterial flora examined by both culture and interference microscopy . The results indicated that the aerobe/anaerobe ratio in the periodontal pocket was 0.23, while it was 0.0022 in the root canal, the large predominance of obligate anaerobes reflecting the anaerobic environment found in the root canal . Morphological classification obtained from interference microscopy showed similar proportions of morphotypes in the two sites . Results of anaerobic culture demonstrated a significantly higher rate of detection of facultative Streptococcus bacteria in the periodontal pocket than in the root canal . The predominant bacterial species common to both regions were Streptococcus, Peptostreptococcus, Eubacterium, Bacteroides, and Fusobacterium for obligate anaerobes . As for facultative anaerobes, Actinomyces and Streptococcus were detected predominantly in the periodontal pocket . The occurrence of micro-organisms common to both sites in this study suggests that the periodontal pocket may be a possible source of root canal infections.

J Bacteriol, 1990 Mar, 172(3), 1284 - 8
Escherichia coli 4.5S RNA gene function can be complemented by heterologous bacterial RNA genes; Struck JC et al.; The essential 4.5S RNA gene of Escherichia coli can be complemented by 4.5S RNA-like genes from three other eubacteria, including both gram-positive and gram-negative organisms . Two of the genes encode RNAs similar in size to the E . coli species; the third, from Bacillus subtilis, specifies an RNA more than twice as large . The heterologous genes are expressed efficiently in E . coli, and the product RNAs resemble those produced by cognate cells . We conclude that the heterologous RNAs can replace E . coli 4.5S RNA and that the essential function of 4.5S RNA is evolutionarily conserved . A consensus structure is presented for the functionally related 4.5S RNA homologs.

Biochem Biophys Res Commun, 1990 Feb 28, 167(1), 154 - 60
Conserved N-terminal sequences in the flagellins of archaebacteria; Kalmokoff ML et al.; Methanococcus voltae produces two flagellins of molecular weight 31,000 and 33,000 . Amino acid analysis as well as peptide mapping with cyanogen bromide, chymotrypsin and Staphylococcus aureus V-8 protease indicates that the two flagellins are distinct . N-terminal sequencing of the 31,000 Mc . voltae flagellin as well as the 24,000 and 25,000 molecular weight flagellins of Methanospirillum hungatei GP1 shows an extensive homology with the reported N-terminus of the flagellins from Halobacterium halobium, deduced from the nucleotide sequence of the cloned genes . However, the N-termini of all three sequenced methanogen flagellins lack a terminal methionine and start at position 13 from the N-terminus of H . halobium flagellins . This initial 12 amino acid stretch may be a leader peptide which is subsequently cleaved to generate the mature flagellin, which could suggest flagellar assembly in archaebacteria occurs by a mechanism distinct from that in eubacteria . The high degree of conservation of the N-terminus of the flagellins from Mc . voltae, Msp . hungatei and H . halobium suggests an important role for this sequence, and that the archaebacteria share a common mechanism for flagellar biosynthesis.

J Biol Chem, 1990 Feb 25, 265(6), 3034 - 9
Organization and nucleotide sequence of a gene cluster coding for eight ribosomal proteins in the archaebacterium Halobacterium marismortui; Arndt E et al.; A DNA fragment containing the genes for the eight ribosomal proteins HmaL3, HL6, HmaL23, HmaL2, HmaS19, HmaL22, HmaS3, and HmaL29 from Halobacterium marismortui has been cloned and sequenced . The organization of this gene cluster in general corresponds to the S10 operon of Escherichia coli although there exists some differences between them . The sequence analysis of the 5'- and 3'-region of the gene cluster revealed three open reading frames (orf1, orf2, and orf3) which do not code for any ribosomal protein whose structure is known . A putative promoter is located upstream of orf1 . Out of the eight ribosomal proteins five have counterparts in eubacteria only, two in both eubacteria and eukaryotes, and one is exclusively related to an eukaryotic ribosomal protein.

J Biol Chem, 1990 Feb 5, 265(4), 1852 - 7
Purification and properties of 5,10-methylenetetrahydromethanopterin reductase, a coenzyme F420-dependent enzyme, from Methanobacterium thermoautotrophicum strain delta H; te Brommelstroet BW et al.; 5,10-Methylenetetrahydromethanopterin reductase was purified 22-fold to apparent homogeneity from the methanogenic bacterium Methanobacterium thermoautotrophicum . The enzyme catalyzes the reduction of 5,10-methylene- to 5-methyltetrahydromethanopterin . The electron carrier coenzyme F420 is specifically used as the cosubstrate . The reductase reaction may proceed in both directions, methylene reduction is, however, thermodynamically favored . In addition, the velocity of the reaction in this direction exceeds the reverse reaction by a factor of 26 . The reductase is composed of a single subunit with an estimated Mr = 35,000 . The active enzyme does not contain a flavin prosthetic group or iron-sulfur clusters, in contrast to 5,10-methylenetetrahydrofolate reductases purified from eukaryotic and eubacterial sources, which catalyze an analogous reaction as the methanogenic reductase.

Proc Natl Acad Sci U S A, 1990 Feb, 87(4), 1586 - 90
Initiation of protein synthesis from a termination codon; Varshney U et al.; We show that the amber termination codon UAG can initiate protein synthesis in Escherichia coli . We mutated the initiation codon AUG of the chloramphenicol acetyltransferase (CAT) gene to UAG (CATam1) and translated mRNA derived from the mutant CAT gene in E . coli S-30 extracts . A full-length CAT polypeptide was synthesized in the presence of tRNA(fMetCUA), a mutant E . coli initiator tRNA which has a change in the anticodon sequence from CAU to CUA . Addition of purified E . coli glutaminyl-tRNA synthetase substantially stimulated synthesis of the CAT polypeptide . Thus, initiation of protein synthesis with UAG and tRNA(fMetCUA) most likely occurs with glutamine and not methionine . The UAG codon also initiates protein synthesis in vivo . To eliminate a weak secondary site of initiation from AUC, the fifth codon, we further mutagenized the CATam1 gene at codons 2 (GAG----GAC) and 5 (AUC----ACC) . Transformation of E . coli with the resultant CATam1.2.5 gene yielded transformants that synthesized CAT polypeptide and were resistant to chloramphenicol only when they were also transformed with the mutant tRNA(fMetCUA) gene . Immunoblot analyses and assays for CAT enzyme activity in extracts from transformed cells indicate that initiation from UAG is efficient, 60-70% of that obtained from AUG . Initiation of protein synthesis from UAG using a mutant initiator tRNA allows tightly regulated expression of specific genes . This may be generally useful for overproduction in E . coli and other eubacteria of proteins which are toxic to these cells.

J Bacteriol, 1990 Feb, 172(2), 579 - 88
A family of genes encode the multiple forms of the Saccharomyces cerevisiae ribosomal proteins equivalent to the Escherichia coli L12 protein and a single form of the L10-equivalent ribosomal protein; Newton CH et al.; The budding yeast Saccharomyces cerevisiae contains a family of genes that encodes four different but related small acidic ribosomal proteins designated L12eIA, L12eIB, L12eIIA, and L12eIIB and a single larger protein designated L10e . These proteins are equivalent (e) to the L12 and L10 proteins of Escherichia coli that assemble as a 4:1 complex onto the large ribosomal subunit . The five yeast genes (or their cDNAs) have been cloned and sequenced (M . Remacha, M . T . Saenz-Robles, M . D . Vilella, and J . P . G . Ballesta, J . Biol . Chem . 263:9044-9101, 1988; K . Mitsui and K . Tsurugi, Nucleic Acids Res . 16:3573, 3574, and 3575, 1988; this work) . Here, the transcripts of these genes were characterized and quantitated and the proteins they encode were compared and aligned . Four of the genes, L12eIA, -IB, -IIA, and L10e, are uninterrupted, whereas the L12eIIB gene contains a 301-nucleotide-long intron between codons 38 and 39 . The transcripts derived from each of these genes were analyzed by Northern (RNA) hybridization, primer extension, and S1 nuclease protection . All five genes are expressed, albeit at different levels . The transcript levels are coordinate and exhibit growth rate-dependent regulation in rich (glucose) and poor (ethanol) media . The five yeast proteins each contain a highly conserved acidic carboxy terminus of about 20 residues in length . This domain of unknown function is also present in archaebacterial but absent from eubacterial L10e and L12e proteins . Comparisons of the factor-binding domains in the yeast and other eucaryotic and archaebacterial L12e proteins indicate that the original duplication to produce the type I and II genes was a very ancient event . The evolutionary relationships between the eucaryotic, archaebacterial, and eubacterial L10e and L12e genes (and proteins) are discussed.

Antonie Van Leeuwenhoek, 1990 Feb, 57(2), 83 - 9
A non-passive mechanism of butyrate excretion operates during acidogenic fermentation of methanol by Eubacterium limosum; Loubiere P et al.; The inhibitory effects of organic acids produced as fermentation end-products during methylotrophic growth of the acidogenic anaerobe, Eubacterium limosum have been investigated . Precise quantification of the intracellular concentrations of acetate and butyrate, together with delta pH measurements indicate that butyrate efflux cannot be explained by a process of passive diffusion . Intracellular concentrations of butyrate were significantly lower than those of the culture broth . It is argued that growth inhibition by butyrate is due to energetic limitations resulting from the energy drain associated with this non-passive efflux mechanism.

Antonie Van Leeuwenhoek, 1990 Feb, 57(2), 63 - 70
Growth stimulation of Treponema denticola by periodontal microorganisms; ter Steeg PF et al.; Previous experiments have indicated that enrichment of subgingival plaque in human serum can lead to the accumulation of Treponema denticola . T . denticola depends on bacterial interactions for its growth in serum . Aim of the present study was to identify specific microorganisms involved in the growth stimulation of T . denticola . To this end, strains isolated from previous plaque enrichment cultures were tested for growth stimulation in co-cultures with T . denticola . In addition, growth of T . denticola was tested in culture filtrates of the same strains, Bacteroides intermedius, Eubacterium nodatum, Veillonella parvula and Fusobacterium nucleatum were found to enhance growth of T . denticola in co-cultures . A continuous co-culture of T . denticola, F . nucleatum and B . intermedius in human serum gave very high levels of T . denticola, up to 3.10(9).ml-1 . Mechanisms involved in growth stimulation may include the ability of B . intermedius and E . nodatum to cleave the protein-core of serum (glyco-)proteins, making these molecules accessible for degradation by T . denticola . In addition, E . nodatum was found to produce a low-molecular weight growth-factor for T . denticola, that was heat-stable and acid as well as alkaline resistant . V . parvula may provide peptidase activities complementary to those of T . denticola . The nature of the growth enhancing activity of F . nucleatum is yet unknown . The data support the dependency of T . denticola on other bacterial species for growth in the periodontal pocket.

J Med Microbiol, 1990 Feb, 31(2), 103 - 8
Pathogenic potential of Eubacterium yurii subspecies; Margaret BS et al.; Several mechanisms that could contribute to the periodontopathogenic potential of Eubacterium yurii were investigated . All 18 strains examined produced RNAase and the metabolites H2S, indole and butyrate . Some strains produced phosphatase and DNAase . Methanol extracts of whole cells of E . yurii subspp . yurii and margaretiae stimulated bone resorption in vitro comparable to that produced by recognised periodontal pathogens . These results suggest that further studies should be performed to elucidate the role of E . yurii in periodontal disease.

Infect Immun, 1990 Feb, 58(2), 523 - 8
Chronic arthritis induced in rats by cell wall fragments of Eubacterium species from the human intestinal flora; Severijnen AJ et al.; To investigate arthritis-inducing properties of Eubacterium species, which are major residents of the human intestinal flora, cell wall fragments (CWF) of several Eubacterium strains were prepared and tested in an animal model . After a single intraperitoneal injection in the rat, CWF of E . aerofaciens, E . contortum, and E . lentum induced a chronic polyarthritis . E . limosum and E . tortuosum CWF induced an acute self-limiting joint inflammation, whereas E . rectale CWF failed to do so . The rhamnose contents of the isolated CWF were not related to their arthritis-inducing properties . Paradoxically, the sensitivity of CWF to lysozyme digestion, which is regarded as a parameter for the clearance of CWF in tissues, appeared to be positively correlated with the ability of Eubacterium CWF to induce chronic joint inflammation . Our findings show the diversity in arthritis-inducing properties among different species of the anaerobic genus Eubacterium and underline the importance of the anaerobic intestinal flora in the induction of joint inflammation.

Mol Microbiol, 1990 Feb, 4(2), 305 - 14
Isolation of a secY homologue from Bacillus subtilis: evidence for a common protein export pathway in eubacteria; Suh JW et al.; Genetic and biochemical studies have shown that the product of the Escherichia coli secY gene is an integral membrane protein with a central role in protein secretion . We found the Bacillus subtilis secY homologue within the spc-alpha ribosomal protein operon at the same position occupied by E . coli secY . B . subtilis secY coded for a hypothetical product 41% identical to E . coli SecY, a protein thought to contain 10 membrane-spanning segments and 11 hydrophilic regions, six of which are exposed to the cytoplasm and five to the periplasm . We predicted similar segments in B . subtilis SecY, and the primary sequences of the second and third cytoplasmic regions and the first, second, fourth, fifth, seventh, and tenth membrane segments were particularly conserved, sharing greater than 50% identity with E . coli SecY . We propose that the conserved cytoplasmic regions interact with similar cytoplasmic secretion factors in both organisms and that the conserved membrane-spanning segments actively participate in protein export . Our results suggest that despite the evolutionary differences reflected in cell wall architecture, Gram-negative and Gram-positive bacteria possess a similar protein export apparatus.

Philos Trans R Soc Lond B Biol Sci, 1990 Jan 30, 326(1236), 367 - 78
Structural aspects of proton-pumping ATPases; Walker JE et al.; ATP synthase is found in bacteria, chloroplasts and mitochondria . The simplest known example of such an enzyme is that in the eubacterium Escherichia coli; it is a membrane-bound assembly of eight different polypeptides assembled with a stoichiometry of alpha 3 beta 3 gamma 1 delta 1 epsilon 1 a1b2c10-12 . The first five of these constitute a globular structure, F1-ATPase, which is bound to an intrinsic membrane domain, F0, an assembly of the three remaining subunits . ATP synthases driven by photosynthesis are slightly more complex . In chloroplasts, and probably in photosynthetic bacteria, they have nine subunits, all homologues of the components of the E . coli enzyme; the additional subunit is a duplicated and diverged relation of subunit b . The mammalian mitochondrial enzyme is more complex . It contains 14 different polypeptides, of which 13 have been characterized . Two membrane components, a (or ATPase-6) and A6L, are encoded in the mitochondrial genome in overlapping genes and the remaining subunits are nuclear gene products that are translated on cytoplasmic ribosomes and then imported into the organelle . The sequence of the proteins of ATP-synthase have provided information about amino acids that are important for its function . For example, amino acids contributing to nucleotide binding sites have been identified . Also, they provide the basis of models of secondary structure of membrane components that constitute the transmembrane proton channel . An understanding of the coupling of the transmembrane potential gradient for protons, delta mu H+, to ATP synthesis will probably require the determination of the structure of the entire membrane bound complex . Crystals have been obtained of the globular domain, F1-ATPase . They diffract to a resolution of 3-4 A and data collection is in progress . As a preliminary step towards crystallization of the entire complex, we have purified it from bovine mitochondria and reconstituted it into phospholipid vesicles.

Rheumatol Int, 1990, 10(1), 21 - 9
Experimental immune mediated arthritis in rhesus monkeys . A model for human rheumatoid arthritis?
Bakker NP, van Erck MG, Zurcher C, Faaber P, Lemmens A, Hazenberg M, Bontrop RE, Jonker M.
The induction of experimental arthritis in rhesus monkeys was studied by intradermal immunization of bovine type II collagen and antigens derived from Mycobacterium tuberculosis, Streptococcus pyogenes, and Eubacterium aerofaciens . The tested bacterial antigens proved to be not arthrogenic . Bovine type II collagen induced clinical arthritis in 50% of the rhesus monkeys . Type II collagen induced arthritis in rhesus monkeys proved to be a potential model to study clinical, serological, histological, genetic, and immunologic features associated with human RA.

Arch Microbiol, 1990, 153(5), 444 - 7
Isolation of lipid activated pseudomurein precursors from Methanobacterium thermoautotrophicum; Hartmann E et al.; From cell extracts of the pseudomurein possessing methanogen Methanobacterium thermoautotrophicum two putative pseudomurein precursors were isolated and characterized: (1) an undecaprenyl pyrophosphate activated disaccharide pentapeptide composed of N-acetylglucosamine, N-acetyltalosaminuronic acid, alanine, glutamic acid and lysine in a molar ratio of 1:1:2:2:1 and (2) the corresponding undecaprenyl pyrophosphate activated tetrapeptide lacking one alanine residue . The isolation of these precursors show that the biosynthesis of the eubacterial murein and the methanobacterial pseudomurein differs not only in the cytoplasmic step, as recently described, but also in the lipid stage.

Arch Microbiol, 1990, 153(3), 241 - 7
Complete nucleotide sequences of seven eubacterial genes coding for the elongation factor Tu: functional, structural and phylogenetic evaluations; Ludwig W et al.; The nucleotide sequences of cloned genes coding for the elongation factor Tu of seven eubacteria have been determined . These genes were from Anacystis nidulans, Bacillus subtilis, Bacteroides fragilis, "Deinonema" spec., Pseudomonas cepacia, Shewanella putrefaciens and Streptococcus oralis . The primary structures of the genes were compared to the available sequences of prokaryotic elongation factors Tu and eukaryotic elongation factors 1 alpha . A conservation profile was determined for homologous amino acid residues . Sites of known or putative functions are usually located at highly conserved positions or within highly conserved sequence stretches . The aligned 24 amino acid sequences were used as basis for a phylogenetic analysis . The phylogenetic tree corroborates the kingdom as well as phylum concept deduced from 16S rRNA data.

Microbios, 1990, 64(260-261), 197 - 202
Sensitivity of oral bacteria to imipenem and other antibiotics; Wilson M et al.; Specimens from a variety of oral and maxillofacial infections were cultured and 88 bacterial isolates belonging to the genera Bacteroides, Actinomyces, Staphylococcus, Veillonella, Eubacterium, Streptococcus and Aerococcus were obtained . These were tested for their sensitivity to imipenem, penicillin, cefuroxime, vancomycin, erythromycin and fusidic acid . Imipenem proved to be the most effective of the antibiotics tested, inhibiting all 88 isolates . Penicillin was the least effective in that it inhibited only 56% of the isolates.

J Gen Microbiol, 1990 Jan, 136 ( Pt 1), 11 - 8
5S rRNA sequences of representatives of the genera Chlorobium, Prosthecochloris, Thermomicrobium, Cytophaga, Flavobacterium, Flexibacter and Saprospira and a discussion of the evolution of eubacteria in general; Van den Eynde H et al.; 5S rRNA sequences were determined for the green sulphur bacteria Chlorobium limicola, Chlorobium phaeobacteroides and Prosthecochloris aestuarii, for Thermomicrobium roseum, which is a relative of the green non-sulphur bacteria, and for Cytophaga aquatilis, Cytophaga heparina, Cytophaga johnsonae, Flavobacterium breve, Flexibacter sp . and Saprospira grandis, organisms allotted to the phylum 'Bacteroides-Cytophaga-Flavobacterium' and relatives as determined by 16S rRNA analyses . By using a clustering algorithm a dendrogram was constructed from these sequences and from all other known eubacterial 5S RNA sequences . The dendrogram showed differences, as well as similarities, with respect to results obtained by 16S RNA analyses . The 5S RNA sequences of green sulphur bacteria were closely related to one another, and to a cluster containing 5S RNA sequences from Bacteroides and its relatives, including Cytophaga aquatilis . 5S RNA sequences of all other representatives of the 'Bacteroides-Cytophaga-Flavobacterium' phylum as distinguished by 16S RNA analysis failed to group with Bacteroides and related clusters . On the basis of 5S RNA sequences, Thermomicrobium roseum clustered with Chloroflexus aurantiacus, as was expected from 16S RNA analysis.

J Gen Microbiol, 1990 Jan, 136 ( Pt 1), 1 - 10
16S ribosomal RNA sequence analysis for determination of phylogenetic relationship among methylotrophs; Tsuji K et al.; 16S ribosomal RNAs (rRNA) of 12 methylotrophic bacteria have been almost completely sequenced to establish their phylogenetic relationships . Methylotrophs that are physiologically related are phylogenetically diverse and are scattered among the purple eubacteria (class Proteobacteria) . Group I methylotrophs can be classified in the beta- and the gamma-subdivisions and group II methylotrophs in the alpha-subdivision of the purple eubacteria, respectively . Pink-pigmented facultative and non-pigmented obligate group II methylotrophs form two distinctly separate branches within the alpha-subdivision . The secondary structures of the 16S rRNA sequences of 'Methylocystis parvus' strain OBBP, 'Methylosinus trichosporium' strain OB3b, 'Methylosporovibrio methanica' strain 81Z and Hyphomicrobium sp . strain DM2 are similar, and these non-pigmented obligate group II methylotrophs form one tight cluster in the alpha-subdivision . The pink-pigmented facultative methylotrophs, Methylobacterium extorquens strain AM1, Methylobacterium sp . strain DM4 and Methylobacterium organophilum strain XX form another cluster within the alpha-subdivision . Although similar in phenotypic characteristics, Methylobacterium organophilum strain XX and Methylobacterium extorquens strain AM1 are clearly distinguishable by their 16S rRNA sequences . The group I methylotrophs, Methylophilus methylotrophus strain AS1 and methylotrophic species DM11, which do not utilize methane, are similar in 16S rRNA sequence to bacteria in the beta-subdivision . The methane-utilizing, obligate group I methanotrophs, Methylococcus capsulatus strain BATH and Methylomonas methanica, are placed in the gamma-subdivision . The results demonstrate that it is possible to distinguish and classify the methylotrophic bacteria using 16S rRNA sequence analysis.

Eur J Biochem, 1989 Dec 22, 186(3), 657 - 61
Identification of phenolyl cobamide from the homoacetogenic bacterium Sporomusa ovata; Stupperich E et al.; Phenolyl cobamide was isolated from cyanide extractions of the anaerobic eubacterium Sporomusa ovata . The proposed corrinoid structure {Co alpha,Co beta-(monocyano,monoaquo)-phenolyl cobamide} has been deduced from 1H NMR, fast-atom-bombardment mass spectroscopy and ultraviolet/visible spectroscopy data . The complete corrinoid resembled p-cresolyl cobamide {Co alpha,Co beta-(monocyano,monoaquo)-p-cresolyl cobamide}, which recently has been obtained from cyanide extractions of the same bacterium . The structures and chemical properties of both cobamides with uncoordinated nucleotides differed significantly from those of vitamin B12 {Co alpha-{alpha-(5,6-dimethylbenzimidazolyl)}-Co beta-cyanocobamide} . Sporomusa synthesized coenzymes of phenolyl cobamide and p-cresolyl cobamide in considerable amounts of 400 nmol/g and 1700 nmol/g dry cells, respectively . More than 90% of the complete corrinoid pool of the homoacetogenic bacterium consisted of these two corrinoids, indicating that they are physiologically important coenzymes of the bacterial metabolism.

Arch Biochem Biophys, 1989 Dec, 275(2), 531 - 9
Metabolism of leukotriene E4 to 5-hydroxy-6-mercapto7,9-trans-11,14-cis-eicosatetraenoic acid by microfloral cysteine-conjugate beta-lyase and rat cecum contents; Bernstrom K et al.; Leukotriene E4 was incubated with cysteine-conjugate beta-lyase isolated from the intestinal bacterium Eubacterium limosum . The reaction was terminated by addition of iodoacetic acid or dimethyl sulfate, and the products formed were isolated by reverse-phase high-performance liquid chromatography . The structures of two adducts of a metabolite were determined by uv spectroscopy, by gas-liquid radiochromatography, and by comparisons with chemically synthesized reference compounds . They were 5-hydroxy-6-S-carboxymethylthio-7,9-trans-11,14-cis-eicosatetraeno ic acid (iodoacetic acid adduct) and 5-hydroxy-6-S-methylthio-7,9-trans-11,14-cis-eicosatetraenoic acid (dimethyl sulfate adduct) indicating that the structure of the underivatized metabolite was 5-hydroxy-6-mercapto-7,9,11,14-eicosatetraenoic acid (5,6-HMETE) . The latter product is formed by beta-lyase-catalyzed cleavage of the cysteine C-S bond in leukotriene E4 . Leukotriene E4 was also metabolized to 5,6-HMETE by rat cecal contents . A product formed was trapped as the iodoacetic acid derivative and identified as 5-hydroxy-6-S-carboxy-methylthio-7,9,11,14-eicosatetraenoic acid . It is concluded that intestinal leukotriene E4, originating from biliary excretion of systemic cysteinyl leukotrienes or produced in the intestine, is converted by microfloral cysteine-conjugate beta-lyase to 5,6-HMETE.

Proc Natl Acad Sci U S A, 1989 Dec, 86(23), 9355 - 9
Evolutionary relationship of archaebacteria, eubacteria, and eukaryotes inferred from phylogenetic trees of duplicated genes; Iwabe N et al.; All extant organisms are though to be classified into three primary kingdoms, eubacteria, eukaryotes, and archaebacteria . The molecular evolutionary studies on the origin and evolution of archaebacteria to date have been carried out by inferring a molecular phylogenetic tree of the primary kingdoms based on comparison of a single molecule from a variety of extant species . From such comparison, it was not possible to derive the exact evolutionary relationship among the primary kingdoms, because the root of the tree could not be determined uniquely . To overcome this difficulty, we compared a pair of duplicated genes, elongation factors Tu and G, and the alpha and beta subunits of ATPase, which are thought to have diverged by gene duplication before divergence of the primary kingdoms . Using each protein pair, we inferred a composite phylogenetic tree with two clusters corresponding to different proteins, from which the evolutionary relationship of the primary kingdoms is determined uniquely . The inferred composite trees reveal that archaebacteria are more closely related to eukaryotes than to eubacteria for all the cases . By bootstrap resamplings, this relationship is reproduced with probabilities of 0.96, 0.79, 1.0, and 1.0 for elongation factors Tu and G and for ATPase subunits alpha and beta, respectively . There are also several lines of evidence for the close sequence similarity between archaebacteria and eukaryotes . Thus we propose that this tree topology represents the general evolutionary relationship among the three primary kingdoms.

J Bacteriol, 1989 Dec, 171(12), 6689 - 95
Natural relationships among sulfate-reducing eubacteria; Devereux R et al.; Phylogenetic relationships among 20 nonsporeforming and two endospore-forming species of sulfate-reducing eubacteria were inferred from comparative 16S rRNA sequencing . All genera of mesophilic sulfate-reducing eubacteria except the new genus Desulfomicrobium and the gliding Desulfonema species were included . The sporeforming species Desulfotomaculum ruminis and Desulfotomaculum orientis were found to be gram-positive organisms sharing 83% 16S rRNA sequence similarity, indicating that this genus is diverse . The gram-negative nonsporeforming species could be divided into seven natural groups: group 1, Desulfovibrio desulfuricans and other species of this genus that do not degrade fatty acids (this group also included "Desulfomonas" pigra); group 2, the fatty acid-degrading "Desulfovibrio" sapovorans; group 3, Desulfobulbus species; group 4, Desulfobacter species; group 5, Desulfobacterium species and "Desulfococcus" niacini; group 6, Desulfococcus multivorans and Desulfosarcina variabilis; and group 7, the fatty acid-oxidizing "Desulfovibrio" baarsii . (The quotation marks are used to indicate the need for taxonomic revision.) Groups 1 to 3 are incomplete oxidizers that form acetate as an end product; groups 4 to 7 are complete oxidizers . The data were consistent with and refined relationships previously inferred by oligonucleotide catalogs of 16S rRNA . Although the determined relationships are generally consistent with the existing classification based on physiology and other characteristics, the need for some taxonomic revision is indicated.

Nature, 1989 Nov 30, 342(6249), 572 - 4
Isolation of a gene regulated by hydrostatic pressure in a deep-sea bacterium; Bartlett D et al.; Barophilic bacteria inhabit the deep oceans, and the specific functional modifications and regulatory mechanisms which govern adaptation to hydrostatic pressure are beginning to be understood . For example, the rate of production of several proteins by some hydrothermal vent archaebacteria and the degree of saturation of membrane lipids in other deep-sea bacteria have been found to change as a result of cultivation at high pressure . We report here the cloning of gene, ompH, which encodes a major pressure-inducible protein of strain SS9, a gram-negative eubacterium isolated from a depth of 2.5 kilometres in the Sulu Sea . Messenger RNA encoded by ompH is expressed when cells are grown at 280 atm but not at 1 atm, indicating that transcription of the ompH gene is controlled by hydrostatic pressure . The function of the OmpH protein in adaptation to high pressure and the use of the ompH gene in studying how bacteria sense and respond to pressure is discussed.

J Mol Biol, 1989 Nov 20, 210(2), 245 - 54
Conserved sequence elements upstream and downstream from the transcription initiation site of the Caulobacter crescentus rrnA gene cluster; Amemiya K; The nucleotide sequence and in vivo transcription start sites for rrnA, one of the two rRNA gene clusters of the eubacterium Caulobacter crescentus, have been determined . Two transcription start sites, a major and minor, for the rRNA gene cluster are located more than 700 nucleotides upstream from the 16 S rRNA gene . Transcription was detected from only the major start site in swarmer cells . But after the swarmer-to-stalked cell transition, transcription was detected from both rRNA start sites and continued throughout the developmental cell cycle when cells were grown in minimal medium . On the other hand, transcription from only the major start site was detected in cells growing in a complex medium . A small open reading frame was found upstream from the rRNA gene transcription start sites and was followed by an inverted repeat sequence . No sequence homology was found between the major rRNA gene transcription start site and the Escherichia coli sigma 70 promoters or the consensus sequence elements reported for C . crescentus fla promoters . However, there were two areas of homology when the major rRNA gene promoter was compared to the nucleotide sequence of the C . crescentus trpFBA promoter . There was a 12 nucleotide sequence centered around the -10 region of both promoters that was closely homologous . In addition, immediately downstream from the transcription start there was a sequence element that was identical in both promoters . These nucleotide sequence elements were not in the temporally expressed fla promoters of C . crescentus.

Eur J Biochem, 1989 Nov 20, 185(3), 685 - 93
Primary structures of five ribosomal proteins from the archaebacterium Halobacterium marismortui and their structural relationships to eubacterial and eukaryotic ribosomal proteins; Hatakeyama T et al.; The complete amino acid sequences of ribosomal proteins L9, L20, L21/22, L24 and L32 from the archaebacterium Halobacterium marismortui were determined . The comparison of the sequences of these proteins with those from other organisms revealed that proteins L21/22 and L24 are homologous to ribosomal protein Yrp29 from yeast and L19 from rat, respectively, and that H . marismortui L20 is homologous to L30 from eubacteria . H . marismortui ribosomal protein L9 showed sequence homology to both L29 from yeast and L15 from eubacteria . No homologous protein was found for H . marismortui L32 . These results are discussed with respect to the phylogenetic relationship between eubacteria, archaebacteria and eukaryotes.

Eur J Biochem, 1989 Nov 6, 185(2), 355 - 64
The 23S ribosomal RNA higher-order structure of Pseudomonas cepacia and other prokaryotes; Hopfl P et al.; A 23S ribosomal RNA gene of Pseudomonas cepacia has been cloned and sequenced . A general higher-order structure model based on earlier published models has been derived from comparative analysis of 23S-like rRNAs of eubacteria, archaebacteria, organelles and eukaryotes . Differences between the previous models were carefully analyzed and controversial regions evaluated . Moderately large insertions and deletions have been found at new points in the secondary structure . The analysis of 50 published as well as unpublished 23S rRNA sequences provide additional proof for six of the seven previously suggested tertiary interactions within the 23S rRNA . P . cepacia is the first representative of the beta subgroup of the Proteobacteria phylum whose 23S rRNA has been sequenced . A tree reflecting evolutionary relationships of prokaryotes was constructed . The topology of this tree is in good agreement with the 16S rRNA tree.

J Bacteriol, 1989 Nov, 171(11), 6323 - 9
Tandem arrangement of photolyase and superoxide dismutase genes in Halobacterium halobium; Takao M et al.; A DNA fragment containing the photolyase gene was cloned from Halobacterium halobium . The deduced amino acid sequence is highly similar to those of four known photolyases from eubacteria and a eucaryote . The cloned gene expressed in Escherichia coli cells the survival of UV-irradiated host cells by photoreactivation . These results indicate that photolyases of eucaryotes, eubacteria, and archaebacteria are derived from a common origin . In this cloned DNA fragment, two additional open reading frames (ORFs), ORF 151 and ORF 200, were found in the 5' and 3' adjacent flanking regions of the photolyase gene . ORF 200 shows unequivocal amino acid sequence homology to all known manganese and iron superoxide dismutases . Northern (RNA) hybridization analysis of H . halobium RNA revealed the existence of three transcripts, one of which covered all three ORFs, indicating that photolyase and superoxide dismutase are partly cotranscribed in this bacterium.

Antimicrob Agents Chemother, 1989 Nov, 33(11), 1883 - 9
Quinolones and coumarins eliminate chloroplasts from Euglena gracilis; Krajcovic J et al.; Quinolones and coumarins were potent eliminators of chloroplasts from Euglena gracilis . There was a remarkable similarity between antichloroplastic and antibacterial activities of DNA gyrase inhibitors . Quinolones produced 100% chloroplast-free cells in concentrations which do not affect cell viability . Optimal conditions were exponential growth, continuous illumination, and neutral or slightly alkaline pH . Coumarins were more toxic than quinolones . Among the quinolones, ofloxacin was the most potent in eliminating chloroplasts . Among the coumarins, coumermycin A1 was the most potent . New quinolones and coumermycin A1 were able to induce the complete inability of originally green cells to form green colonies after 24 h of drug exposure, while clorobiocin and novobiocin required several days of exposure . Darkness, heat shock (42 degrees C, 10 min), or simultaneous treatment with chloramphenicol or rifampin decreased the potency of DNA gyrase inhibitors for producing chloroplast-free cells . Remarkably, in cells in which division was blocked by three different methods (resting medium, hyperthermic conditions {37 degrees C}, or addition of cycloheximide), new quinolones and coumermycin A1 nevertheless eliminated chloroplasts . The antichloroplastic activity of DNA gyrase inhibitors is additional data suggesting an evolutionary relationship between chloroplasts and eubacteria.

J Mol Evol, 1989 Nov, 29(5), 448 - 62
Sequence alignment and evolutionary comparison of the L10 equivalent and L12 equivalent ribosomal proteins from archaebacteria, eubacteria, and eucaryotes; Shimmin LC et al.; The genes corresponding to the L10 and L12 equivalent ribosomal proteins (L10e and L12e) of Escherichia coli have been cloned and sequenced from two widely divergent species of archaebacteria, Halobacterium cutirubrum and Sulfolobus solfataricus . The deduced amino acid sequences of the L10e and L12e proteins have been compared to each other and to available eubacterial and eucaryotic sequences . We have identified the human P0 protein as the eucaryotic L10e . The L10e proteins from the three kingdoms were found to be colinear . The eubacterial L10e protein is much shorter than the archaebacterial-eucaryotic proteins because of two large deletions, one internal and one at the carboxy terminus . The archaebacterial and eucaryotic L12e proteins were also colinear; the eubacterial protein is homologous to the archaebacterial and eucaryotic L12e proteins, but has suffered rearrangement through what appear to be gene fusion events . Intraspecies comparisons between L10e and L12e sequences indicate the archaebacterial and eucaryotic L10e proteins contain a partial copy of the L12e protein fused to their carboxy terminus . In the eubacteria most of this fusion has been removed by the carboxy terminal deletion . Within the L12e-derived region, a 26-amino acid-long internal modular sequence reiterated thrice in the archaebacterial L10e, twice in the eucaryotic L10e, and once in the eubacterial L10e was discovered . This modular sequence also appears to be present as a single copy in all L12e proteins and may play a role in L12e dimerization, L10e-L12e complex formation, and the function of L10e-L12e complex in translation.(ABSTRACT TRUNCATED AT 250 WORDS)

J Bacteriol, 1989 Nov, 171(11), 6106 - 16
Chemiosmotic energy conversion of the archaebacterial thermoacidophile Sulfolobus acidocaldarius: oxidative phosphorylation and the presence of an F0-related N,N'-dicyclohexylcarbodiimide-binding proteolipid; Lubben M et al.; The energy-transducing mechanism of the thermoacidophilic archaebacterium Sulfolobus acidocaldarius DSM 639 has been studied, addressing the question whether chemiosmotic proton gradients serve as an intermediate energy store driving an F0F1-analogous ATP synthase . At pH 3.5, respiring S . acidocaldarius cells developed an electrochemical potential of H+ ions, consisting mainly of a proton gradient and a small inside-negative membrane potential . The steady-state proton motive force of 140 to 160 mV was collapsed by protonophores, while N,N'-dicyclohexylcarbodiimide (DCCD) caused a hyperpolarization of the membrane, as expected for a reagent commonly used to inhibit the flux through proton channels of F0F1-type ATP synthases . Cellular ATP content was strongly related to the proton motive force generated by respiration and declined rapidly, either by uncoupling or by action of DCCD, which in turn induced a marked respiratory control effect . This observation strongly supports the operation of chemiosmotic ATP synthesis with H+ as the coupling ion . The inhibition of ATP synthesis by {14C}DCCD was correlated with covalent reactions with membrane proteins . The extraction of labeled membranes with organic solvents specifically yielded a readily aggregating proteolipid of 6 to 7 kilodaltons apparent molecular mass . Its amino acid composition revealed significant similarity to the proteolipid found in eubacteria, such as Escherichia coli, as an extremely hydrophobic constituent of the F0 proton channel . Moreover, the N-terminal amino acid sequence of the Sulfolobus proteolipid displays a high degree of homology to eubacterial sequences, as well as to one derived from nucleic acid sequencing of another Sulfolobus strain (K . Denda, J . Konishi, T . Oshima, T . Date, and M . Yoshida, J . Biol . Chem . 264:7119-7121, 1989) . Despite certain structural similarities between eucaryotic vacuolar ATPases and the F1-analogous ATPase from Sulfolobus sp . described earlier, the results reported here promote the view that the archaebacterial ATP-synthesizing complex functionally belongs to the F0F1 class of ATPases . These may be considered as phylogenetically conserved catalysts of energy transduction present in all kingdoms of organisms.

J Am Vet Med Assoc, 1989 Oct 15, 195(8), 1104 - 7
Isolation of Eubacterium suis from sows with cystitis; Walker RL et al.; Eubacterium suis was isolated from the bladders of 3 of 64 sows (4.7%) examined at slaughter . Two of the sows had cystitis similar to other descriptions of E suis infections in sows . There was no histologic evidence of cystitis in the third sow from which E suis was recovered . In all 3 instances, a mixed infection was present . Other bacteria present were predominantly gram-positive organisms . Eubacterium suis cystitis may be more prevalent than previously suspected . The special culture conditions required to grow this organism may account for the paucity of reports in the United States.

Gene, 1989 Oct 15, 82(1), 65 - 75
Phylogenetic comparative analysis and the secondary structure of ribonuclease P RNA--a review; Pace NR et al.; The most incisive a priori approach to inferring the higher order structure of large RNAs has proven to be the use of phylogenetic comparisons . This article provides guidelines to the method, using as an illustration the elucidation of the secondary structure of the catalytic RNA subunit of ribonuclease P (RNase P) . The resultant structure is compared to the possibilities that are predicted thermodynamically for the RNase P RNA sequences of nine eubacteria.

J Mol Biol, 1989 Oct 5, 209(3), 345 - 58
Ribosomal RNA operon anti-termination . Function of leader and spacer region box B-box A sequences and their conservation in diverse micro-organisms; Berg KL et al.; All Escherichia coli rrn operons show a common motif in which anti-terminator box B-box A sequences occur twice, first in the leader and again in the 16 S-23 S spacer . In this study we have analyzed several aspects of rrn anti-termination by leader and spacer anti-terminator sequences . Using DNA synthesis and a plasmid test system, we incorporated random changes into the leader anti-terminator region and examined these mutations for their ability to read through a strong terminator . We also examined anti-termination by synthetic box A and by rrn spacer region sequences . Information derived from these experiments was used to search the rrn sequences of other micro-organisms for possible anti-termination features . Our principal conclusions were that: (1) box A was sufficient for terminator readthrough; (2) we could show no positive requirement for box B in our test system; (3) many of the negative anti-terminator mutations caused a promoter up-effect in the absence of a terminator; (4) the search of rrn operons from other micro-organisms revealed that anti-terminator-like box B-box A sequences exist in leader and spacer regions of both eubacteria and archaebacteria . The frequent occurrence of this pattern suggested that the E . coli rrn anti-termination motif is widespread in nature and has been conserved in microbial evolution.

J Bacteriol, 1989 Oct, 171(10), 5720 - 8
Beta-lactam biosynthesis in a gram-negative eubacterium: purification and characterization of isopenicillin N synthase from Flavobacterium sp . strain SC 12.154; Palissa H et al.; The occurrence, localization, and extraction of isopenicillin N-synthase (IPNS) were investigated in the gram-negative low-level beta-lactam producer Flavobacterium sp . strain SC 12.154, which forms deacetoxycephalosporin and excretes the cephabacin 7-formamidocephalosporin . IPNS was detected with anti-IPNS antibodies raised against the Cephalosporium acremonium enzyme . The flavobacterium enzyme, whose molecular mass (38 kilodaltons) and cofactor requirements resemble those of the fungal and Streptomyces enzymes, is formed at the transition from growth to the stationary phase . It was extracted into the polyethylene glycol phase of a polyethylene glycol-Ficoll-dextran three-phase system and was purified by quaternary aminoethyl ion-exchange chromatography, gel filtration, covalent chromatography on cystamine-Sepharose, and fast-protein liquid chromatography on Mono Q . The enzyme was characterized with respect to sulfhydryl requirement, inhibition by disulfides and metal ions, pH and temperature dependence, and stimulation by polyethylene glycol and low Triton X-100 concentrations, as well as by several amino acids, including alpha-aminoadipic acid and cysteine . The Km for alpha-aminoadipyl-cysteinyl-D-valine was 0.08 mM . An inactive membrane-associated form of IPNS was detected together with a beta-lactamase active on isopenicillin N . The system has been suggested as a model for the study of endogenous functions of beta-lactams in bacteria.

Proc Natl Acad Sci U S A, 1989 Oct, 86(20), 7746 - 50
Sensory rhodopsins I and II modulate a methylation/demethylation system in Halobacterium halobium phototaxis; Spudich EN et al.; This work demonstrates that phototaxis stimuli in the archaebacterium Halobacterium halobium control a methylation/demethylation system in vivo through photoactivation of sensory rhodopsin I (SR-I) in either its attractant or repellent signaling form as well as through the repellent receptor sensory rhodopsin II (SR-II, also called phoborhodopsin) . The effects of positive stimuli that suppress swimming reversals (i.e., an increase in attractant or decrease in repellent light) and negative stimuli that induce swimming reversals (i.e., a decrease in attractant or increase in repellent light) through each photoreceptor were monitored by assaying release of volatile {3H}methyl groups . This assay has been used to measure {3H}methanol produced during the process of adaptation to chemotactic stimuli in eubacteria . In H . halobium positive photostimuli produce a transient increase in the rate of demethylation followed by a decrease below the unstimulated value, whereas negative photostimuli cause an increase followed by a rate similar to that of the unstimulated value . Photoactivation of the SR-I attractant and simultaneous photoactivation of the SR-II repellent receptors cancel in their effects on demethylation, demonstrating the methylation system is regulated by an integrated signal . Analysis of mutants indicates that the source for the volatile methyl groups is intrinsic membrane proteins distinct from the chromoproteins that share the membrane . A methyl-accepting protein (94 kDa) previously correlated in amount with the SR-I chromoprotein (25 kDa) is shown here to be missing in a recently isolated SR-I-SR-II+ mutant (Flx3b), thus confirming the association of this protein with SR-I . Photoactivated SR-II in mutant Flx3b controls demethylation, predicting the existence of a photomodulated methyl-accepting component distinct from the 94-kDa protein of SR-I . We present a model in which the three known phototaxis signaling receptor states (the attractant receptor SR-I587, its repellent form S373, and the repellent receptor SR-II490) are coupled to two distinct transducers the demethylation of which is controlled by one integrated signal.

J Mol Biol, 1989 Sep 5, 209(1), 21 - 36
Organization and structure of the Methanococcus transcriptional unit homologous to the Escherichia coli "spectinomycin operon" . Implications for the evolutionary relationship of 70 S and 80 S ribosomes; Auer J et al.; By means of an immunological approach and a subsequent chromosome-walking strategy a chromosomal region encoding ribosomal proteins in the archaebacterium Methanococcus vannielii was cloned . The determination of the nucleotide sequence of the 7.8 x 10(3) base DNA fragment revealed the existence of 14 putative ribosomal protein genes and two unidentified open reading frames . They are organized in a transcriptional unit that is very similar to the Escherichia coli "spectinomycin operon" in respect of both gene composition and gene order . The Methanococcus transcriptional unit contains, in addition to those genes whose products have a homologue in the E . coli operon, three genes whose products share sequence similarity with eukaryotic 80 S but not with eubacterial ribosomal proteins . The Methanococcus ribosomal proteins almost exclusively exhibit a higher sequence similarity to eukaryotic 80 S ribosomal proteins than to those of eubacteria and many of them have a size intermediate between those of their eukaryotic and eubacterial homologues . These results are discussed in terms of a hypothesis that implies that the recent eubacterial ribosome developed by a "minimization" process from a more complex organelle and that the archaebacterial ribosome has maintained features of this ancestor.

J Mol Biol, 1989 Sep 5, 209(1), 37 - 54
Codon recognition patterns as deduced from sequences of the complete set of transfer RNA species in Mycoplasma capricolum . Resemblance to mitochondria; Andachi Y et al.; The nucleotide sequences of the complete set of tRNA species in Mycoplasma capricolum, a derivative of Gram-positive eubacteria, have been determined . This bacterium represents the first genetic system in which the sequences of all the tRNA species have been determined at the RNA level . There are 29 tRNA species: three for Leu, two each for Arg, Ile, Lys, Met, Ser, Thr and Trp, and one each for the other 12 amino acids as judged from aminoacylation and the anticodon nucleotide sequences . The number of tRNA species is the smallest among all known genetic systems except for mitochondria . The tRNA anticodon sequences have revealed several features characteristic of M . capricolum . (1) There is only one tRNA species each for Ala, Gly, Leu, Pro, Ser and Val family boxes (4-codon boxes), and these tRNAs all have an unmodified U residue at the first position of the anticodon . (2) There are two tRNAThr species having anticodons UGU and AGU; the first positions of these anticodons are unmodified . (3) There is only one tRNA with anticodon ICG in the Arg family box (CGN); this tRNA can translate codons CGU, CGC and CGA . No tRNA capable of translating codon CGG has been detected, suggesting that CGG is an unassigned codon in this bacterium . (4) A tRNATrp with anticodon UCA is present, and reads codon UGA as Trp . On the basis of these and other observations, novel codon recognition patterns in M . capricolum are proposed . A comparatively small total, 13, of modified nucleosides is contained in all M . capricolum tRNAs . The 5' end nucleoside of the T psi C-loop (position 54) of all tRNAs is uridine, not modified to ribothymidine . The anticodon composition, and hence codon recognition patterns, of M . capricolum tRNAs resemble those of mitochondrial tRNAs.

DICP, 1989 Sep, 23(9), 668 - 70
Erythromycin-induced digoxin toxicity; Morton MR et al.; The potential interaction between certain antibiotics and digoxin has been discussed in the literature; however, few cases of actual erythromycin-induced digoxin toxicity have been reported . We present a case in which an 86-year-old woman who was taking digoxin 0.25 mg/d developed probably digoxin toxicity after the administration of erythromycin for the treatment of otitis media and streptococcal pharyngitis . Her digoxin concentration increased from a trough of 1.9 to 5.1 nmol/L six days after the erythromycin was started . Digoxin was discontinued and restarted approximately six weeks later when the patient's atrial fibrillation and congestive heart failure recurred . Her digoxin dose at this time was 0.125 mg/d and resulted in steady-state concentrations of 1.2, 1.4, and 1.2 nmol/L over the next year . Erythromycin inhibition of Eubacterium lentum, which converts digoxin into digoxin-reduction products in the gut, is the proposed mechanism of this interaction.

J Bacteriol, 1989 Sep, 171(9), 5025 - 30
Evidence for unlinked rrn operons in the Planctomycete Pirellula marina; Liesack W et al.; Southern hybridization of rRNAs to chromosomal BamHI-digested DNA of the eubacterium Pirellula marina revealed the presence of two sets of 16S and 23S rRNA genes . The two copies of the 23S rRNA genes, located on 11- and about 13-kilobase (kb) inserts, were isolated from a lambda bacteriophage Charon 35 library . The 11-kb fragment was cloned directly into pBR322, while a 5.4-kb BamHI-PstI rDNA subfragment of the approximately 13-kb insert was cloned into pUC18 . Both recombinant plasmids, pPI1100 and pPI540, were characterized by restriction enzyme mapping and Southern hybridization with the large rRNA species . Restriction fragments from both inserts were subcloned into phage M13 mp18 and mp19 . Correlation of genomic hybridization data with physical characterization of recombinant plasmids showed that, in contrast to the general organization of rrn operons in eubacteria, the 16S rRNA genes of P . marina are separated by at least 8.5 (pPI540) and 4.4 (pPI1100) kb, respectively, from the closely linked 23S-5S rRNA genes . Comparison of the flanking regions from both 23S-5S rRNA genes with published consensus sequences of structural elements indicates the presence of putative transcription signals, i.e., a single Pribnow box, discriminator, antitermination boxes A, B, and C, and a Rho-independent terminator.

Trends Genet, 1989 Sep, 5(9), 294 - 9
The evolutionary origins of organelles; Gray MW; Analysis of organellar genomes strongly supports the idea that chloroplasts and mitochondria originated in evolution as eubacteria-like endosymbionts, whose closest contemporaries are cyanobacteria and purple photosynthetic bacteria, respectively . However, there is still much debate about whether a single endosymbiotic event or multiple ones gave rise to each organelle in different eukaryotes, and considerable uncertainty about what has happened to the genomes of chloroplasts and mitochondria since their appearance in the eukaryotic cell.

Proc Natl Acad Sci U S A, 1989 Sep, 86(17), 6661 - 5
Evolution of the vacuolar H+-ATPase: implications for the origin of eukaryotes; Gogarten JP et al.; Active transport across the vacuolar components of the eukaryotic endomembrane system is energized by a specific vacuolar H+-ATPase . The amino acid sequences of the 70- and 60-kDa subunits of the vacuolar H+-ATPase are approximately equal to 25% identical to the beta and alpha subunits, respectively, of the eubacterial-type F0F1-ATPases . We now report that the same vacuolar H+-ATPase subunits are approximately equal to 50% identical to the alpha and beta subunits, respectively, of the sulfur-metabolizing Sulfolobus acidocaldarius, an archaebacterium (Archaeobacterium) . Moreover, the homologue of an 88-amino acid stretch near the amino-terminal end of the 70-kDa subunit is absent from the F0F1-ATPase beta subunit but is present in the alpha subunit of Sulfolobus . Since the two types of subunits (alpha and beta subunits; 60- and 70-kDa subunits) are homologous to each other, they must have arisen by a gene duplication that occurred prior to the last common ancestor of the eubacteria, eukaryotes, and Sulfolobus . Thus, the phylogenetic tree of the subunits can be rooted at the site where the gene duplication occurred . The inferred evolutionary tree contains two main branches: a eubacterial branch and an eocyte branch that gave rise to Sulfolobus and the eukaryotic host cell . The implication is that the vacuolar H+-ATPase of eukaryotes arose by the internalization of the plasma membrane H+-ATPase of an archaebacterial-like ancestral cell.

Nucleic Acids Res, 1989 Aug 25, 17(16), 6463 - 71
Hypervariability of simple sequences as a general source for polymorphic DNA markers; Tautz D; Short simple sequence stretches occur as highly repetitive elements in all eukaryotic genomes and partially also in prokaryotes and eubacteria . They are thought to arise by slippage like events working on randomly occurring internally repetitive sequence stretches . This predicts that they should be generally hypervariable in length . I have used the polymerase chain reaction (PCR) process to show that several randomly chosen simple sequence loci with different nucleotide composition and from different species show extensive length polymorphisms . These simple sequence length polymorphisms (SSLP) may be usefully exploited for identity testing, population studies, linkage analysis and genome mapping.

Biochem J, 1989 Aug 1, 261(3), 973 - 7
Purification and characterization of glucose dehydrogenase from the thermoacidophilic archaebacterium Thermoplasma acidophilum; Smith LD et al.; Glucose dehydrogenase was purified to homogeneity from the thermoacidophilic archaebacterium Thermoplasma acidophilum . The enzyme is a tetramer of polypeptide chain Mr 38,000 +/- 3000, it is catalytically active with both NAD+ and NADP+ cofactors, and it is thermostable and remarkably resistant to a variety of organic solvents . The amino acid composition was determined and compared with those of the glucose dehydrogenases from the archaebacterium Sulfolobus solfataricus and the eubacteria Bacillus subtilis and Bacillus megaterium . The N-terminal amino acid sequence of the Thermoplasma acidophilum enzyme was determined to be: (S/T)-E-Q-K-A-I-V-T-D-A-P-K-G-G-V-K-Y-T-T-I-D-M-P-E.

Proc Natl Acad Sci U S A, 1989 Aug, 86(15), 5969 - 73
Megabase-sized linear DNA in the bacterium Borrelia burgdorferi, the Lyme disease agent; Ferdows MS et al.; Using pulsed-field gel electrophoresis we examined the genome of Borrelia burgdorferi, a eubacterium of the spirochete phylum and the agent of Lyme disease . A population of this species' cells was lysed in situ in agarose blocks . An abundant DNA form that behaved as a linear duplex molecule under different electrophoretic conditions was found . The estimated size of the molecule was 950 kilobases . DNA from two other genera of spirochetes did not enter the gel under these conditions . These studies indicate that Borrelia spirochetes, perhaps uniquely among prokaryotic organisms, have linear chromosomes.

J Bacteriol, 1989 Aug, 171(8), 4202 - 6
Phylogenetic diversity of the Rickettsiae; Weisburg WG et al.; Small subunit rRNA sequences have been determined for representative strains of six species of the family Rickettsiaceae: Rickettsia rickettsii, Rickettsia prowazekii, Rickettsia typhi, Coxiella burnetii, Ehrlichia risticii, and Wolbachia persica . The relationships among these sequences and those of other eubacteria show that all members of the family Rickettsiaceae belong to the so-called purple bacterial phylum . The three representatives of the genus Rickettsia form a tight monophyletic cluster within the alpha subdivision of the purple bacteria . E . risticii also belongs to the alpha subdivision and shows a distant yet specific relationship to the genus Rickettsia . However, the family as a whole is not monophyletic, in that C . burnetii and W . persica are members of the gamma subdivision . The former appears to show a specific, but rather distant, relationship to the genus Legionella.

Protein Seq Data Anal, 1989 Aug, 2(5), 395 - 402
Examination of protein sequence homologies . VI . The evolution of Escherichia coli L7/L12 equivalent ribosomal proteins ('A' proteins), and the tertiary structure; Otaka E et al.; Sequence homologies among 23 complete and two partial sequences of ribosomal 'A' proteins from eukaryotes, metabacteria, eubacteria and chloroplasts, equivalent to Escherichia coli L7/L12, were examined using a correlation method that evaluates sequence similarity quantitatively . Examination of 325 comparison matrices prepared for possible combinations of the sequences indicates that 'A' protein sequences can be classified into two types: one is the "prototype" from eubacteria and chloroplasts, and the other is the "transposition type" from eukaryotes and metabacteria, which must have resulted from the internal transposition of the prototype sequence . The transposition type of eukaryotes can further be classified into P1 and P2 lines . Sequences of the P1 line are closer to those of metabacteria than to those of the P2 line . Eleven gaps, as deletion or insertion sites of amino acid residues, are necessary for an alignment of all the sequences . According to the crystallographic data for the C-terminal fragment (CTF) from E . coli L7, all the gaps involved in the CTF are located between segments that correspond to structural and functional elements such as alpha helix, beta strand, turning loop or hinge part . The existence of specific "preservation units" in these molecules is suggested . In contrast, the transposition site is located at the center of an alpha helix element that is involved in a folding domain, indicating that the transposition event was extremely drastic.

Proc Natl Acad Sci U S A, 1989 Aug, 86(16), 6121 - 5
Template supercoiling during ATP-dependent DNA helix tracking: studies with simian virus 40 large tumor antigen; Yang L et al.; Incubation of topologically relaxed plasmid DNA with simian virus 40 (SV40) large tumor antigen (T antigen), ATP, and eubacterial DNA topoisomerase I resulted in the formation of highly positively supercoiled DNA . Eukaryotic DNA topoisomerase I could not substitute for eubacterial DNA topoisomerase 1 in this reaction . Furthermore, the addition of eukaryotic topoisomerase I to a preincubated reaction mixture containing both T antigen and eubacterial topoisomerase I caused rapid relaxation of the positively supercoiled DNA . These results suggest that SV40 T antigen can introduce topoisomerase-relaxable supercoils into DNA in a reaction coupled to ATP hydrolysis . We interpret the observed T antigen supercoiling reaction in terms of a recently proposed twin-supercoiled-domain model that describes the mechanics of DNA helix-tracking processes . According to this model positive and negative supercoils are generated ahead of and behind the moving SV40 T antigen, respectively . The preferential relaxation of negative supercoils by eubacterial DNA topoisomerase I explains the accumulation of positive supercoils in the DNA template . The supercoiling assay using DNA conformation-specific eubacterial DNA topoisomerase I may be of general use for the detection of ATP-dependent DNA helix-tracking proteins.

FEMS Microbiol Lett, 1989 Aug, 51(3), 255 - 9
delta-Aminolevulinic acid biosynthesis in Escherichia coli and Bacillus subtilis involves formation of glutamyl-tRNA; O'Neill GP et al.; Cell-free extracts of Escherichia coli and Bacillus subtilis catalyzed the tRNA-dependent, RNase A-sensitive formation of delta-aminolevulinic acid (ALA) from glutamate . Cell extracts prepared from cultures of E . coli grown under aerobic or anaerobic conditions had similar levels of ALA biosynthetic activity . Both the tRNA-stimulated conversion of glutamate to ALA and the conversion of glutamate-1-semialdehyde to ALA were inhibited by gabaculin . However, gabaculin had no effect on the growth of either E . coli or B . subtilis . The tRNA-dependent transformation of glutamate to ALA in E . coli and B . subtilis thus appears to be very similar to the pathway found in cyanobacteria, certain obligate anaerobic eubacteria, archaebacteria and in the chloroplasts of algae and higher plant species.

J Biol Chem, 1989 Jul 25, 264(21), 12253 - 8
Evolution and regulation of the gene encoding superoxide dismutase from the archaebacterium Halobacterium cutirubrum; May BP et al.; The gene encoding the manganese-containing superoxide dismutase (SOD) of Halobacterium cutirubrum was isolated and characterized . The gene and 5'- and 3'-untranslated regions were located on a genomic DNA fragment of 1127 nucleotides . The deduced amino acid sequence is 200 residues long and has 39-42% identity with manganese-containing SODs of eubacteria and mitochondria . This homology may be due to either lateral transfer of the gene between eubacteria and archaebacteria or to high amino acid sequence conservation in the enzyme during the separate evolution of eubacteria and archaebacteria . Transcription of the gene initiates only about three nucleotides upstream of the translation initiation codon . The 5' end of the transcript does not contain a purine-rich Shine-Dalgarno sequence, and the promoter region does not contain consensus sequences found in other archaebacterial promoters . Termination of transcription occurs at 5 consecutive thymine residues that are preceded by a GC-rich region . The gene is basally expressed in anaerobically grown cells but is also inducible by paraquat, a generator of oxygen radicals . The same transcription initiation site is used in both types of expression, suggesting that one promoter is responsible for both basal and regulated expression . In addition to the single copy of the authentic SOD gene, the genome of H . cutirubrum contains a sequence that is very closely related to but does not code for the previously purified SOD of this organism.

FEBS Lett, 1989 Jul 17, 251(1-2), 132 - 6
The H+ ATPase regulatory subunit of Methanococcus thermolithotrophicus: amplification of an 800 bp fragment by polymerase chain reaction; Bernasconi P et al.; An 800 bp fragment of Methanococcus thermolithotrophicus genomic DNA was amplified by the polymerase chain reaction method using primers designed from conserved regions of the V-type H+ ATPase regulatory subunits from the archaebacterium Sulfolobus, and several eukaryotes . Although more than one product was obtained, only one of them had the expected size and was exclusively amplified in the presence of the left and right primers . The DNA and the deduced protein sequences of the putative Methanococcus H+ ATPase subunit revealed homology to the corresponding sequences in Sulfolobus and eukaryotes (about 60% identical residues) and a less evident homology to the eubacterial F1-ATPase alpha-subunit (22% identical residues with E . coli).

FEBS Lett, 1989 Jul 3, 250(2), 416 - 8
A small basic ribosomal protein in Sulfolobus solfataricus equivalent to L46 in yeast: structure of the protein and its gene; Ramirez C et al.; The structure of the gene for a small, very basic ribosomal protein in Sulfolobus solfataricus has been determined and the structure of the protein coded by this gene (L46e) has been confirmed by partial amino acid sequencing . The protein shows substantial sequence homology to the eukaryotic ribosomal proteins L39 in rat and L46 in yeast . There is no sequence homology to any of the eubacterial ribosomal proteins suggesting that this protein is absent in the eubacterial ribosome.

J Lipid Res, 1989 Jul, 30(7), 1033 - 9
Mechanism of intestinal formation of deoxycholic acid from cholic acid in humans: evidence for a 3-oxo-delta 4-steroid intermediate; Bjorkhem I et al.; 12 alpha-Hydroxy-3-oxo-4-cholenoic acid coupled to an adenosine nucleotide has been shown to be a metabolite of cholic acid in the intestinal anaerobic bacteria, Eubacterium species VPI 12708 (1987 . J . Biol . Chem . 262: 4701-4707) and it has been suggested that this may be an intermediate in the conversion of cholic acid into deoxycholic acid . The possibility that the intestinal conversion of cholic acid into deoxycholic acid involves a 3-oxo-delta 4-steroid as an intermediate has been studied in the present work by use of {3 beta-3H}- and {5-3H}-labeled cholic acid . Whole cells as well as cell extracts of Eubacterium sp . VPI 12708 catalyzed conversion of {3 beta-3H} + {24-14C}cholic acid into deoxycholic acid with loss of about 50% of 3H label . When unlabeled chenodeoxycholic acid (20 microM) was added along with {3 beta-3} + {24-14C}cholic acid, then approximately 85% of the {3 beta-3H}-labeled was lost from deoxycholic acid . After administration of the same mixture to two healthy volunteers, deoxycholic acid could be isolated that had lost 81 and 84%, respectively, of the 3H label . Conversion of a mixture of {5-3H}- and {24-14C}labeled cholic acid by the above intestinal bacteria or cell extracts led to loss of 79-94 of the {5-3H} label.(ABSTRACT TRUNCATED AT 250 WORDS)

FEMS Microbiol Lett, 1989 Jul 1, 51(1), 101 - 5
N-terminal amino acid sequence of the Borrelia burgdorferi flagellin; Gassmann GS et al.; The 41 kDa flagellar protein of Borrelia burgdorferi appears to be an immunodominant antigen producing an early and strong response in most, if not all, individuals during infection in humans . It would represent a very good antigen for serodiagnosis of Lyme disease, if its crossreactivity with flagella of other bacteria was low . To gain information on this point we isolated the B . burgdorferi flagellin by preparative two-dimensional electrophoresis for N-terminal amino acid analysis . By comparing the N-terminal amino acid sequences of flagellar proteins from other eubacteria we found that the first six out of twenty nine amino acids were identical to the Treponema pallidum and Treponema phagedenis 'class B' flagellins . All 29 N-terminal residues exhibited a moderate inter-genus homology (44-55%), in contrast to the high degree (67-95%) of inter-species conservation of the treponemal 'class B' flagellar N-terminal sequences . There was little similarity to other flagellins except the B . subtilis flagellar protein.

Z Naturforsch {C}, 1989 Jul-Aug, 44(7-8), 641 - 50
Molecular evolution of H+-ATPases . I . Methanococcus and Sulfolobus are monophyletic with respect to eukaryotes and Eubacteria; Gogarten JP et al.; The classification of methanogenic bacteria as archaebacteria based on 16 s rRNA sequence analysis is currently in dispute . To provide an alternative molecular marker, the polymerase chain reaction technique was used to amplify a 930 bp fragment of Methanococcus thermolithotrophicus genomic DNA corresponding to the catalytic domain of the membrane H+-ATPase . The deduced amino acid sequence was 54-58% identical to the approximately 70 kDa subunits of Sulfolobus acidocaldarius and the eukaryotic vacuolar-type H+-ATPase, and only 29% identical to the beta subunit of the eubacterial-type F0F1-ATPases . Interestingly, a highly conserved aspartate residue in the phosphorylation domain of E1E2-ATPases (P-type) is conserved in the Methanococcus sequence, but is absent from all other known vacuolar and F0F1-ATPases . This suggests that the H+-ATPase of M . thermolithotrophicus, like that of M . voltae, may have a phosphorylated intermediate, despite belonging to the vacuolar-type class of proton pumps . Phylogenetic analysis using Felsenstein's maximum likelihood method and Lake's evolutionary parsimony method confirmed that the H+-ATPases of the two archaebacteria, Methanococcus and Sulfolobus, when compared to eukaryotic vacuolar-type ATPases and eubacterial F0F1-ATPases, form a monophyletic group.

J Mol Evol, 1989 Jul, 29(1), 20 - 7
Organization and nucleotide sequence of a transcriptional unit of Methanococcus vannielii comprising genes for protein synthesis elongation factors and ribosomal proteins; Lechner K et al.; By a chromosome walking strategy the DNA region from Methanococcus vannielii flanking the genes for protein synthesis elongation factor (EF) 1 alpha and EF-2 was cloned and sequenced . A gene organization of 5' - beta' - open reading frame (ORF) 1 - ORF2 - S12 - S7 - EF-2 - EF-1 alpha - S10 - ORF3 - ORF4 - 3' was found where beta', S12, S7, S10, EF-2, and EF-1 alpha represent gene products with sequences similar to the beta' subunit of RNA polymerase, ribosomal proteins S12, S7, and S10, and EF-G and EF-Tu from Escherichia coli, respectively . ORF1-4 represent gene products with no known eubacterial counterparts . Northern blot analysis of transcripts and nuclease S1 mapping showed that transcription initiates between beta' and ORF1 and terminates at the 3' side of the S10 gene and that the genes from ORF1 to S10 are cotranscribed . Apart from the presence of two additional ORFs, ORF1 and ORF2, and of the gene for S10, this organization is identical to that of the eubacterial "streptomycin operon." ORF1 displays sequence similarity to rat liver ribosomal protein L30 and may represent one of the "additional" ribosomal proteins of Methanococcus . The sequenced part of the beta' gene and the EF-2 and EF-1 alpha gene products from Methanococcus are more similar to their eukaryotic than to their eubacterial counterparts . It appears, therefore, that the genetic organization of the translational components resembles the situation in eubacteria, whereas their primary structures are more eukaryotic in nature.

Gene, 1989 Jun 30, 79(1), 33 - 46
Organization and nucleotide sequence analysis of a ribosomal RNA gene cluster from Streptomyces ambofaciens; Pernodet JL et al.; The Streptomyces ambofaciens genome contains four rRNA gene clusters . These copies are called rrnA, B, C and D . The complete nucleotide (nt) sequence of rrnD has been determined . These genes possess striking similarity with other eubacterial rRNA genes . Comparison with other rRNA sequences allowed the putative localization of the sequences encoding mature rRNAs . The structural genes are arranged in the order 16S-23S-5S and are tightly linked . The mature rRNAs are predicted to contain 1528, 3120 and 120 nt, for the 16S, 23S and 5S rRNAs, respectively . The 23S rRNA is, to our knowledge, the longest of all sequenced prokaryotic 23S rRNAs . When compared to other large rRNAs it shows insertions at positions where they are also present in archaebacterial and in eukaryotic large rRNAs . Secondary structure models of S . ambofaciens rRNAs are proposed, based upon those existing for other bacterial rRNAs . Positions of putative transcription start points and of a termination signal are suggested . The corresponding putative primary transcript, containing the 16S, 23S and 5S rRNAs plus flanking regions, was folded into a secondary structure, and sequences possibly involved in rRNA maturation are described . The G + C content of the rRNA gene cluster is low (57%) compared with the overall G + C content of Streptomyces DNA (73%).

Science, 1989 Jun 30, 244(4912), 1569 - 71
The design and catalytic properties of a simplified ribonuclease P RNA; Waugh DS et al.; Ribonuclease P (RNase P) RNA is the catalytic moiety of the ribonucleoprotein enzyme that removes precursor sequences from the 5' ends of pre-transfer RNAs in eubacteria . Phylogenetic variation according to recently proposed secondary structure models was used to identify structural elements of the RNase P RNA that are dispensable for catalysis . A simplified RNase P RNA that consists only of evolutionarily conserved features was designed, synthesized, and characterized . Although the simplified RNA (Min 1 RNA) is only 263 nucleotides in length, in contrast to the 354 to 417 nucleotides of naturally occurring RNase P RNAs, its specificity of pre-tRNA cleavage is identical to that of the native enzymes . Moreover, the catalytic efficiencies of the Min 1 RNA and the native RNA enzymes are similar . These results focus the search for the catalytic elements of RNase P RNAs to their conserved structure.

Nucleic Acids Res, 1989 Jun 26, 17(12), 4517 - 34
Organization and nucleotide sequence of the genes encoding the large subunits A, B and C of the DNA-dependent RNA polymerase of the archaebacterium Sulfolobus acidocaldarius; Puhler G et al.; The genes for the three large subunits A, B and C, of the DNA-dependent RNA polymerase of the archaebacterium Sulfolobus acidocaldarius DSM 639, were identified and characterized . The three genes follow each other immediately in the order B-A-C, which corresponds to that found in the rpoBC operon of the Escherichia coli genome . The transcription products formed in vivo were studied by Northern analysis and the start-points were determined by S1-nuclease mapping and primer directed extension analysis . The three RNA polymerase subunit genes were co-transcribed together with an open reading frame (ORF) of 88 amino acid residues length situated immediately upstream of the B gene and two ORFs of 104 and 130 amino acid residues following the C gene (together 8500 nucleotides) . The following ORF, encoding a protein of 118 amino acids homologous to the ribosomal protein S12 of E . coli, was weakly transcribed with the large co-transcript and strongly from an own promoter . The derived amino acid sequence of the B-subunit was found to be homologous to the B- (second largest) subunits of the eukaryotic nuclear polymerases I, II and III and to the eubacterial beta-subunit . The combined A + C-subunits correspond to the A- (largest) subunits of the eukaryotic RNA polymerases I, II and III and to the eubacterial beta'-subunit . The amino acid sequence similarity of the Sulfolobus subunits to the eukaryotic components is clearly higher than to the E . coli subunit.

Oral Microbiol Immunol, 1989 Jun, 4(2), 82 - 8
The effect of subminimal inhibitory concentrations of antimicrobial agents on three bacterial mixtures; Lacroix JM et al.; A test tube technique was developed to screen bacterial mixtures to detect interbacterial interactions that play a role in determining sensitivity to antimicrobial agents . We found 3 mixtures where these bacterial interactions change the sensitivity to antimicrobials or change the proportions of each bacterial species in the mixture . The mixtures were: Fusobacterium nucleatum 102.3 and Bacteroides endodontalis ATCC 35406; F . nucleatum 102.3 and B . endodontalis BN11 a-f; and Capnocytophaga ochracea 1956c and Eubacterium saburreum 162.4 . The antimicrobials used were metronidazole for the first 2 mixtures and tetracycline for the last . F . nucleatum seems to protect B . endodontalis from the action of metronidazole . Conversely, the growth inhibition of C . ochracea by E . saburreum was lifted when tetracycline was present . We also found that the growth of C . ochracea can then permit the subsequent growth of E . saburreum . The test tube method permits the evaluation both of interbacterial interactions and the detection of any protective mechanism against antimicrobial agents in a bacterial mixture . We found that F . nucleatum 102.3 can decrease the metronidazole level in the culture medium, and by the use of 14C-metronidazole we demonstrated that acetamide is produced from metronidazole.

Appl Environ Microbiol, 1989 Jun, 55(6), 1630 - 4
Inducible bacteriophages from ruminal bacteria; Klieve AV et al.; The incidence of temperate bacteriophage in a wide range of ruminal bacteria was investigated by means of induction with mitomycin C . Supernatant liquid from treated cultures was examined for phagelike particles by using transmission electron microscopy . Of 38 ruminal bacteria studied, nine organisms (23.7%) representing five genera (Eubacteria, Bacteroides, Butyrivibrio, Ruminococcus, and Streptococcus) produced phagelike particles . Filamentous particles from Butyrivibrio fibrisolvens are the first of this morphological type reported from ruminal bacteria . All of the other particles obtained possessed polyhedral heads and long, noncontractile tails (group B-type phage) . The limited range of morphological types produced by mitomycin C induction cannot yet account for the much wider range of types found in ruminal contents by direct examination . The presence of viral genetic material in a significant percentage of the bacteria tested, as well as in a range of different genera, indicates that viral genetic material may be a normal constituent of the genome of appreciable numbers of ruminal bacteria.

Biochimie, 1989 Jun, 71(6), 701 - 9
Evolution of large subunit rRNA structure . The 3' terminal domain contains elements of secondary structure specific to major phylogenetic groups; Bachellerie JP et al.; Refined secondary structure models supported by phylogenetic evidence have been derived for the 3' terminal domain of large subunit rRNA (the region that exists as a separate 4.5 S molecular entity in chloroplast ribosomes) through a comparative analysis of all the pro- and eukaryotic sequences at present available . While several universally conserved features of secondary structure are found, a few diversified structural elements are also detected which are specific to one of the primary kingdoms, eubacteria, archaebacteria, or eukaryotes . Remarkably, some appear to be selectively preserved during the evolution of the primary kindgom, suggesting they represent functionally important structures . Thus, although the role of this 3' terminal domain in ribosomal function still remains unknown, its mode of sequence variation clearly points to a significant diversification of its function among the primary kindgoms.

Proc Natl Acad Sci U S A, 1989 Jun, 86(12), 4569 - 73
Archaebacterial DNA-dependent RNA polymerases testify to the evolution of the eukaryotic nuclear genome; Puhler G et al.; Genes for DNA-dependent RNA polymerase components B, A, and C from the archaebacterium Sulfolobus acidocaldarius and for components B", B', A, and C from the archaebacterium Halobacterium halobium were cloned and sequenced . They are organized in gene clusters in the order above, which corresponds to the order of the homologous rpoB and rpoC genes in the corresponding operon of the Escherichia coli genome . Derived amino acid sequences of archaebacterial components A and C were aligned with each other and with the sequences of corresponding (largest) subunits from the archaebacterium Methanobacterium thermoautotrophicum, with sequences of various eukaryotic nuclear RNA polymerases I, II, and III, and with the sequence of the beta' component from E . coli polymerase . The archaebacterial genes for component A are homologous to about the first two-thirds of genes for the eukaryotic component A and the eubacterial component beta', and the archaebacterial genes for component C are homologous to the last third of the genes for the eukaryotic component A and the eubacterial component beta' . Unrooted phylogenetic dendrograms derived from both distance matrix and parsimony analyses show the archaebacteria are a coherent group closely related to the eukaryotic nuclear RNA polymerase II and/or III lineages . The eukaryotic polymerase I lineage appears to arise separately from a bifurcation with the eubacterial beta' component lineage.

Science, 1989 May 26, 244(4907), 986 - 9
Prevention of translational frameshifting by the modified nucleoside 1-methylguanosine; Bjork GR et al.; The methylated nucleoside 1-methylguanosine (m1G) is present next to the 3' end of the anticodon (position 37) in all transfer RNAs (tRNAs) that read codons starting with C except in those tRNAs that read CAN codons . All of the three proline tRNA species, which read CCN codons in Salmonella typhimurium, have been sequenced and shown to contain m1G in position 37 . A mutant of S . typhimurium that lacks m1G in its tRNA when grown at temperatures above 37 degrees C, has now been isolated . The mutation (trmD3) responsible for this methylation deficiency is in the structural gene (trmD) for the tRNA(m1G37)methyltransferase . Therefore, the three proline tRNAs in the trmD3 mutant have an unmodified guanosine at position 37 . Furthermore, the trmD3 mutation also causes at least one of the tRNAPro species to frequently shift frame when C's are present successively in the message . Thus, m1G appears to prevent frameshifting . The data from eubacteria apply to both eukaryotes and archaebacteria.

J Mol Biol, 1989 May 20, 207(2), 417 - 31
Computer modeling from solution data of spinach chloroplast and of Xenopus laevis somatic and oocyte 5 S rRNAs; Westhof E et al.; Detailed atomic models of a eubacterial 5 S rRNA (spinach chloroplast 5 S rRNA) and of a eukaryotic 5 S rRNA (somatic and oocyte 5 S rRNA from Xenopus laevis) were built using computer graphic . Both models integrate stereochemical constraints and experimental data on the accessibility of bases and phosphates towards several structure-specific probes . The base sequence was first inserted on to three-dimensional structural fragments picked up in a specially devised databank . The fragments were modified and assembled interactively on an Evans & Sutherland PS330 . Modeling was finalized by stereochemical and energy refinement . In spite of some uncertainty in the relative spatial orientation of the substructures, the broad features of the models can be generalized and several conclusions can be reached: (1) both models adopt a distorted Y-shape structure, with helices B and D not far from colinearity; (2) no tertiary interactions exist between loop c and region d or loop e; (3) the internal loops, in particular region d, contain several non-canonical base-pairs of A.A, U.U and A.G types; (4) invariant residues appear to be more important for protein or RNA binding than for maintaining the tertiary structure . The models are corroborated by footprinting experiments with ribosomal proteins and by the analysis of various mutants . Such models help to clarify the structure-function relationship of 5 S rRNA and are useful for designing site-directed mutagenesis experiments.

Science, 1989 May 12, 244(4905), 673 - 9
How old is the genetic code? Statistical geometry of tRNA provides an answer; Eigen M et al.; The age of the molecular organization of life as expressed in the genetic code can be estimated from experimental data . Comparative sequence analysis of transfer RNA by the method of statistical geometry in sequence space suggests that about one-third of the present transfer RNA sequence divergence was present at the urkingdom level about the time when archaebacteria separated from eubacteria . It is concluded that the genetic code is not older than, but almost as old as our planet . While this result may not be unexpected, it was not clear until now that interpretable data exist that permit inferences about such early stages of life as the establishment of the genetic code.

Proc Natl Acad Sci U S A, 1989 May, 86(9), 3031 - 5
A hydrogenase-linked gene in Methanobacterium thermoautotrophicum strain delta H encodes a polyferredoxin; Reeve JN et al.; The genes mvhDGA, which encode the subunit polypeptides of the methyl viologen-reducing hydrogenase in Methanobacterium thermoautotrophicum strain delta H, have been cloned and sequenced . These genes, together with a fourth open reading frame designated mvhB, are tightly linked and appear to form an operon that is transcribed starting 42 base pairs upstream of mvhD . The organization and sequences of the mvhG and mvhA genes indicate a common evolutionary ancestry with genes encoding the small and large subunits of hydrogenases in eubacterial species . The product of the mvhB gene is predicted to contain six tandomly repeated bacterial-ferredoxin-like domains and, therefore, is predicted to be a polyferredoxin that could contain as many as 48 iron atoms in 12 Fe4S4 clusters.

Mol Gen Genet, 1989 May, 217(1), 97 - 104
Characterization of the str operon genes from Spirulina platensis and their evolutionary relationship to those of other prokaryotes; Buttarelli FR et al.; A 5.3 kb DNA segment containing the str operon (ca . 4.5 kb) of the cyanobacterium Spirulina platensis has been sequenced . The str operon includes the structural genes rpsL (ribosomal protein S12), rpsG (ribosomal protein S7), fus (translation elongation factor EF-G) and tuf (translation elongation factor EF-Tu) . From the nucleotide sequence of this operon, the primary structures of the four gene products have been derived and compared with the available corresponding structures from eubacteria, archaebacteria and chloroplasts . Extensive homologies were found in almost all cases and in the order S12 greater than EF-Tu greater than EF-G greater than S7; the largest homologies were generally found between the cyanobacterial proteins and the corresponding chloroplast gene products . Overall codon usage in S . platensis was found to be rather unbiased.

J Mol Evol, 1989 May, 28(5), 418 - 26
Structural comparison of 26S rRNA-binding ribosomal protein L25 from two different yeast strains and the equivalent proteins from three eubacteria and two chloroplasts; Raue HA et al.; The sequences of Saccharomyces carlsbergensis ribosomal protein (r-protein) SL25* and its equivalents from Candida utilis (CL25), Escherichia coli (EL23), Bacillus stearothermophilus (BL23), Mycoplasma capricolum (ML23), Marchantia polymorpha chloroplasts (McpL23), and Nicotiana tabacum chloroplasts (NcpL23) were examined using a computer program that evaluates the extent of sequence similarity by calculating correlation coefficients for each pair of residues in two proteins from a number of physical properties of individual amino acids . Comparison matrices demonstrate that the prokaryotic sequences (including McpL23 and NcpL23) can be aligned unambiguously by introducing small internal deletions/insertions at three specific positions . A similar comparison brought to light a clear evolutionary relationship between the prokaryotic and the yeast proteins despite the fact that visual inspection of these sequences revealed only limited similarity . The alignment deduced from this comparison shows the two yeast r-proteins to have acquired a long (50-60 amino acids) N-terminal extension as well as a 13-amino acid-long deletion near the C-terminus . The significance of these findings in terms of the evolution of r-proteins in general and the biological function of various parts of the SL25 protein in particular is discussed.

FEBS Lett, 1989 Apr 24, 247(2), 259 - 62
Archaebacterial malate dehydrogenase: the amino-terminal sequence of the enzyme from Sulfolobus acidocaldarius is homologous to the eubacterial and eukaryotic malate dehydrogenases; Gorisch H et al.; 42 residues of the N-terminal amino acid sequence of malate dehydrogenase from the thermoacidophilic archaebacterium Sulfolobus acidocaldarius have been determined as VKVAFIGVGRGVGQTIAYNTIVNGYADEVMLYDVVPELPTKK . In eubacterial and eukaryotic enzymes this region is known to encompass residues involved in pyridine nucleotide binding . In the archaebacterial enzyme the residues Gly-7, Gly-11 and Asp-33 are also present . The data suggest that in the enzyme from S . acidocaldarius like in the other malate dehydrogenases the binding domain for NAD(H) is localized at the N-terminal part of the polypeptide chain . The archaebacterial enzyme is homologous to the other malate dehydrogenases, of which the amino acid sequences are known, however, it is only distantly related to the mitochondrial/E . coli group and the cytosolic/Thermus flavus group.

Eur J Biochem, 1989 Apr 15, 181(1), 41 - 6
Purification of isoleucyl-tRNA synthetase from Methanobacterium thermoautotrophicum by pseudomonic acid affinity chromatography; Rechsteiner T et al.; The isoleucyl-tRNA synthetase of the archaebacterium Methanobacterium thermoautotrophicum was purified 1500-fold to electrophoretic homogeneity by a procedure based on affinity chromatography on Sepharose-bound pseudomonic acid, a strong competitive inhibitor of this enzyme . The purified enzyme is a monomer with a molecular mass of 120 kDa . In this respect and in its Km values for the PPi-ATP exchange, and aminoacylation reactions, it resembles the isoleucyl-tRNA synthetases from eubacterial and eukaryotic sources . Its aminoacylation activity is optimal at pH 8.0 and at 55 degrees C . Pseudomonic acid is a strong competitive inhibitor of the aminoacylation reaction with respect to both L-isoleucine (KiIle 10 nM) and ATP (KiATP 20 nM).

FEBS Lett, 1989 Apr 10, 247(1), 147 - 53
The progenitor of ATP synthases was closely related to the current vacuolar H+-ATPase; Nelson H et al.; The gene encoding the proteolipid of the vacuolar H+-ATPase of yeast was cloned and sequenced . The deduced amino acid sequence of the yeast protein is highly homologous to that of the proteolipid from bovine chromaffin granules . In contrast to other membrane proteins the transmembrane segments of the bovine and yeast proteolipids were much more conserved than the hydrophilic parts . The fourth transmembrane segment, which contains the DCCD-binding site, was conserved 100% . Comparison of vacuolar and eubacterial proteolipids revealed a homology which pointed to a common ancestral gene that underwent gene duplication to form the vacuolar proteolipids . Additional support for this notion came from the amino acid sequences of subunits involved in the catalytic sectors of archaebacterial ATP synthase and plant and yeast vacuolar H+-ATPases, which reveal extensive sequence homology . Slight, but significant, homology between the archaebacterial and eubacterial ATP synthases was observed . These observations might suggest that the progenitor of ATP synthases was closely related to the present vacuolar H+-ATPases.

Akush Ginekol (Mosk), 1989 Apr, (4), 36 - 9
{Microbiological aspects of suppurative-inflammatory complications (pericultitis) after hysterectomy}; Danilov AIu; Bacteriologic findings in 101 patients before and after hysterectomy for benign tumors, inflammations, uterine prolapse are presented . Changes in aerobic and anaerobic vaginal and endocervical microflora species were demonstrated with respect to the phase of the cycle and the age of the patient . Pre- and postoperative genital mifroflora is represented by aerobic and anaerobic opportunity organisms, such as enterococcus, epidermal staphylococcus, E . coli, lactobacilli, eubacteria, and bacteroides . Postoperative complications are caused by associations of aerobic and anaerobic bacteria, with anaerobic organisms as the leading causative agent . Categories of risk with respect to infectious postoperative complications have been identified on the basis of microbiologic parameters.

J Bacteriol, 1989 Apr, 171(4), 2083 - 9
Identification and nucleotide sequence of the Leptospira biflexa serovar patoc trpE and trpG genes; Yelton DB et al.; Leptospira biflexa is a representative of an evolutionarily distinct group of eubacteria . In order to better understand the genetic organization and gene regulatory mechanisms of this species, we have chosen to study the genes required for tryptophan biosynthesis in this bacterium . The nucleotide sequence of the region of the L . biflexa serovar patoc chromosome encoding the trpE and trpG genes has been determined . Four open reading frames (ORFs) were identified in this region, but only three ORFs were translated into proteins when the cloned genes were introduced into Escherichia coli . Analysis of the predicted amino acid sequences of the proteins encoded by the ORFs allowed us to identify the trpE and trpG genes of L . biflexa . Enzyme assays confirmed the identity of these two ORFs . Anthranilate synthase from L . biflexa was found to be subject to feedback inhibition by tryptophan . Codon usage analysis showed that there was a bias in L . biflexa towards the use of codons rich in A and T, as would be expected from its G + C content of 37% . Comparison of the amino acid sequences of the trpE gene product and the trpG gene product with corresponding gene products from other bacteria showed regions of highly conserved sequence.

J Bacteriol, 1989 Apr, 171(4), 1967 - 73
Cyanobacterial RNA polymerase genes rpoC1 and rpoC2 correspond to rpoC of Escherichia coli; Xie WQ et al.; The DNA-dependent RNA polymerase (ribonucleoside triphosphate:RNA nucleotidyltransferase, EC 2.7.7.6) of cyanobacteria contains a unique core component, gamma, which is absent from the RNA polymerases of other eubacteria (G . J . Schneider, N . E . Tumer, C . Richaud, G . Borbely, and R . Haselkorn, J . Biol . Chem . 262:14633-14639, 1987) . We present the complete nucleotide sequence of rpoC1, the gene encoding the gamma subunit, from the heterocystous cyanobacterium Nostoc commune UTEX 584 . The derived amino acid sequence of gamma (621 residues) corresponds with the amino-terminal portion of the beta' polypeptide of Escherichia coli RNA polymerase . A second gene in N . commune UTEX 584, rpoC2, encodes a protein which shows correspondence with the carboxy-terminal portion of the E . coli beta' subunit . The rpoBC1C2 genes of N . commune UTEX 584 are present in single copies and are arranged in the order rpoBC1C2, and the coding regions are separated by short AT-rich spacer regions which have the potential to form very stable secondary structures . Our data indicate the occurrence of divergent evolution of structure in the eubacterial DNA-dependent RNA polymerase.

J Biol Chem, 1989 Mar 15, 264(8), 4423 - 7
Purification and properties of phloroglucinol reductase from Eubacterium oxidoreducens G-41; Haddock JD et al.; Phloroglucinol reductase was purified 90-fold to homogeneity from the anaerobic rumen organism Eubacterium oxidoreducens strain G-41 . The enzyme is stable in the presence of air and is found in the soluble fraction after ultracentrifugation of cell extract . Ion-exchange, hydrophobic interaction, and affinity chromatography were used to purify the enzyme . The native Mr is 78,000, and the subunit Mr is 33,000 indicating an alpha 2 homodimer . The enzyme is specific for phloroglucinol and NADPH . The Km and Vmax are 600 microM and 640 mumol min-1 mg-1 (pH 7.2) for phloroglucinol, and 6.7 microM and 550 mumol min-1 mg-1 (pH 6.8) for NADPH; the Km and Vmax for the reverse direction are 290 microM and 140 mumol min-1 mg-1 (pH 7.2) for dihydrophloroglucinol, and 27 microM and 220 mumol min-1 mg-1 (pH 7.2) for NADP . Temperature and pH optima are 40 degrees C and 7.8 in the forward direction . The pure enzyme is colorless in solution and flavins are absent . Analysis for cobalt, manganese, molybdenum, vanadium, tungsten, selenium, copper, nickel, iron, and zinc indicated that these metals are not components of the phloroglucinol reductase . Cupric chloride, n-ethylmaleimide, and p-chloromercuribenzoate are potent inhibitors of enzyme activity . The properties of phloroglucinol reductase indicate that it functions in the pathway of anaerobic degradation of trihydroxybenzenes by catalyzing reduction of the aromatic nucleus prior to ring fission.

Nucleic Acids Res, 1989 Mar 11, 17(5), 1907 - 14
Transfer RNA genes frequently serve as integration sites for prokaryotic genetic elements; Reiter WD et al.; The DNA sequences were determined at the boundaries of the integrated copy of the archaebacterial genetic element SSV1 . A 44 bp sequence present as a single copy on the 15.5 kb circular SSV1 DNA flanked the integrated copy as a direct DNA sequence repeat, suggesting that SSV1 integration occurred by recombination between this 44 bp SSV1 sequence and an identical sequence on the bacterial chromosome . At the left attachment site, a region encompassing the 44 bp attachment core sequence and the 31 nucleotides upstream of it displayed all characteristics expected for an arginine tRNA gene . An analysis of published attachment site sequences of other systems revealed that tRNA genes also constitute the bacterial attachment site in the case of three temperate phages and two transmissible plasmids in eubacteria, indicating a widespread occurrence of tRNA genes as integration target sites . This finding may be important for the understanding of mechanisms and evolution of site-specific recombination.

Science, 1989 Mar 10, 243(4896), 1360 - 3
Phylogenetic stains: ribosomal RNA-based probes for the identification of single cells; DeLong EF et al.; Rapid phylogenetic identification of single microbial cells was achieved with a new staining method . Formaldehyde-fixed, intact cells were hybridized with fluorescently labeled oligodeoxynucleotides complementary to 16S ribosomal RNA (rRNA) and viewed by fluorescence microscopy . Because of the abundance of rRNA in cells, the binding of the fluorescent probes to individual cells is readily visualized . Phylogenetic identification is achieved by the use of oligonucleotides (length 17 to 34 nucleotides) that are complementary to phylogenetic group-specific 16S rRNA sequences . Appropriate probes can be composed of oligonucleotide sequences that distinguish between the primary kingdoms (eukaryotes, eubacteria, archaebacteria) and between closely related organisms . The simultaneous use of multiple probes, labeled with different fluorescent dyes, allows the identification of different cell types in the same microscopic field . Quantitative microfluorimetry shows that the amount of an rRNA-specific probe that binds to Escherichia coli varies with the ribosome content and therefore reflects growth rate.

J Bacteriol, 1989 Mar, 171(3), 1744 - 6
Mutation of Bacillus firmus OF4 to duramycin resistance results in substantial replacement of membrane lipid phosphatidylethanolamine by its plasmalogen form; Clejan S et al.; Mutant strains of alkalophilic Bacillus firmus OF4 that were selected for resistance to duramycin had greatly reduced levels of membrane diacylphosphatidylethanolamine, as had been found in studies of such mutants of Bacillus subtilis . In the B . firmus strains, however, substantial levels of plasmenylethanolamine were found . This is an unusual membrane component for an aerobic eubacterium, but the presence of trace amounts even in the wild type was confirmed in experiments with 32Pi-labeled growth medium . The membrane lipid composition of the duramycin-resistant strains had several other changes that also left alkalophilic growth unimpaired.

J Endod, 1989 Mar, 15(3), 112 - 6
Antimicrobial susceptibilities of Eubacterium, Peptostreptococcus, and Bacteroides isolated from root canals of teeth with periapical pathosis; Yamamoto K et al.; Eubacterium, Peptostreptococcus, and Bacteroides were isolated in high frequency from root canals with acute periapical inflammation . The antimicrobial susceptibilities of these strains were studied by determining minimum inhibitory concentrations of different agents . Although all three kinds of isolates were susceptible to penicillins, the isolates other than black-pigmented Bacteroides were less susceptible to cephems, tetracyclines, and macrorides with several resistant strains . All strains were uniformly resistant to aminoglycosides . Some differences in susceptibilities were observed among species of Eubacterium and Peptostreptococcus, while penicillins were effective for both species . Black-pigmented Bacteroides showed good susceptibilities to all agents except for aminoglycosides . The susceptibility of Bacteroides gingivalis was superior to that of Bacteroides intermedius . There were many resistant strains in non-black-pigmented but not in black-pigmented Bacteroides isolates . Penicillins were the most effective for Eubacterium, Peptostreptococcus, and Bacteroides, indicating that penicillins are suitable for treatment of root canals with acute apical periodontitis.

J Bacteriol, 1989 Mar, 171(3), 1524 - 30
Mutations in the Escherichia coli fnr and tgt genes: control of molybdate reductase activity and the cytochrome d complex by fnr; Frey B et al.; In eubacteria, the tRNA transglycosylase (Tgt) in specific tRNAs exchanges a guanine in the anticodon for 7-aminomethyl-7-deazaguanine, which is finally converted to queuosine . The tgt gene of Escherichia coli has been mapped at 9 min on the genome, and mutant pairs containing an intact or mutated tgt allele were obtained after transduction of the tgt locus by P1 bacteriophages into a genetically defined E . coli strain (S . Noguchi, Y . Nishimura, Y . Hirota, and S . Nishimura, J . Biol . Chem . 257:6544-6550, 1982) . These tgt mutants grew anerobically with fumarate as an electron acceptor, while nitrate or trimethylamine N-oxide could not be reduced . Furthermore, molybdate reductase activity was almost lacking and the characteristic absorption maxima, corresponding to cytochrome a1 and the cytochrome d complex, were not detectable in low-temperature reduced-minus-oxidized difference spectra in anaerobically grown cells . Transduction of the mutated tgt locus into another E . coli recipient resulted in tgt mutants without anaerobic defects . Transformation of the original tgt mutants with an fnr gene-containing plasmid reversed the anaerobic defects . Clearly, the original tgt mutants harbor a second mutation, affecting the anaerobic regulator protein Fnr . The results suggest that fnr is involved in anaerobic control of components of the cytochrome d complex and of the redox system that transfers electrons to molybdate . F' plasmids containing a fused lacI-lacZ gene with the nonsense codon UAG at different positions in the lacI part were transferred to E . coli strains with a mutated or nonmutated tgt locus but intact in fnr . A twofold increase in the frequency of incorrect readthrough of the UAG codon, dependent on the codon context, was observed in the tgt mutant and is suggested to be caused by a tRNA(Tyr) with G in place of queuosine.

Mol Gen Genet, 1989 Feb, 215(3), 381 - 7
Structure of the dnaA region of Pseudomonas putida: conservation among three bacteria, Bacillus subtilis, Escherichia coli and P . putida; Fujita MQ et al.; We have cloned from Pseudomonas putida a gene homologous to Escherichia coli dnaA, and determined the sequence of the gene and its neighboring region . The dnaA gene and at least three other genes, dnaN, recF and gyrB, were found to be highly homologous to the genes in the dnaA regions of the E . coli and Bacillus subtilis chromosomes . A non-translatable region of some 600 bp immediately upstream of the dnaA gene is also conserved in the three bacteria and contains 3, 12, and 14 DnaA-boxes (TTATCCACA and closely related sequences) in E . coli, P . putida and B . subtilis, respectively . The present results confirm our hypothesis that the dnaA region is the replication origin region of the ancestral bacterium and that the essential feature of the dnaA and DnaA-box combination is conserved in most eubacteria and plays a central role in initiation of chromosomal replication.

Biochim Biophys Acta, 1989 Jan 23, 1007(1), 36 - 43
Isolation and characterization of DNA-binding proteins from the cyanobacterium Synechococcus sp . PCC 7002 (Agmenellum quadruplicatum) and from spinach chloroplasts; Crevel G et al.; Basic, low-molecular-weight DNA-binding proteins were isolated from the unicellular cyanobacterium Synechococcus sp . PCC 7002 (Agmenellum quadruplicatum) and from the chloroplasts of spinach (Spinacia oleacera) . In Synechococcus, two major proteins which bind to double-strand DNA (10 and 16 kDa, respectively) were purified . The 10 kDa protein, named HAq, resembles strongly, in amino-acid composition, eubacterial HU-type proteins . The 16 kDa protein is slightly basic . Its characteristics are compared to those of E . coli protein H1 and 17K . In spinach chloroplasts, a major protein HC (10 kDa), which also binds to ds-DNA, was purified . As observed for known archaebacterial and mitochondrial DNA-binding proteins, its amino-acid composition differs significantly from those of eubacterial HU . The comparison of the amino-terminal sequence (27 residues) with other chloroplast peptidic sequences is discussed.

FEMS Microbiol Lett, 1989 Jan 15, 48(2), 125 - 8
Cloning and restriction analysis of ribosomal RNA genes from Mycobacterium smegmatis; Bercovier H et al.; A genomic library of Mycobacterium smegmatis DNA was constructed in phage EMBL3 . A clone (gamma HB85) containing rRNA genes was isolated using as probes, fragments of E . coli rRNA cistron B . This cloned DNA fragment was mapped by restriction analysis and was shown to contain one complete set of rRNA genes (rRNA A) . The physical mapping of the second set of rRNA genes of M . smegmatis (rRNA B) was done by restriction analysis of total chromosomal DNA . The two sets of rRNA genes showed highly conserved restriction sites within the respective sets but not in the flanking regions . The two rRNA sets of genes are organised like in the other eubacteria in the order 16S-23S-5S.

Nucleic Acids Res, 1989 Jan 11, 17(1), 31 - 6
The 4.5S RNA gene from Pseudomonas aeruginosa; Toschka HY et al.; The essential 4.5S RNA of Escherichia coli contains a structural motif, which is also present in RNAs from other organisms, i.e . Bacillus subtilis scRNA, Halobacterium halobium 7S RNA and eukaryotic 7SL RNAs . This suggests a common function in all organisms, which could be related to protein translocation, since 7SL RNA is essential for this process in eukaryotes . We have analysed the structure and expression of the 4.5S RNA gene from another gram-negative eubacterium, Pseudomonas aeruginosa . The single copy gene encodes a 113 nucleotides long RNA, which shares 75% sequence homology to the E . coli 4.5S RNA and also exhibits the completely conserved hairpin structure of the corresponding RNAs of B . subtilis and E . coli . Transcription initiates 24 nucleotides upstream from the mature 5' end and exceeds beyond the 4.5S RNA coding region . A distal open reading frame, similar to that described for E . coli, does not exist downstream from the P . aeruginosa 4.5S RNA gene.

Science, 1989 Jan 6, 243(4887), 75 - 7
Phylogenetic meaning of the kingdom concept: an unusual ribosomal RNA from Giardia lamblia; Sogin ML et al.; An analysis of the small subunit ribosomal RNA (16S-like rRNA) from the protozoan Giardia lamblia provided a new perspective on the evolution of nucleated cells . Evolutionary distances estimated from sequence comparisons between the 16S-like rRNAs of Giardia lamblia and other eukaryotes exceed similar estimates of evolutionary diversity between archaebacteria and eubacteria and challenge the phylogenetic significance of multiple eukaryotic kingdoms . The Giardia lamblia 16S-like rRNA has retained many of the features that may have been present in the common ancestor of eukaryotes and prokaryotes.

Rev Infect Dis, 1989 Jan-Feb, 11 Suppl 1, S68 - 73; discussion S73-4
Sequential assessment of vaginal microflora in healthy women randomly assigned to tampon or napkin use; Chow AW et al.; The effect of tampon usage on the vaginal microflora of 35 healthy women was determined following their random allocation to either tampon or napkin use for three consecutive menstrual cycles . Sequential and semiquantitative vaginal cultures were obtained on days 3 +/- 2, 15 +/- 2, and 25 +/- 2 of the menstrual cycle (day 1, first day of menses) before and after randomization . Before randomization, the rate of isolation and median counts of facultative lactobacilli were significantly higher (P less than .05) and that of eubacteria was significantly lower (P = .026) among regular tampon users than among exclusive napkin users . After randomization, only median counts of coagulase-negative staphylococci were significantly increased (P = .025) during tampon use compared with the rates for the same women during napkin use . These shifts in vaginal microflora occurred only in samples obtained during menstruation and not in those obtained at other sampling times . The data presented here support the notion that the use of tampons may result in alterations in the autochthonous vaginal microflora . It remains to be determined if these ecologic shifts in the vaginal microflora may adversely affect resistance to colonization by potential pathogens in the lower female genital tract.

J Bacteriol, 1989 Jan, 171(1), 93 - 8
Isolation and characterization of an archaebacterial viruslike particle from Methanococcus voltae A3; Wood AG et al.; Small amounts of a 23-kilobase covalently closed circular DNA molecule were isolated from unwashed cells of Methanococcus voltae A3 . Further investigation indicated the presence of greater quantities of the circular DNA in the culture supernatant, complexed with protein in a manner rendering the DNA resistant to DNase . Electron-microscopic examination of supernatant material revealed the presence of particles which morphologically resemble virus . Phenol extraction of viruslike particle preparations resulted in the recovery of DNase-sensitive open-circular DNA molecules . As many as 30 viruslike particles per cell were recovered from some cultures . Hybridization data clearly indicated the presence of a chromosomally integrated copy of the viruslike particle DNA . Although M . voltae PS was not observed to produce viruslike particles, DNA homologous to the viruslike particle DNA was detected in its chromosome . A mutant of M . voltae A3 was isolated which produced no particles; its DNA was deleted for 80% of the integrated viruslike particle DNA . Despite any similarities to lysogenic bacteriophages of eubacteria, neither infectivity nor inducibility of the viruslike particles could be demonstrated.

Arch Microbiol, 1989, 152(2), 206 - 8
Nucleotide sequence of 16S rRNA and phylogenetic position of the green sulfur bacterium Clathrochloris sulfurica; Witt D et al.; The almost complete primary structure of the 16S rRNA from the green sulfur bacterium "Clathrochloris sulfurica" was determined by reverse transcriptase sequencing . Comparison of defined invariable parts of the molecule from representatives of 9 major lines of descent from the eubacterial kingdom shows C . sulfurica to be highly related to Chlorobium vibrioforme . The relationship between "Clathrochloris" and Chlorobium is in accord with the present allocation of these two genera into the family Chlorobiaceae.

Arch Microbiol, 1989, 151(4), 353 - 8
Chemical composition of Eubacterium nodatum cell wall peptidoglycan; Severin AI et al.; The structure of Eubacterium nodatum cell wall peptidoglycan was investigated . The peptide subunit of E . nodatum peptidoglycan has the following structure: L-Ala-D-Glu (Gly)-L-Orn-D-Ala . The carboxyl group of alanine occupying position 4 is attached to the delta-amino group of ornithine of an other subunit by the cross-linking bridge L-Ala-L-Ala-L-Orn . All glycine molecules are connected with the alpha-carboxyl group of glutamic acid with the ratio being 0.5-1 . The hydrolysis of E . nodatum peptidoglycan by the S . albus G enzyme proceeds primarily due to the activity of alanyl-alanine endopeptidase, ornithyl-ornithine endopeptidase, ornithyl-alanine endopeptidase, N-acetyl-muramyl-alanine amidase, N-acetylmuramidase and N-acetylglucosaminidase.

Arch Microbiol, 1989, 151(4), 348 - 52
Chemical composition of Eubacterium alactolyticum cell wall peptidoglycan; Severin AI et al.; The mechanism of lysis of Eubacterium alactolyticum cell walls by Streptomyces albus G enzyme was studied . The analysis of the peptide terminal groups and peptide subunits isolated from the cell wall digest, released during solubilization of the cell walls, revealed that lytic action of S . albus G enzyme was mainly due to D-alanyl-A2pm endopeptidase, N-acetylmuramyl-L-alanine amidase, N-acetylmuramidase and N-acetylglucosaminidase . E . alactolyticum cell wall peptidoglycan is composed mainly of glucosamine, muramic acid, D-glutamic acid, L- and D-alanine, meso-diaminopimelic acid and glycine . The peptide subunit consists of L-alanyl-D-glutamyl-meso-A2pm-D-alanine . D-Alanine is connected directly with the amino group of the meso-A2pm residue of another peptide subunit . All of the L-amino groups of meso-diaminopimelic acid are involved in cross-linking . The possible structure of the peptide moiety of E . alactolyticum cell wall peptidoglycan is presented.

Can J Microbiol, 1989 Jan, 35(1), 171 - 5
The expression of the superoxide dismutase gene in Halobacterium cutirubrum and Halobacterium volcanii; May BP et al.; The gene encoding the Mn-containing superoxide dismutase (SOD) from Halobacterium cutirubrum has been cloned and sequenced . The deduced amino acid sequence is homologous to the sequences of Fe and Mn SODs from eubacteria . The high degree of amino acid identity between the archaebacterial and eubacterial proteins suggests that a SOD gene may have been laterally transferred between eubacteria and archaebacteria sometime after the accumulation of atmospheric oxygen . Consensus elements of halobacterial promoters are found upstream of the coding region, however, the spacing between them and the transcription start site is greater than in other genes . Termination of transcription occurs in five consecutive T residues that are preceded by a GC-rich sequence that has short inverted repeats . In addition to the authentic SOD gene, H . cutirubrum also contains a putative pseudogene . The SOD levels and growth rates of H . cutirubrum and Halobacterium volcanii were tested in response to treatment by paraquat, an intracellular generator of superoxide . In H . volcanii the growth rate slowed, and SOD was strongly induced throughout prolonged treatment with paraquat . In H . cutirubrum the same effects were noticed initially, but after 48 h exposure to the drug, the growth rate increased and the SOD level decreased . Production of paraquat resistant mutants of H . cutirubrum may play a part in this process, however, some type of physiological adaptation is also probably required.

Appl Environ Microbiol, 1989 Jan, 55(1), 11 - 6
Development of a differential medium for bile salt hydrolase-active Lactobacillus spp; Dashkevicz MP et al.; An agar plate assay was developed to detect bile salt hydrolase activity in lactobacilli . On Lactobacillus-selective MRS or Rogosa SL medium supplemented with taurodeoxycholic, taurocholic, or taurochenodeoxycholic acids, bile salt hydrolysis was manifested at two intensities: (i) the formation of precipitate halos around colonies or (ii) the formation of opaque granular white colonies . Sixty-six lactobacilli were tested for bile salt hydrolase activity by both the plate assay and a sensitive radiochemical assay . No false-positive or false-negative results were detected by the plate assay . Based on results of experiments with Eubacterium lentum and Bacteroides species, the plate assay was dependent on two factors: (i) the presence of bile salt hydrolytic activity and (ii) the ability of the organism to sufficiently acidify the medium to protonate free bile acids . The availability of a differential medium for determination of bile salt hydrolase activity will provide a rapid method for determining shifts in a specific functional activity of intestinal Lactobacillus species and provide a rapid screening capability for identifying bile salt hydrolase-deficient mutants . The latter application should allow bile salt hydrolase activity to be used as a marker enzyme in genetic experiments.

Can J Microbiol, 1989 Jan, 35(1), 200 - 4
Gene organization and structure of two transcriptional units from Methanococcus coding for ribosomal proteins and elongation factors; Auer J et al.; Two transcriptional units coding for ribosomal proteins and protein synthesis elongation factors in Methanococcus vannielii have been cloned and analysed in detail . They correspond to the "streptomycin operon" and "spectinomycin operon" of the Escherichia coli chromosome . The following general conclusions can be drawn from comparison of the nucleotide and the derived amino acid sequences of ribosomal proteins from Methanococcus with those from eubacteria and eukaryotes . (i) Ribosomal protein and elongation factor genes in Methanococcus are clustered in transcriptional units corresponding closely to E . coli ribosomal protein operons with respect to both gene composition and organization . (ii) These transcriptional units contain, in addition, a few open reading frames whose putative gene products share sequence similarity with eukaryotic 80S but not with eubacterial, ribosomal proteins . They may correspond to "additional" ribosomal proteins of the Methanococcus ribosome, there being no functional homologues in the eubacterial ribosome . (iii) Methanococcus ribosomal proteins and elongation factors almost exclusively exhibit a higher sequence similarity to eukaryotic 80S ribosomal proteins than to those of eubacteria . (iv) Many Methanococcus ribosomal proteins have a size intermediate between those of their eukaryotic and eubacterial homologues . These results are discussed in terms of a hypothesis which implies that the recent eubacterial ribosome developed by a "minimization" process from a more complex organelle and that the archaebacterial ribosome has maintained features of this ancestor.

Can J Microbiol, 1989 Jan, 35(1), 73 - 80
The phylogenetic relations of DNA-dependent RNA polymerases of archaebacteria, eukaryotes, and eubacteria; Zillig W et al.; Unrooted phylogenetic dendrograms were calculated by two independent methods, parsimony and distance matrix analysis, from an alignment of the derived amino acid sequences of the A and C subunits of the DNA-dependent RNA polymerases of the archaebacteria Sulfolobus acidocaldarius and Halobacterium halobium with 12 corresponding sequences including a further set of archaebacterial A+C subunits, eukaryotic nuclear RNA polymerases, pol I, pol II, and pol III, eubacterial beta' and chloroplast beta' and beta" subunits . They show the archaebacteria as a coherent group in close neighborhood of and sharing a bifurcation with eukaryotic pol II and (or) pol IIIA components . The most probable trees show pol IA branching off from the tree separately at a bifurcation with the eubacterial beta' lineage . The implications of these results, especially for understanding the possibly chimeric origin of the eukaryotic nuclear genome, are discussed.

Can J Microbiol, 1989 Jan, 35(1), 228 - 33
Studies on DNA polymerases and topoisomerases in archaebacteria; Forterre P et al.; We have isolated DNA polymerases and topoisomerases from two thermoacidophilic archaebacteria: Sulfolobus acidocaldarius and Thermoplasma acidophilum . The DNA polymerases are composed of a single polypeptide with molecular masses of 100 and 85 kDa, respectively . Antibodies against Sulfolobus DNA polymerase did not cross react with Thermoplasma DNA polymerase . Whereas the major DNA topoisomerase activity in S . acidocaldarius is an ATP-dependent type I DNA topoisomerase with a reverse gyrase activity, the major DNA topoisomerase activity in T . acidophilum is a ATP-independent relaxing activity . Both enzymes resemble more the eubacterial than the eukaryotic type I DNA topoisomerase . We have found that small plasmids from halobacteria are negatively supercoiled and that DNA topoisomerase II inhibitors modify their topology . This suggests the existence of an archaebacterial type II DNA topoisomerase related to its eubacterial and eukaryotic counterparts . As in eubacteria, novobiocin induces positive supercoiling of halobacterial plasmids, indicating the absence of a eukaryotic-like type I DNA topoisomerase that relaxes positive superturns.

Can J Microbiol, 1989 Jan, 35(1), 81 - 5
Sequence comparison of glyceraldehyde-3-phosphate dehydrogenases from the three urkingdoms: evolutionary implication; Hensel R et al.; The primary structure of the glyceraldehyde-3-phosphate dehydrogenase from the archaebacteria shows striking deviation from the known sequences of eubacterial and eukaryotic sequences, despite unequivocal homologies in functionally important regions . Thus, the structural similarity between the eubacterial and eukaryotic enzymes is significantly higher than that between the archaebacterial enzymes and the eubacterial and eukaryotic enzymes . This preferred similarity of eubacterial and eukaryotic glyceraldehyde-3-phosphate dehydrogenase structures does not correspond to the phylogenetic distances among the three urkingdoms as deduced from comparisons of ribosomal ribonucleic acid sequences . Indications will be presented that the closer relationship of the eubacterial and eukaryotic glyceraldehyde-3-phosphate dehydrogenase resulted from a gene transfer from eubacteria to eukaryotes after the segregation of the three urkingdoms.

Can J Microbiol, 1989 Jan, 35(1), 52 - 7
Conserved gene structures and expression signals in methanogenic archaebacteria; Allmansberger R et al.; A comparative analysis of cotranscribed gene clusters comprising the structural genes mcrA, mcrB, mcrC, mcrD, and mcrG was carried out in three species of methanogens . mcrA, mcrB, and mcrG are the structural genes for the three subunits of methyl coenzyme M reductase, while the two other genes encode polypeptides of unknown functions . The degree of conservation of the mcr gene products among different species of methanogens varies . No correlation was found between the conservation of the G+C contents of the homologous genes and of the amino acid sequences of their products among the different bacteria . The comparison of RNA polymerase core subunit genes of Methanobacterium thermoautotrophicum as evolutionary markers with their equivalents in Escherichia coli, Saccharomyces cerevisiae, and Drosophila melanogaster showed that homologous polypeptide domains are encoded by different numbers of genes suggesting gene fusion of adjacent genes in the course of evolution . The archaebacterial subunits exhibit much stronger homology with their eukaryotic than with their eubacterial equivalents on the polypeptide sequence level . All the analyzed genes are preceded by ribosome binding sites of eubacterial type . In addition to known putative promoter sequences, conserved structural elements of the DNA were detected surrounding the transcription initiation sites of the mcr genes.

Can J Microbiol, 1989 Jan, 35(1), 234 - 44
Structure and evolution of the L11, L1, L10, and L12 equivalent ribosomal proteins in eubacteria, archaebacteria, and eucaryotes; Ramirez C et al.; The genes corresponding to the L11, L1, L10, and L12 equivalent ribosomal proteins (L11e, L1e, L10e, and L12e) of Escherichia coli have been cloned and sequenced from two widely divergent species of archaebacteria, Halobacterium cutirubrum and Sulfolobus solfataricus, and the L10 and four different L12 genes have been cloned and sequenced from the eucaryote Saccharomyces cerevisiae . Alignments between the deduced amino acid sequences of these proteins and to other available homologous proteins of eubacteria and eucaryotes have been made . The data suggest that the archaebacteria are a distinct coherent phylogenetic group . Alignment of the proline-rich L11e proteins reveals that the N-terminal region, believed to be responsible for interaction with release factor 1, is the most highly conserved region and that there is specific conservation of most of the proline residues, which may be important in maintaining the highly elongated structure of the molecule . Although L11 is the most highly methylated protein in the E . coli ribosome, the sites of methylation are not conserved in the archaebacterial L11e proteins . The L1e proteins of eubacteria and archaebacteria show two regions of very high similarity near the center and the carboxy termini of the proteins . The L10e proteins of all kingdoms are colinear and contain approximately three fourths of an L12e protein fused to their carboxy terminus, although much of this fusion has been lost in the truncated eubacterial protein . The archaebacterial and eucaryotic L12e proteins are colinear, whereas the eubacterial protein has suffered a rearrangement through what appear to be gene fusion events . Within the L12e derived region of the L10e proteins there exists a repeated module of 26 amino acids, present in two copies in eucaryotes, three in archaebacteria, and one in eubacteria . This modular sequence is apparently also present in the L12e proteins of all kingdoms and may play a role in L12e dimerization, L10e-L12e complex formation, and the function of the L10e-L12e complex in translation.

Can J Microbiol, 1989 Jan, 35(1), 164 - 70
Organization of genes encoding the L11, L1, L10, and L12 equivalent ribosomal proteins in eubacteria, archaebacteria, and eucaryotes; Shimmin LC et al.; Archaebacterial and eucaryotic cytoplasmic ribosomes contain proteins equivalent to the L11, L1, L10, and L12 proteins of the eubacterium Escherichia coli . In E . coli the genes encoding these ribosomal proteins are clustered, cotranscribed, and autogenously regulated at the level of mRNA translation . Genomic restriction fragments encoding the L11e, L1e, L10e, and L12e (equivalent) proteins from two divergent archaebacteria . Halobacterium cutirubrum and Sulfolobus solfataricus, and the L10e and L12e proteins from the eucaryote Saccharomyces cerevisiae have been cloned, sequenced, and analyzed . In the archaebacteria, as in eubacteria, the four genes are clustered and the L11e, L1e, L10e, and L12e order is maintained . The transcription pattern of the H . cutirubrum cluster is different from the E . coli pattern and the flanking genes on either side of the tetragenic clusters in E . coli, H . cutirubrum, and Sulfolobus solfataricus are all unrelated to each other . In the eucaryote Saccharomyces cerevisiae there is a single L10e gene and four separate L12e genes that are designated L12eIA, L12eIB, L12eIIA, and L12eIIB . These five genes are not closely linked and each is transcribed as a monocistronic mRNA; the L10e, L12eIA, L12eIB, and the L12eIIA genes are contiguous and uninterrupted, whereas the L12eIIB gene is interrupted by a 301 nucleotide long intron located between codons 38 and 39.

Can J Microbiol, 1989 Jan, 35(1), 119 - 23
A brief note concerning archaebacterial phylogeny; Olsen GJ et al.; Critical analysis of the recently proposed alternative to the normal archaebacterial tree, the new eocyte tree, shows that the latter's central topology, in which the eubacteria branch from an entirely different section of the unrooted archaebacterial tree than the eukaryotes, is consistent with an artifact . The effects of the alignment used and the particular composition of the sequence quartets analyzed to infer this tree are discussed in detail.

Can J Microbiol, 1989 Jan, 35(1), 11 - 20
Comparative studies of ribosomal proteins and their genes from Methanococcus vannielii and other organisms; Kopke AK et al.; Using data from a partial protein sequence analysis of ribosomal proteins derived from the archaebacterium Methanococcus vannielii, oligonucleotide probes were synthesized . The probes enabled us to localize several ribosomal protein genes and to determine their nucleotide sequences . The amino acid sequences that were deduced from the genes correspond to proteins L12 and L10 from the rif operon, according to the genome organization in Escherichia coli, and to proteins L23 and L2, which have comparable locations, as in the Escherichia coli S10 operon . Various degrees of similarity were found when the four proteins were compared with the corresponding ribosomal proteins of prokaryotic or eukaryotic organisms . The highest sequence homology was found in counterparts from other archaebacteria, such as Halobacterium marismortui, Halobacterium halobium, or Sulfolobus . In general, the M . vannielii protein sequences were more related to the eukaryotic kingdom than to the Gram-positive or Gram-negative eubacteria . On the other hand, the organization of the ribosomal protein genes clearly follows the operon structure of the Escherichia coli genome and is different from the monocistronic eukaryotic gene arrangements . The protein coding regions were not interrupted by introns . Furthermore, the Shine-Dalgarno type sequences of methanogenic bacteria are homologous with those of eubacteria, and also their terminator regions are similar.

J Bacteriol, 1989 Jan, 171(1), 581 - 4
Duplication of the tuf gene: a new insight into the phylogeny of eubacteria; Sela S et al.; The conservation and duplication of the tuf gene encoding the elongation factor EF-Tu were used to define phylogenetic relationships among eubacteria . When the tufA gene of Escherichia coli was used as a probe in hybridization experiments, duplicate tuf genes were found in gram-negative bacteria from three major phyla: purple bacteria, bacteroides, and cyanobacteria . Only a single copy of tuf was found in gram-positive bacteria, including mycobacteria and mycoplasmas . Gram-positive clostridia were found to carry two copies of tuf.

Acta Microbiol Pol, 1989, 38(3-4), 207 - 15
The physiological role of eubacterial histone-like proteins; Paterczyk B; This minireview reflects the change in views on the role of histone-like proteins which has occurred in the 1980 s . Initially these proteins were regarded as analogous to eukaryotic histones though distinguished from the latter by much lower strength of binding to DNA . This was attributed to the greater dynamics of the structure of bacterial chromatin . The accumulation in recent years of results testifying to the absence of protein HU in central region of the nucleoid and the participation of this protein in almost all processes involving the recognition of specific DNA sequences by regulatory proteins forces the rejection of the concept of "histone-likeness" and favors the concept of a general, non-specific enhancer of the recognition of sequences acting by causing changes in the secondary structure of a given DNA region.

Can J Microbiol, 1989 Jan, 35(1), 153 - 9
Ribosomal protein gene cluster of Halobacterium halobium: nucleotide sequence of the genes coding for S3 and L29 equivalent ribosomal proteins; Spiridonova VA et al.; A 1643 base pair fragment encoding the S3 and L29 equivalent ribosomal proteins has been sequenced from the archaebacterium Halobacterium halobium . The incomplete open reading frame present upstream from the S3 gene encodes a protein homologous to the eubacterial ribosomal protein L22 . The initiation codons of the S3 and L29 genes overlap with the termination codons of the upstream genes . A tight physical organization suggests that these genes are transcribed as a polycistronic operon . Peculiarities of the protein structure and gene organization are discussed.

FEMS Microbiol Lett, 1989 Jan 1, 48(1), 19 - 24
Broad range DNA probes for detecting and amplifying eubacterial nucleic acids; Chen K et al.; In this report we describe and characterize two oligomer probes that are broadly homologous to conserved eubacterial 16S ribosomal RNA (rRNA) sequences not present in human 18 rRNA or human mitochondrial 12S rRNA . One or both of the probes can detect all of 23 phylogenetically diverse eubacterial nucleic acids against which they were tested by dot blot hybridization . A sensitivity of about 1 bacterium per 10 eukaryotic cells was achieved . By using these oligomer sequences or their complements as primers in the polymerase chain reaction (PCR), the equivalent of 1 pg of E . coli DNA was detected in the presence of a large excess of eukaryotic DNA . Information useful for partial phylogenetic classification of detected organisms may be obtained by direct sequence analysis of the amplified DNA and comparison with known sequences or catalogs . Such broadly homologous probes offer advantages over more narrowly specific probes for detecting organisms whose identity is unknown . They could thus be employed for recognizing infection by organisms that cannot be cultured as may occur, for example, in tissue culture or in plant or animal diseases of unknown cause, provided the probes fail to hybridize with host nucleic acids.

J Bacteriol, 1989 Jan, 171(1), 272 - 9
Nucleotidylation, not phosphorylation, is the major source of the phosphotyrosine detected in enteric bacteria; Foster R et al.; The majority of the phosphotyrosine recovered from partial acid hydrolysates of 32P-labeled Escherichia coli is derived from a single prominent protein . We show here by biochemical, genetic, and immunological criteria that this protein is actually glutamine synthetase adenylylated (not phosphorylated) at tyrosine . Furthermore, all of the phosphotyrosine detectable in partial acid hydrolysates of 32P-labeled Salmonella typhimurium was eliminated in a strain deficient in both glutamine synthetase and uridylyltransferase, an enzyme which uridylylates the regulatory protein PII at a tyrosine residue . These results suggest that protein-tyrosine phosphorylation represents a rare modification in eubacterial cells.

Infect Immun, 1989 Jan, 57(1), 245 - 54
Lipid-modified surface protein antigens expressing size variation within the species Mycoplasma hyorhinis; Boyer MJ et al.; Monoclonal antibodies (MAbs) previously shown to recognize distinct epitopes selectively expressed on the surface of some Mycoplasma hyorhinis strains were used to define two discrete sets of lipid-modified membrane surface proteins showing marked size variation within this species . Sodium dodecyl sulfate-polyacrylamide gel electrophoresis and immunoblot analysis of Triton X-114 phase-fractionated proteins from six isolates of M . hyorhinis defined a set of amphiphilic integral membrane proteins of 23, 50, and 55 kilodaltons (kDa) recognized on respective isolates by one MAb and a second set of integral proteins of 88, 120, and 100 to 150 kDa recognized by another MAb . The first group of proteins all contained a common, amphiphilic 18-kDa limit tryptic polypeptide bearing the epitope . The size- and strain-variant surface antigens identified by the MAbs were shown to be lipid-modified proteins . Phase fractionation of {3H}palmitate-labeled organisms revealed numerous 3H-labeled proteins in all isolates, which partitioned exclusively into the hydrophobic phase . These proteins generally showed pronounced size variation among isolates and included the antigen variants recognized by the two MAbs, as demonstrated directly by immunoprecipitation of correspondingly sized 3H-labeled proteins from each isolate . A third MAb recognized an invariant, lipid-associated surface protein of 70 kDa on all M . hyorhinis isolates . Covalent modification of lipid-associated proteins was confirmed by identifying 3H-labeled methyl palmitate after acid methanolysis of Triton X-114 phase proteins derived from {3H}palmitate-labeled organisms . However, removal of covalently bound lipid from chloroform-methanol-extracted proteins by alkaline hydroxylamine was selective; complete removal was observed with only a few proteins, possibly including the 120-kDa form of one antigen variant . This suggested potential differences in the nature of covalent linkage among lipid-modified M . hyorhinis surface antigens . Intraspecies antigen variants described here in M . hyorhinis share some characteristics with size-variant antigens reported in phylogenetically related gram-positive eubacteria and may contribute to phenotypic diversification and differences in pathogenicity of mycoplasmas.

Proc Natl Acad Sci U S A, 1988 Dec, 85(23), 9101 - 5
Tandemly repeated tRNA pseudogenes in photobacterium; Giroux S et al.; A region distal to three tRNA genes in Photobacterium phosphoreum, a Gram-negative eubacterium, unexpectedly contains a high number of repeated DNA segments that are closely related to the adjacent tRNAPro gene . The 5' to 3' order of this cluster is tRNAPro-tRNAHis-tRNAPro followed by eight tRNAPro-like structures interspersed by rho-independent terminators . The two tRNAPro genes, which are identical, and the tRNAHis gene have 86% and 87% positional identity, respectively, to their counterparts in the argT operon of Escherichia coli . The facts that these tRNA-like structures are not transcribed, in contrast to the tRNA retropseudogenes of eukaryotes, and that these structures are clustered near their progenitor suggest they are an unusual class of tRNA pseudogenes that arose by tandem duplication.

J Mol Evol, 1988 Dec-1989 Feb, 28(1-2), 98 - 112
The evolutionary relationships among known life forms; Cedergren R et al.; Sequences of small subunit (SSU) and large subunit (LSU) ribosomal RNA genes from archaebacteria, eubacteria, and the nucleus, chloroplasts, and mitochondria of eukaryotes have been compared in order to identify the most conservative positions . Aligned sets of these positions for both SSU and LSU rRNA have been used to generate tree diagrams relating the source organisms/organelles . Branching patterns were evaluated using the statistical bootstrapping technique . The resulting SSU and LSU trees are remarkably congruent and show a high degree of similarity with those based on alternative data sets and/or generated by different techniques . In addition to providing insights into the evolution of prokaryotic and eukaryotic (nuclear) lineages, the analysis reported here provides, for the first time, an extensive phylogeny of the mitochondrial lineage.

J Membr Biol, 1988 Dec, 106(3), 183 - 202
Proline porters effect the utilization of proline as nutrient or osmoprotectant for bacteria; Wood JM; Proline is utilized by all organisms as a protein constituent . It may also serve as a source of carbon, energy and nitrogen for growth or as an osmoprotectant . The molecular characteristics of the proline transport systems which mediate the multiple functions of proline in the Gram negative enteric bacteria, Escherichia coli and Salmonella typhimurium, are now becoming apparent . Recent research on those organisms has provided both protocols for the genetic and biochemical characterization of the enzymes mediating proline transport and molecular probes with which the degree of homology among the proline transport systems of archaebacteria, eubacteria and eukaryotes can be assessed . This review has provided a detailed summary of recent research on proline transport in E . coli and S . typhimurium; the properties of other organisms are cited primarily to illustrate the generality of those observations and to show where homologous proline transport systems might be expected to occur . The characteristics of proline transport in eukaryotic microorganisms have recently been reviewed (Horak, 1986).

J Bacteriol, 1988 Dec, 170(12), 5601 - 6
Highly repetitive tRNA(Pro)-tRNA(His) gene cluster from Photobacterium phosphoreum; Giroux S et al.; A DNA fragment comprising the four tRNA gene sequences of the Escherichia coli argT locus hybridized with two Sau3A-generated DNA fragments from the vibrio Photobacterium phosphoreum (ATCC 11040) . Detailed sequence analysis of the longer fragment shows the following gene organization: 5'-promoter-tRNA(Pro)-tRNAPro-tRNA(Pro)-tRNA(His)-tRNA(Pro)-tRNA(Pro)- tRNA(His)-tRNA(Pro)-five pseudogenes derived from the upstream tRNAPro interspersed by putative Rho-independent terminators . This sequence demonstrates the presence of highly repetitive, tandem tRNA genes in a bacterial genome . Furthermore, a stretch of 304 nucleotides from this cluster was found virtually unchanged in the other (shorter) fragment which was previously sequenced . The two clusters together contain eight tRNA(Pro) pseudogenes and eight fully intact tRNA(Pro) genes, an unusually high number for a single eubacterial isoacceptor tRNA . These results show that the organization of some tRNA operons is highly variable in eubacteria.

FEBS Lett, 1988 Nov 21, 240(1-2), 21 - 8
The primary structures of ribosomal proteins L16, L23 and L33 from the archaebacterium Halobacterium marismortui; Hatakeyama T et al.; The complete amino acid sequences of ribosomal proteins L16, L23 and L33 from the archaebacterium Halobacterium marismortui were determined . The sequences were established by manual sequencing of peptides produced with several proteases as well as by cleavage with dilute HCl . Proteins L16, L23 and L33 consist of 119, 154 and 69 amino acid residues, and their molecular masses are 13,538, 16,812 and 7620 Da, respectively . The comparison of their sequences with those of ribosomal proteins from other organisms revealed that L23 and L33 are related to eubacterial ribosomal proteins from Escherichia coli and Bacillus stearothermophilus, while protein L16 was found to be homologous to a eukaryotic ribosomal protein from yeast . These results provide information about the special phylogenetic position of archaebacteria.

Science, 1988 Nov 18, 242(4881), 1040 - 2
Multiple principal sigma factor homologs in eubacteria: identification of the "rpoD box"; Tanaka K et al.; Genes for the principal sigma factor (rpoD genes) of various eubacteria were identified with a synthetic oligonucleotide probe corresponding to a conserved sequence in rpoD gene products of Escherichia coli and Bacillus subtilis . Multiple rpoD homologs were found in the strains of Micrococcus, Pseudomonas, and Streptomyces, whereas single genes were detected in E . coli, B . subtilis, and Staphylococcus aureus . The four rpoD homologs of Streptomyces coelicolor A3(2) were cloned and sequenced . A homologous portion with 13 amino acids was found in the rpoD genes of S . coelicolor A3(2), E . coli, and B . subtilis and was named the "rpoD box."

FEBS Lett, 1988 Nov 7, 239(2), 313 - 8
Sequence of the gene for ribosomal protein L23 from the archaebacterium Methanococcus vannielii; Kopke AK et al.; The N-terminal sequence of HPLC-purified protein L23 from the Methanococcus vannielii ribosome has been determined by automated liquid-phase Edman degradation . Using the N-terminal amino acid sequence, an oligonucleotide probe complementary to the 5'-end of the gene was synthesized . The 26-mer oligonucleotide, containing two inosines, was used for hybridization with digested M . vannielii chromosomal DNA . The hybridizing band from HpaII-digested genomic DNA was ligated into pUC18 to yield plasmid pMvaZ1 containing the entire gene of protein L23 . The nucleotide sequence complemented the partial amino acid sequence, and the gene codes for a protein of 9824 Da . The amino acid sequence of protein L23 form M . vannielii was compared to that of ribosomal proteins from other archaebacteria as well as from eubacteria and eukaryotes . The number of identical amino acids is highest when the M . vannielii protein is compared to the homologous protein from yeast and lowest vs that from tobacco chloroplasts . Interestingly, the secondary structures of the proteins as predicted by computer programs are more conserved than the primary structures.

Mutat Res, 1988 Nov, 206(3), 335 - 42
Conversion of IQ, a dietary pyrolysis carcinogen to a direct-acting mutagen by normal intestinal bacteria of humans; Carman RJ et al.; The dietary carcinogen, 2-amino-3-methyl-3H-imidazo{4,5-f}quinoline (IQ) is mutagenic in the Salmonella/microsomal mutagenicity assay when activated by microsomal enzymes . IQ is found in many cooked foods, notably fried beef and pork . In laboratory rodents IQ is carcinogenic . We showed that mixed and pure cultures of human intestinal anaerobes, notably Eubacterium spp., metabolized IQ to 2-amino-3,6-dihydro-3-methyl-7H-imidazo{4,5-f}quinoline-7-one (HOIQ) . Unlike IQ, both the synthetic and bacterially produced HOIQ were direct-acting mutagens, i.e . active without microsomal activation . This new direct-acting mutagen, from the bacterial metabolism of a dietary pyrolysis carcinogen, raises new concerns about the possible role of this class of genotoxins in the etiology of human cancer.

Mutat Res, 1988 Nov, 194(3), 219 - 26
Polyclonal antibodies against O6-methylguanine-DNA methyltransferase in adapted bacteria; Ceccoli J et al.; The similarity of the adaptive response and the methyltransferase component in bacterial strains from different phylogenic groups was investigated . An adaptive response with induction of transferase activity was found for the first time in the soil bacteria P . aeruginosa and X . maltophilia . Polyclonal antibodies against the E . coli ada protein were used to investigate the structural similarity of the transferases from several bacterial strains with adaptive responses and inducible transferase activity . These antibodies cross-reacted with transferase from M . luteus and P . aeruginosa but not with proteins from other related bacteria, and not with human transferase . The phylogenic relationships of bacteria with adaptive responses suggest that the response likely was present in the common ancestor of eubacteria . The restricted antibody cross-reactivity may reflect the dual role of the E . coli ada protein not only in DNA repair but in positive gene regulation.

Biol Chem Hoppe Seyler, 1988 Nov, 369(11), 1259 - 66
Archaebacterial ATPase: studies on subunit composition and quaternary structure of the F1-analogous ATPase from Sulfolobus acidocaldarius; Lubben M et al.; A modified procedure for the purification of soluble ATPase from the thermoacidophilic archaebacterium Sulfolobus acidocaldarius is described . In addition to (alpha) 65 and (beta) 51 kDa polypeptides, further subunits gamma * (20 kDa) and delta * (12 kDa) are demonstrated to be components of the enzyme, exhibiting a total molecular mass of 380 kDa . Molecular electron microscopic images of the native enzyme indicate a quaternary structure probably formed by the gamma *, delta *-complex as a central mass surrounded by a pseudohexagon of the peripherally arranged larger alpha and beta subunits . As can be derived from both molecular mass and electron microscopy data, the archaebacterial Sulfolobus-ATPase emerges to exist as an alpha 3 beta 3-quaternary structure with respect to the larger subunits . This is normally found in typical F1-ATPases of eubacterial and eukaryotic organisms . Therefore it is postulated that F1- and F0F1-ATPases, respectively, can occur ubiquitously in all urkingdoms of organisms as functional units of energy-transducing membranes.

J Vet Intern Med, 1988 Oct-Dec, 2(4), 171 - 6
Central nervous system infection associated with anaerobic bacteria in two dogs and two cats; Dow SW et al.; Central nervous system (CNS) infection caused by anaerobic bacteria (including Bacteroides, Fusobacterium, Peptostreptococcus, and Eubacterium) was diagnosed in two dogs and two cats . In one dog there was extensive meningomyeloencephalitis, presumably the result of hematogenous spread of bacteria from lung abscesses and bacterial endocarditis . Subdural empyema of unknown origin was found in a second dog and two cats . Clinical signs in all four animals included mental depression and focal neurologic deficits, without fever.

J Bacteriol, 1988 Oct, 170(10), 4555 - 61
Evidence for a multigene family involved in bile acid 7-dehydroxylation in Eubacterium sp . strain VPI 12708; White WB et al.; Eubacterium sp . strain VPI 12708 is a human intestinal isolate which has an inducible bile acid 7-dehydroxylation activity . At least two cholic acid-induced polypeptides, with molecular masses of 27,000 and 45,000 daltons, respectively, coelute with bile acid 7-dehydroxylation activity . The 45,000-dalton polypeptide appears to be encoded by a cholic acid-induced mRNA species of greater than 6 kilobases, which suggests that the gene coding for this polypeptide is part of a larger operon . A gene has been cloned which flanks the gene encoding the 45,000-dalton polypeptide, in the upstream (5') direction . This gene appears to encode a second 27,000-dalton polypeptide . The gene bears striking homology at both the nucleotide (80%) and deduced amino acid sequence (89%) levels with the gene which encodes the 27,000-dalton polypeptide that has been shown previously to be involved in the bile acid 7-dehydroxylation reaction sequence . The implications of this homology and the possible function(s) of the two homologous genes in bile acid 7-dehydroxylation are discussed . Evidence is presented which suggests that the two homologous genes involved in bile acid 7-dehydroxylation may be part of a larger multigene family in Eubacterium sp . strain VPI 12708.

J Bacteriol, 1988 Oct, 170(10), 4608 - 12
Analysis and characterization of the folates in the nonmethanogenic archaebacteria; White RH; A detailed analysis of the folate coenzymes in the nonmethanogenic archaebacteria has been performed . By using the Lactobacillus casei microbiological assay for folates, the levels of folates in Sulfolobus solfataricus and Sulfolobus acidocaldarius were found to be 3.7 and 8.3 ng/g (dry weight) of cells, respectively, compared with 88,000 and 28,000 ng/g (dry weight) of cells in Halobacterium halobium and Halobacterium strain GN-1, respectively . The levels of folates found in the Sulfolobus spp . were approximately 100 times less than those found in the typical eubacterium, whereas the levels in the halobacteria were approximately 10 times higher . The folate in Sulfolobus solfataricus was shown to consist of only 5-formyltetrahydropteroylglutamate, and the folate in Halobacterium strain GN-1 was shown to consist of only pteroyldiglutamate . The low folate levels in the Sulfolobus spp . are the same as those found in the methanogenic bacteria, suggesting that another C1 carrier may function in these cells.

J Gen Microbiol, 1988 Oct, 134 ( Pt 10), 2815 - 21
Beta-subunit of ATP-synthase: a useful marker for studying the phylogenetic relationship of eubacteria; Amann R et al.; The genes encoding the beta-subunits of ATP-synthases (ATPases) from Bacteroides fragilis DSM 2151, Cytophaga lytica DSM 2039 and 'Taxeobacter ocellatus' were cloned . The nucleotide sequences were determined completely for the genes of the first two organisms and to a major part for that of 'T . ocellatus' . The predicted amino acid sequences were compared with previously published amino acid sequences of beta-subunits . Two characteristic insertions were found in genes from organisms belonging to the so-called bacteroides-cytophaga-flavo-bacterium group . The remaining structure shows a high degree of sequence similarity within this group . These data support the conclusions drawn from comparative 16S rRNA sequence analyses that organisms in this phenotypically heterogeneous group are phylogenetically related . A phylogenetic tree was constructed based on a distance matrix of optimally aligned amino acid sequences of beta-subunits of ATPases of various eubacteria, chloroplasts and mitochondria . It is in good agreement with a tree derived from 16S rRNA sequence analyses.

Biofactors, 1988 Oct, 1(3), 237 - 40
Methanogen factor 390 formation: species distribution, reversibility and effects of non-oxidative cellular stresses; Gloss LM et al.; Factor 390 (F390), an adenylylated or guanylylated derivative of the methanogen coenzyme factor 420 (F420), was previously detected in Methanobacterium thermoautotrophicum cells exposed to air . Of six other methanogenic species that have now been tested, only Methanobacterium formicicum was found to produce F390 upon oxygen exposure . Aerobic conditions led to an immediate cessation of methanogenesis, whereas only 51% of cellular F420 was slowly converted to F390 over 4 h in Mb.formicicum at 37 degrees C . F390 formation is reversible . When oxidized cells were re-introduced into anoxic medium, F390 reverted to F420 prior to recovery of methanogenesis . Anaerobic cultures of Mb.formicicum were subjected to alternative stresses such as exposure to heavy metals, methanogenesis inhibitors and eubacterial alarmone-producing chemicals; however, only oxygen was found to induce F390 formation.

J Bacteriol, 1988 Oct, 170(10), 4548 - 54
Nucleotide sequence analysis of a gene cloned from Leptospira biflexa serovar patoc which complements an argE defect in Escherichia coli; Zuerner RL et al.; The genus Leptospira, as a member of the order Spirochaetales, forms one of the most ancient evolutionary branches of the eubacteria . These spirochetes are morphologically and physiologically different from most eubacteria, and little is known about Leptospira genetics . In this communication, we report the first nucleotide sequence of a Leptospira gene . A gene which complements an argE mutation in Escherichia coli was isolated from a plasmid-based genomic library composed of Leptospira biflexa serovar patoc DNA . The functional region for the complementing activity was localized by transposon mutagenesis and restriction enzyme mapping and by subcloning . Nucleotide sequence analysis indicated a single open reading frame within the region containing argE complementing activity . The size of the predicted protein, 31,071 daltons, was in excellent agreement with data obtained from coupled transcription-translation reactions primed with cloned L . biflexa DNA . One surprising result was that the predicted amino acid sequence of this protein closely resembles portions of the beta' subunits of RNA polymerases from bacteria and chloroplasts.

J Mol Biol, 1988 Sep 5, 203(1), 15 - 27
Mutations in rpoD that increase expression of genes in the mal regulon of Escherichia coli K-12; Hu JC et al.; The sigma subunits of eubacterial RNA polymerases determine the site selectivity of initiation of transcription at promoters . Mutations in rpoD, the gene that encodes sigma 70, the major sigma factor in Escherichia coli, should be useful in determining the molecular details of the process of transcription initiation . However, such mutations are likely to be deleterious or lethal, since sigma70 is an essential gene product . We designed a system for the rapid isolation and fine structure mapping of mutations in rpoD, which allows selection of mutations that would otherwise be deleterious to the cell . We used this system to isolate a new class of mutations in rpoD, mutations that relieve the requirement for CAP-cAMP for initiation at promoters in the mal regulon . These mutations, which we designate rpoD(Mal) mutations, occur in two clusters in the rpoD gene within regions previously suggested by amino acid sequence comparisons to be important for sigma structure or function . We cannot distinguish whether the rpoD(Mal) mutations affect mal expression by altering interaction between RNA polymerase and mal promoters or between RNA polymerase and the accessory transcription factor MalT . However, the effects of the mutations on activator-independent transcription from the lac promoter (4 rpoD(Mal) mutations decrease CAP-independent expression of the lac promoter in vivo) suggest that the regions of sigma identified by our mutations may be directly involved in promoter recognition.

J Bacteriol, 1988 Sep, 170(9), 4280 - 5
Methyl-accepting protein associated with bacterial sensory rhodopsin I; Spudich EN et al.; In vivo radiolabeling of Halobacterium halobium phototaxis mutants and revertants with L-{methyl-3H} methionine implicated seven methyl-accepting protein bands with apparent molecular masses from 65 to 150 kilodaltons (kDa) in adaptation of the organism to chemo and photo stimuli, and one of these (94 kDa) was specifically implicated in phototaxis . The lability of the radiolabeled bands to mild base treatment indicated that the methyl linkages are carboxylmethylesters, as is the case in the eubacterial chemotaxis receptor-transducers . The 94-kDa protein was present in increased amounts in an overproducer of the apoprotein of sensory rhodopsin I, one of two retinal-containing phototaxis receptors in H . halobium . It was absent in a strain that contained sensory rhodopsin II and that lacked sensory rhodopsin I and was also absent in a mutant that lacked both photoreceptors . Based on the role of methyl-accepting proteins in chemotaxis in other bacteria, we suggest that the 94-kDa protein is the signal transducer for sensory rhodopsin I . By {3H}retinal labeling studies, we previously identified a 25-kDa retinal-binding polypeptide that was derived from photochemically reactive sensory rhodopsin I . When H . halobium membranes containing sensory rhodopsin I were treated by a procedure that stably reduced {3H}retinal onto the 25-kDa apoprotein, a 94-kDa protein was also found to be radiolabeled . Protease digestion confirmed that the 94-kDa retinal-labeled protein was the same as the methyl-accepting protein that was suggested above to be the signal transducer for sensory rhodopsin I . Possible models are that the 25- and 94-kDa proteins are tightly interacting components of the photosensory signaling machinery or that both are forms of sensory rhodopsin I.

J Bacteriol, 1988 Sep, 170(9), 4136 - 40
RNA polymerase subunit homology among cyanobacteria, other eubacteria and archaebacteria; Schneider GJ et al.; RNA polymerase purified from vegetative cells of the cyanobacterium Anabaena sp . strain PCC 7120 contains a dissociable sigma factor and a core of five subunits: the beta', beta, and two alpha subunits characteristic of all eubacteria and an additional 66,000-molecular-weight polypeptide called gamma . Fifteen of fifteen strains of unicellular and filamentous cyanobacteria tested contained a serologically related gamma protein . Antiserum to gamma reacted with Escherichia coli beta' and the A subunit of RNA polymerase of the archaebacterium Sulfolobus acidocaldarius . Thus the evolution of the RNA polymerase beta' subunit has followed different paths in three groups of procaryotes: cyanobacteria, other eubacteria, and archaebacteria.

Mol Microbiol, 1988 Sep, 2(5), 569 - 79
Transcriptional analysis of the 16S rRNA gene of the rrnD gene set of Streptomyces coelicolor A3(2); Baylis HA et al.; The nucleotide sequence of 2.5 kb of the Streptomyces coelicolor A3(2) rRNA gene set rrnD, extending from upstream of the 16S rRNA gene to the putative 5' end of the 23S rRNA gene, has been determined (Baylis and Bibb, 1987; this paper) . In addition to locating the 5' end of the 16S rRNA gene, nuclease S1 mapping identified seven RNA 5' end-points upstream of the 16S rRNA gene; four of these were coincident with transcriptional initiation points for S . coelicolor A3(2) RNA polymerase in vitro and were consequently regarded as in vivo transcription start points for promoters p1 to p4 . One end-point identified by nuclease S1 mapping localized a putative processing site analogous to those found upstream of 16S rRNA genes in other eubacteria . Sequence motifs similar to those discovered in low G+C Gram-positive bacteria were found associated with two of the promoters and the processing site . A probable protein coding region was observed upstream of the promoter region.

Nucleic Acids Res, 1988 Aug 25, 16(16), 7817 - 26
Gene for the diphtheria toxin-susceptible elongation factor 2 from Methanococcus vannielii; Lechner K et al.; Protein synthesis elongation factor 2 (EF-2) from all archaebacteria so far analysed, is susceptible to inactivation by diphtheria toxin, a property which it shares with EF-2 from the eukaryotic 8OS translation system . To resolve the structural basis of diphtheria toxin susceptibility, the structural gene for the EF-2 from an archaebacterium, Methanococcus vannielii, was cloned and its nucleotide sequence determined . It was found that (i) this gene is closely linked to that coding for elongation factor 1 alpha-(EF-1 alpha), (ii) the size of the gene product, as derived from the nucleotide sequence, lies between those for EF-2 from eukaryotes and eubacteria, (iii) it displays a higher sequence similarity to eukaryotic EF-2 than to eubacterial homologues, and (iv) the histidine residue which is modified to diphthamide and then ADP-ribosylated by diphtheria toxin is present in a sequence context similar to that of eukaryotic EF-2 but it is not conserved in eubacterial EF-G . The EF-2 gene from Methanococcus is expressed in transformed Saccharomyces cerevisiae but is not ADP-ribosylated by diphtheria toxin . This indicates that the Saccharomyces enzyme system is unable to post-translationally convert the respective histidine residue from the Methanococcus EF-2 into diphthamide.

Nucleic Acids Res, 1988 Aug 25, 16(16), 8113 - 28
Relatedness of archaebacterial RNA polymerase core subunits to their eubacterial and eukaryotic equivalents; Berghofer B et al.; The sequence of the genes encoding the four largest subunits of the RNA polymerase of the archaebacterium Methanobacterium thermoautotrophicum was determined and putative translation signals were identified . The genes are more strongly homologous to eukaryotic than to eubacterial RNA polymerase genes . Analysis of the polypeptide sequences revealed colinearity of two pairs of adjacent archaebacterial genes encoding the B" and B' or A and C genes, respectively, with two eubacterial and two eukaryotic genes each encoding the two largest RNA polymerase subunits . This difference in sequence organization is discussed in terms of gene fusion in the course of evolution . The degree of conservation is much higher between the archaebacterial and the eukaryotic polypeptides than between the archaebacterial and the eubacterial enzyme . Putative functional domains were identified in two of the subunits of the archaebacterial enzyme.

Antimicrob Agents Chemother, 1988 Aug, 32(8), 1271 - 3
Tetracycline resistance and TetM in oral anaerobic bacteria and Neisseria perflava-N . sicca; Roberts MC et al.; Tetracycline-resistant organisms isolated from six patients with periodontal disease included Bacteroides spp., Eubacterium spp., Fusobacterium nucleatum, Neisseria perflava-N . sicca, Peptostreptococcus anaerobius, Veillonella parvula, and facultative streptococci . All but the Bacteroides spp . and Eubacterium spp . hybridized with the TetM determinant . An additional 417 bacterial strains were screened, and 4% of both the oral streptococci and the Fusobacterium spp . hybridized with the TetM probe.

J Bacteriol, 1988 Aug, 170(8), 3584 - 92
Evolutionary relationships among cyanobacteria and green chloroplasts; Giovannoni SJ et al.; The 16S rRNAs from 29 cyanobacteria and the cyanelle of the phytoflagellate Cyanophora paradoxa were partially sequenced by a dideoxynucleotide-terminated, primer extension method . A least-squares distance matrix analysis was used to infer phylogenetic trees that include green chloroplasts (those of euglenoids, green algae, and higher plants) . The results indicate that many diverse forms of cyanobacteria diverged within a short span of evolutionary distance . Evolutionary depth within the surveyed cyanobacteria is substantially less than that separating the major eubacterial taxa, as though cyanobacterial diversification occurred significantly after the appearance of the major eubacterial groups . Three of the five taxonomic sections defined by Rippka et al . (R . Rippka, J . Deruelles, J . B . Waterbury, M . Herdman, and R . Y . Stanier, J . Gen . Microbiol . 111:1-61, 1979) (sections II {pleurocapsalean}, IV {heterocystous, filamentous, nonbranching}, and V {heterocystous, filamentous, branching}) are phylogenetically coherent . However, the other two sections (I {unicellular} and III {nonheterocystous, filamentous}) are intermixed and hence are not natural groupings . Our results not only support the conclusion of previous workers that the cyanobacteria and green chloroplasts form a coherent phylogenetic group but also suggest that the chloroplast lineage, which includes the cyanelle of C . paradoxa, is not just a sister group to the free-living forms but rather is contained within the cyanobacterial radiation.

FEBS Lett, 1988 Aug 1, 235(1-2), 241 - 6
AsnI: a novel class II restriction endonuclease from Arthrobacter sp., strain N-CM, recognizing 5'-AT/TAAT-3'; Rexer BU et al.; A new class II restriction endonuclease, AsnI, with a novel sequence specificity was isolated from the Gram-positive eubacterium Arthrobacter species, strain N-CM . AsnI recognizes the unambiguously defined palindromic hexanucleotide (Formula: see text) consisting of A- and T-residues . The novel enzyme in the presence of Mg2+ cleaves specifically both strands as indicated by the arrows . The staggered cuts generate 5'-protruding ends with single-stranded 5'-TA-3' dinucleotide extensions . The novel enzyme may be a useful tool for cloning experiments by complementation of the few enzymes such as PstI and PvuI cutting only once in the Ampr-gene of plasmids pBR322 and pBR328.

Proc Natl Acad Sci U S A, 1988 Aug, 85(15), 5521 - 4
cDNA sequence encoding the 16-kDa proteolipid of chromaffin granules implies gene duplication in the evolution of H+-ATPases; Mandel M et al.; Vacuolar H+-ATPases function in generating protonmotive force across the membranes of organelles connected with the vacuolar system of eukaryotic cells . This family of H+-ATPases is distinct from the two other families of H+-ATPases, the plasma membrane-type and the eubacterial-type . One of the subunits of the vacuolar H+-ATPase binds N,N'-dicyclohexylcarbodiimide (DCCD) and has been implicated in the proton-conducting activity of these enzymes . We have cloned and sequenced the gene encoding the DCCD-binding protein (proteolipid) of the H+-ATPase of bovine chromaffin granules . The gene encodes a highly hydrophobic protein of 15,849 Da . Hydropathy plots revealed four transmembrane segments, one of which contains a glutamic residue that is the likely candidate for the DCCD binding site . Sequence homology with the vacuolar proteolipid and with the proteolipids of eubacterial-type H+-ATPases was detected . The proteolipids from Escherichia coli, spinach chloroplasts, and yeast mitochondria matched better to the NH2-terminal part of the vacuolar protein . The proteolipids of bovine mitochondria and Neurospora mitochondria matched better to the COOH-terminal end of the vacuolar proteolipid . These findings suggest that the proteolipids of the vacuolar H+-ATPases were evolved in parallel with the eubacterial proteolipid, from a common ancestral gene that underwent gene duplication.

J Mol Biol, 1988 Jul 5, 202(1), 45 - 58
Mapping and sequencing of mutations in the Escherichia coli rpoB gene that lead to rifampicin resistance; Jin DJ et al.; Rifampicin is an antibiotic that inhibits the function of RNA polymerase in eubacteria . Mutations affecting the beta subunit of RNA polymerase can confer resistance to rifampicin . A large number of rifampicin-resistant (hereafter called Rifr) mutants have been isolated in Escherichia coli to probe the involvement of RNA polymerase in a variety of physiological processes . We have undertaken a comprehensive analysis of Rifr mutations to identify their structural and functional effects on RNA polymerase . Forty-two Rifr isolates with a variety of phenotypes were mapped to defined intervals within the rpoB gene using a set of deletions of the rpoB gene . The mutations were sequenced . Seventeen mutational alterations affecting 14 amino acid residues were identified . These alleles are located in three distinct clusters in the center of the rpoB gene . We discuss the implications of our results with regards to the structure of the rifampicin binding site.

Eur J Biochem, 1988 Jul 1, 174(4), 637 - 40
Biosynthesis of vitamin B-12 in anaerobic bacteria . Experiments with Eubacterium limosum and D-erythrose 14C-labeled in different positions; Vogt JR et al.; In anaerobic microorganisms the origin of C atoms 2 and 4-7 of the 5,6-dimethylbenzimidazole moiety of vitamin B-12 is still unknown . In order to tackle this problem we added several 14C-labeled putative precursors to Eubacterium limosum fermentations . The degradation of the isolated vitamin B-12 revealed that only D-erythrose, 14C-labeled in different positions, was efficiently incorporated into the 5,6-dimethylbenzimidazole part . The 5,6-dimethylbenzimidazole obtained from an experiment with D-{U-14C}erythrose was further degraded . It was found that C-2 was unlabeled, whereas half of the label was located in C-5 plus C-6, and the other half in C-4 plus C-7 . These results demonstrate that in E . limosum D-erythrose is a precursor of C-atoms 4, 5, 6 and 7 of the 5,6-dimethylbenzimidazole part of vitamin B-12.

Proc Natl Acad Sci U S A, 1988 Jul, 85(14), 4976 - 80
Protein kinase and phosphoprotein phosphatase activities of nitrogen regulatory proteins NTRB and NTRC of enteric bacteria: roles of the conserved amino-terminal domain of NTRC; Keener J et al.; The NTRC protein (ntrC product) of enteric bacteria activates transcription of nitrogen-regulated genes by a holoenzyme form of RNA polymerase that contains the ntrA product (sigma 54) as sigma factor . Although unmodified NTRC will bind to DNA, it must be phosphorylated to activate transcription . Both phosphorylation and dephosphorylation of NTRC occur in the presence of the NTRB protein (ntrB product) . We here demonstrate rigorously that it is the NTRB protein that is a protein kinase by showing that NTRB can phosphorylate itself, whereas NTRC cannot . Phosphorylated NTRC (NTRC-P) is capable of autodephosphorylation with a first-order rate constant of 0.14-0.19 min-1 (t 1/2 of 5.0-3.6 min) at 37 degrees C . In addition, there is regulated dephosphorylation of NTRC-P . By contrast to the autophosphatase activity, regulated dephosphorylation requires three components in addition to NTRC-P: the PII regulatory protein, NTRB, and ATP . NTRC is phosphorylated within its amino-terminal domain, which is conserved in one partner of a number of two-component regulatory systems in a wide variety of eubacteria . A purified amino-terminal fragment of NTRC (approximately equal to 12.5 kDa) is sufficient for recognition by NTRB and is autodephosphorylated at the same rate as the native protein.

J Bacteriol, 1988 Jun, 170(6), 2472 - 9
Characterization of the pyrogallol-phloroglucinol isomerase of Eubacterium oxidoreducens; Krumholz LR et al.; Cell extracts of Eubacterium oxidoreducens, in the presence of dimethyl sulfoxide, catalyzed the conversion of pyrogallol to phloroglucinol with methyl sulfide as a product . The isomerization reaction also proceeded when 1,2,3,5-benzenetetrol was present rather than dimethyl sulfoxide . An assay to quantitate this activity was developed . The assay followed the disappearance of 1,2,4-benzenetriol as determined colorimetrically after incubation with sodium molybdate at neutral pH . The products of this reaction were resorcinol and 2,6-dihydroxyquinone . The enzyme(s) catalyzing this reaction was purified fivefold from cells grown on gallate plus H2 . The purification procedure involved treatment with 40% acetone, precipitation with ammonium sulfate, DEAE-cellulose chromatography, concentration by ultrafiltration (molecular weight cutoff, greater than 100,000), and hydroxylapatite chromatography . This preparation had a specific activity of 14.7 mumol/min per mg of protein and a pH optimum of about 7.3 . It was strongly inhibited by p-chloromercuribenzoate . The mechanism of the reaction involved oxidation of the pyrogallol followed by introduction of water . The benzenetetrol intermediate was then reduced and dehydrated to phloroglucinol.

J Gen Microbiol, 1988 Jun, 134 ( Pt 6), 1561 - 4
Outer membrane protein pattern of Eubacterium plautii; Baardsen R et al.; The outer membrane SDS-PAGE pattern of Eubacterium plautii was characterized by a large number of surface exposed low- and high-molecular-mass proteins . Silver stainable carbohydrate was not present . The pattern was clearly distinct from those of outer membrane preparations of Eubacterium saburreum and Fusobacterium nucleatum . The results are compatible with a Gram-positive cell wall structure in E . plautii.

Arch Biol Med Exp (Santiago), 1988 Jun, 21(1), 219 - 29
Methylation of proteins from the translational apparatus: an overview; Toledo H et al.; Several of the translational apparatus proteins are methylated in all kinds of organisms . Although most of the modified proteins play key roles during protein biosynthesis, the biological function of these chemical modifications still remains elusive . Our recent data indicate a highly conserved pattern of ribosomal protein methylation in eubacteria, with methylated proteins being both structurally and functionally homologous in several microorganisms . Chloroplast ribosomes also appear to have a rather eubacterial pattern of ribosomal protein methylation . On the other hand, there is an apparently ubiquitous methylation of some of the translational factors in several organisms . These findings suggest an important, albeit unknown role for the post-synthetic methylation of the translational machinery . The analysis of the sequences of known methylation target sites and the search of similar sites in other proteins of known sequence, allows to predict those ribosomal proteins or translational factors that may be subjected to post-translational modifications with one or more methyl groups . Although a definitive answer with respect to the biological role of these N-methylations is still missing, a direct correlation between the methylation of some proteins and their biological activity is just beginning to emerge.

J Biol Chem, 1988 May 25, 263(15), 7087 - 93
Microsequence analysis of DNA-binding proteins 7a, 7b, and 7e from the archaebacterium Sulfolobus acidocaldarius; Choli T et al.; DNA-binding proteins in eubacteria, such as Escherichia coli NS1 and NS2, are generally small basic molecules . In contrast, the archaebacterium Sulfolobus acidocaldarius contains three groups of DNA-binding proteins which have molecular masses of 7, 8, and 10 kDa . In the first group, five proteins (7a-7e) have been identified, while in the second and third group only two proteins each are present, denoted 8a and 8b and 10a and 10b, respectively . In this paper, we present the primary structures of proteins 7a, 7b, and 7e from the first group . All three proteins contain lysyl residues which are monomethylated to different extents . The modified lysines are found in the NH2-terminal regions of all 7-kDa proteins and in the COOH-terminal part of protein 7e . The sequences of the 7-kDa group are highly similar to each other . All of these macromolecules have been shown to interact specifically with DNA . Protein 7e of the 7-kDa group shows the tightest binding to DNA.

J Biol Chem, 1988 May 15, 263(14), 6538 - 46
Primary structure of the archaebacterial Methanococcus vannielii ribosomal protein L12 . Amino acid sequence determination, oligonucleotide hybridization, and sequencing of the gene; Strobel O et al.; The primary structure of ribosomal protein L12 from Methanococcus vannielii has been determined by direct amino acid sequence analysis with automated liquid phase Edman degradation of the entire protein and manual 4-N,N'-dimethylaminoazobenzene-4'-isothiocyanate/phenylisothiocyanate sequencing of fragments obtained by enzymatic digestion and by partial acid hydrolysis . The knowledge of the amino acid sequences of these various fragments allowed the synthesis of two oligonucleotide probes complementary to the 5'- and the 3'-end of the gene, and they were used for hybridization with digested M . vannielii chromosomal DNA . Both oligonucleotide probes gave similar and clear hybridization signals . The plasmid pMvaX1 containing the entire gene of protein L12 was obtained . The nucleotide sequence complemented the partial amino acid sequence, and it is in full agreement with the protein sequence and the amino acid analysis . Comparison of secondary structural elements and hydrophobicity plots of the M . vannielii protein L12 with the known L12 sequences derived from other archaebacterial and eukaryotic sources show strong homologies among these sequences . They contain an exceptional highly conserved hydrophilic sequence area in the C-terminal part of the proteins . In comparison with eubacterial L12 proteins, the conservation is reduced to single amino acid residues . However, the eubacterial L12 proteins have hydrophilic regions similar to those of L12 from M . vannielii . These regions are predicted to be located at the surface of the proteins, as has been proven to be the case in crystallized Escherichia coli L12 protein . It is possible that the strongly conserved hydrophilic sequence regions form part of the factor-binding domain.

Eur J Biochem, 1988 May 2, 173(3), 473 - 82
Comparative evaluation of gene expression in archaebacteria; Zillig W et al.; Gene organization, gene structure, especially regarding transcription and translation signals, and the structure of essential components of the gene expression machinery of archaebacteria are compared with those of eubacteria and eukaryotes . Many features of the genetic machinery of archaebacteria are shared either with eubacteria or with eukaryotes . For example, the translation signals including ribosome-binding sites are the same as in eubacteria, but the consensus sequence of archaebacterial promoters closely resembles that of the eukaryotic polymerase II promoters . Archaebacterial genes can be organized in transcription units resembling those of eubacteria . But the sequences of several protein components of the genetic machinery have strikingly more homology with those of their eukaryotic than with those of their eubacterial correspondents . The sequences of the large components of DNA-dependent RNA polymerases of archaebacteria closely resemble those of the eukaryotic RNA polymerases II and, somewhat less, III . In a dendrogram calculated from percentage homology data, the eukaryotic RNA polymerase I component A shares a branching point with the eubacterial component . The implications of these findings for the origin and the evolution of the eukaryotic ancestry are discussed.

J Bacteriol, 1988 May, 170(5), 2078 - 82
New function of vitamin B12: cobamide-dependent reduction of epoxyqueuosine to queuosine in tRNAs of Escherichia coli and Salmonella typhimurium; Frey B et al.; Queuosine (Q), 7-{(4,5-cis-dihydroxy-2-cyclopentene-1-yl)-amino)methyl)-7- deazaguanosine, and Q derivatives usually replace guanosine in the anticodon of tRNAs(GUN) of eubacteria and of cytoplasmic and mitochondrial tRNAs of lower and higher eucaryotes except yeasts . Q appears to be synthesized de novo exclusively in eubacteria, and the free-base queuine serves as a nutrient factor for eucaryotes . Recently, a Q derivative, oQ, containing a 2,3-epoxy-4,5-dihydroxycyclopentane ring, has been identified in Escherichia coli tRNA(Tyr) . Here we show that oQ is formed when E . coli or Salmonella typhimurium is grown in glucose-salt medium . The formation of oQ was independent of molecular oxygen, and oQ-tRNAs were converted to Q-tRNAs by adding cobalamin to the growth medium . Under strictly anaerobic conditions, considerable amounts of Q were present in E . coli and S . typhimurium tRNAs when the bacteria were grown in the presence of cobalt ions with glycerol as the carbon source and fumarate as the electron acceptor . Under these conditions, the biosynthesis of cobalamin was induced . The results suggest that oQ is derived from ribose and that oQ is finally reduced to Q by a cobamide-dependent enzyme.

Biochem Cell Biol, 1988 May, 66(5), 325 - 48
Organelle origins and ribosomal RNA; Gray MW; As the detailed molecular biology of organelle genomes has unfolded, there has been a general acceptance of the view that plastids and mitochondria are of endosymbiotic, eubacterial origin . Plastid genes are strikingly similar to their eubacterial (particularly cyanobacterial) counterparts in sequence, organization, and mode of expression, and such features strongly support the hypothesis that the plastid and its genome were derived in evolution from a blue-green alga-like endosymbiont . Mitochondria, on the other hand, are problematic: mitochondrial genes are organized and expressed in remarkably diverse ways in the different major groups of eukaryotes, and in no case are these features particularly characteristic of either bacterial or nuclear genomes . There is, however, clear evidence derived from gene sequence supporting the eubacterial ancestry of mitochondria, and some of the most compelling data have come from analyses of mitochondrial ribosomal RNA (rRNA) . Plant mitochondrial rRNA genes diverge in sequence at a particularly slow rate, and these genes have proven to be especially supportive of the endosymbiont hypothesis, pointing to an origin of mitochondria from within the alpha subdivision of the purple bacteria . Ribosomal RNA sequences provide a basis for the construction of global phylogenetic trees that probe the evolutionary history of organelles, and that address the question of whether mitochondria and plastids are monophyletic or polyphyletic in origin . Such studies raise the possibility that the rRNA genes of plant mitochondria originated separately from the mitochondrial rRNA genes of other eukaryotes.

J Bacteriol, 1988 May, 170(5), 2070 - 7
Nucleotide sequence and regulation of a gene involved in bile acid 7-dehydroxylation by Eubacterium sp . strain VPI 12708; Coleman JP et al.; Eubacterium sp . strain VPI 12708 is an anaerobic intestinal bacterium that has inducible bile acid 7-dehydroxylation activity . At least four new polypeptides were synthesized after addition of primary bile acids to the growth medium . One of these, of molecular weight 27,000 (P-27), was shown to be involved in the 7-dehydroxylation reaction sequence . The gene coding for P-27 was cloned, and the entire DNA sequence for the protein-coding region was determined . In addition, sequence information was obtained for 294 bases upstream from the translational start codon and 329 bases downstream from the stop codon . Induction studies with a synthetic oligonucleotide probe (16-mer) revealed the presence of a cholic acid-inducible mRNA species approximately 900 bases long . A 5' terminus of this mRNA was detected by primer extension analysis, and the location of the 3' terminus of the mRNA was estimated by using S1 nuclease mapping . The 3' terminus of the mRNA contained a large element with dyad symmetry of unknown function . The open reading frame contained 249 codons, and the corresponding polypeptide had a calculated molecular weight of 26,745 . The amino acid sequence of P-27 showed significant homology to several previously described alcohol-polyol dehydrogenases ("nonzinc" dehydrogenases), especially in the region believed to contain a pyridine nucleotide-binding domain . The implications of this homology and the possible function of P-27 in bile acid 7-dehydroxylation are discussed.

J Gen Microbiol, 1988 May, 134 ( Pt 5), 1197 - 204
DNA probes with different specificities from a cloned 23S rRNA gene of Micrococcus luteus; Regensburger A et al.; A 7500 bp PstI restriction fragment of chromosomal DNA from Micrococcus luteus containing a 23S rRNA gene was cloned in vector pHE3 in E . coli RR 28 (the recombinant plasmid was designated pAR1) . A recombinant phage (pAR5) hybridizing to all eubacteria tested was constructed by shotgun subcloning of the PstI fragment in phage M13mp8 . Further subcloning of the fragments of the 23S rRNA gene in the vectors pTZ18R and pTZ19R using selected restriction sites of the gene enabled us to select cloned fragments of the 23S rRNA gene representing different specificities . Probes specific for Micrococcus luteus-Micrococcus lylae (pAR28), for the Arthrobacter-Micrococcus group (pAR27), for eubacteria (pAR5), and for the detection of eu- and archaebacteria (the so-called universal probe pAR17) were constructed . The specificity of each probe was analysed by dot hybridization to the chromosomal DNAs of representatives of most of the main phyla of eu- and archaebacteria.

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