J Protozool, 1987 Nov, 34(4), 435 - 8 Effect of biogenic amines on phagocytosis in Tetrahymena thermophila; Quinones-Maldonado V et al.; Stimulation of phagocytosis by serotonin and catecholamines in Tetrahymena grown in proteose-peptone medium proved to be concentration dependent, the optimal concentrations being approximately 0.1 to 1.0 microM . The serotonergic antagonists, spiperone, and metergoline, also stimulated the process, whereas the beta- and alpha-adrenergic antagonists, propranolol, alprenolol, and ergocryptine, had no effect or inhibited phagocytosis . A wide variety of derivatives of the biogenic amines had no effect on phagocytosis, demonstrating the specificity of recognition mechanism for neurohormones in Tetrahymena . Such hormones act by at least two independent mechanisms, one for adrenergic agonists, another for dopamine . Presumably, recognition mechanisms for hormones in protozoa resemble in some respects those in multicellular organisms, therefore bespeaking a common origin. J Exp Biol, 1987 Nov, 133, 169 - 82 Thermal dependence of passive electrical properties of lizard muscle fibres; Adams BA; 1 . The thermal dependence of passive electrical properties was determined for twitch fibres from the white region of the iliofibularis (IF) muscle of Anolis cristatellus (15-35 degrees C) and Sceloporus occidentalis (15-40 degrees C), and for twitch fibres from the white (15-45 degrees C) and red (15-40 degrees C) regions of the IF of Dipsosaurus dorsalis . These species differ in thermal ecology, with Anolis being the least thermophilic and Dipsosaurus the most thermophilic . 2 . Iliofibularis fibres from the three species reacted similarly to changing temperature . As temperature was increased, input resistance (Rin) decreased (average R10 = 0.7), length constant (L) decreased (average R10 = 0.9), time constant (tau) decreased (average R10 = 0.8), sarcoplasmic resistivity (Rs) decreased (average R10 = 0.8) and apparent membrane resistance (Rm) decreased (average R10 = 0.7) . In contrast, apparent membrane capacitance (Cm) increased with increasing temperature (average R10 = 1.3) . 3 . Rin, L, tau and apparent Rm were lowest in fibres from Anolis (the least thermophilic species) and highest in fibres from Dipsosaurus (the most thermophilic species) . Anolis had the largest and Dipsosaurus the smallest diameter fibres (126 and 57 micron, respectively) . Apparent Cm was highest in fibres from Sceloporus, which had fibres of intermediate diameter (101 micron) . Rs did not differ significantly among species . 4 . The effect of temperature on the passive electrical properties of these lizard fibres was similar to that reported for muscle fibres from other ectothermic animals (crustaceans, insects, fish and amphibians) but qualitatively different from that reported for some mammalian (cat tenuissimus, goat intercostal) fibres . The changes that occur in the passive electrical properties render the fibres less excitable as temperature increases. J Protozool, 1987 Nov, 34(4), 416 - 7 Self-splicing RNA and an RNA enzyme in Tetrahymena; Zaug AJ et al.; The RNA molecules transcribed from many eukaryotic genes are interrupted by intervening sequences, which are removed by a process called RNA splicing . One structurally related group of intervening sequences, the group I intervening sequences, are found in a variety of microorganisms . Some of these, including the group I intervening sequence from the ribosomal RNA precursor of Tetrahymena thermophila, have been shown to mediate their own splicing in an RNA-catalyzed reaction . Following its excision from the ribosomal RNA precursor, the Tetrahymena intervening sequence acts as an enzyme, cutting and rejoining RNA substrates. Biochem Int, 1987 Nov, 15(5), 953 - 60 Three-dimensional crystals of ribosomes and their subunits from eu- and archaebacteria; Glotz C et al.; Ordered three-dimensional crystals of 70S ribosomes as well as of 30S and 50S ribosomal subunits from various bacteria (E . coli, Bacillus stearothermophilus, Thermus thermophilus and Halobacterium marismortui) have been grown by vapour diffusion in hanging drops using mono- and polyalcohols . A new compact crystal form of 50S subunits has been obtained, and it is suitable for crystallographic studies at medium resolution . In addition, from one crystal form large crystals could be grown in X-ray capillaries . In all cases the crystals were obtained from functionally active ribosomal particles, and the particles from dissolved crystals retained their integrity and biological activity. Genetics, 1987 Nov, 117(3), 451 - 66 Genomic organization and developmental fate of adjacent repeated sequences in a foldback DNA clone of Tetrahymena thermophila; Tschunko AH et al.; DNA sequence elimination and rearrangement occurs during the development of somatic cell lineages of eukaryotes and was first discovered over a century ago . However, the significance and mechanism of chromatin elimination are not understood . DNA elimination also occurs during the development of the somatic macronucleus from the germinal micronucleus in unicellular ciliated protozoa such as Tetrahymena thermophila . In this study foldback DNA from the micronucleus was used as a probe to isolate ten clones . All of those tested (4/4) contained sequences that were repetitive in the micronucleus and rearranged in the macronucleus . The presence of inverted repeated sequences was clearly demonstrated in one of them by electron microscopy . DNA sequence analysis showed that the left portion of this clone contains three tandem, directly repeated copies of a 340-bp sequence, a 120-bp portion of which appears in inverted orientation at a 1.6-kb distance . This clone, pTtFB1, was subjected to a detailed analysis of its developmental fate . Subregions were subcloned and used as probes against Southern blots of micronuclear and macronuclear DNA . We found that all subregions defined repeated sequence families in the micronuclear genome . A minimum of four different families was defined, two of which are retained in the macronucleus and two of which are completely eliminated . The inverted repeat family is retained with little rearrangement . Two of the families, defined by subregions that do not contain parts of the inverted repeat, one in the "loop" and one in the "right flanking region," are totally eliminated during macronuclear development--and contain open reading frames . A fourth family occurs in the "loop" region and is rearranged extensively during development . The two gene families that are eliminated are stable in the micronuclear genome but are not clustered together as evidenced by experiments in which DNAs from nullisomic strains are used to map family members to specific micronuclear chromosomes . The inverted repeat family is also stable in the micronuclear genome and is dispersed among several chromosomes . The significance of retained inverted repeats to the process of elimination is discussed. J Biol Chem, 1987 Oct 25, 262(30), 14672 - 82 Site-directed mutagenesis of core sequence elements 9R', 9L, 9R, and 2 in self-splicing Tetrahymena pre-rRNA; Williamson CL et al.; The intron within the Tetrahymena thermophila nuclear large rRNA precursor is the best studied example of group I self-splicing introns . In this paper, we examine the structural and functional roles of four internal sequence elements which are characteristic of group I introns in the RNA-catalyzed processing reactions . Oligonucleotide-directed mutagenesis was used to generate mutations in sequence elements 9R', 9L, 9R and 2 of the Tetrahymena intervening sequence . Self-splicing activities of variant precursor RNAs were characterized by in vitro splicing following transcription with T7 or SP6 RNA polymerase . First, we confirm the proposed base pairing of sequence elements 9R and 9R' by construction and analysis of compensatory mutations . Mutations in elements 9R (G272A C274G) and 9R' (G100C C102U) each disrupt the pairing and eliminate self-splicing activity . A compensatory 9R/9R' mutation (G100C C102U G272A C274G) restores pairing and normal splicing activity . We conclude that 9R X 9R' pairing is a requirement for self-splicing . Second, we show that self-splicing activity is very sensitive to both nucleotide sequence and RNA secondary structure in the pairing segments of elements 9L and 2 . Mutations within these regions at positions 266, 268, 307, and 309 can increase as well as decrease activity relative to wild type . Third, a mutation in the highly conserved nonpairing segment of element 9L (U259A A261C) increases KM for GTP from 29 to 120 microM, but does not otherwise affect splicing activity . The primary consequence of this mutation is a decrease in GTP binding energy of approximately 0.9 kcal/mol . Last, we show that a mutation in the highly conserved nonpairing segment of element 2 (A301C A302G G303C) eliminates transesterification activity, but does not affect 3' splice site hydrolysis. FEBS Lett, 1987 Oct 19, 223(1), 92 - 6 Amino acid sequence of iron-superoxide dismutase from Pseudomonas ovalis; Isobe T et al.; The amino acid sequence of iron-superoxide dismutase from Pseudomonas ovalis was deduced by the analyses of peptides derived from limited hydrolysis of the aminoethylated or pyridylethylated apoprotein with trypsin, Staphylococcus aureus V8 protease, and dilute acid hydrolysis . The polypeptide chain contains 195 amino acid residues and has a calculated Mr of 21,421 . The sequence is highly homologous (65% identity) to the recently published sequence of the iron-superoxide dismutase from Photobacterium leiognathi . It is also homologous to the known sequences of the manganese-superoxide dismutase by sharing 33-53% identical residues . Alignment of the superoxide dismutase sequences and the available structural information from X-ray crystallography suggest that the ligands to the iron in the P . ovalis superoxide dismutase are His-26, His-74, Asp-156 and His-160, which align with the ligands to the manganese in the Thermus thermophilus manganese-superoxide dismutase . The sequence information of the P . ovalis dismutase will facilitate refinement of the X-ray crystallographic data that are now available at 2.9 A resolution. Biochem Biophys Res Commun, 1987 Oct 14, 148(1), 397 - 402 A 31P-NMR study on multilamellar liposomes formed from the lipids of a thermophilic bacterium; Pinheiro TJ et al.; The membrane lipids of a thermophilic bacterium, Thermus SPS11, isolated from thermal springs in Sao Pedro do Sul, Portugal, were fractionated by chromatography on silica gel . The total lipid extract was found to contain one major phospholipid (PL), which accounts for about 90% of the total lipid phosphorous, and one major glycolipid (GL), which accounts for about 95% of the total carbohydrate in the non-phospholipid fraction . The membranes also contain about 11% by weight of a complex mixture of carotenoids (CA) . Multilamellar liposomes, in excess water, formed from PL and mixtures of PL with GL and CA in proportions found in the natural membrane were investigated by proton-decoupled 31P-nuclear magnetic resonance (NMR) spectroscopy and X-ray diffraction . All mixtures examined were found to be in a lamellar phase with disordered hydrophobic chains with no evidence for "non-bilayer structures" between 23 degrees and 85 degrees C . Compared to bilayers formed from pure PL or mixtures of PL and CA, significantly larger values for the chemical shift anisotropy of the 31P-NMR powder patterns were obtained from bilayers formed from mixtures of PL and GL, at temperatures above 75 degrees C, and mixtures of PL, GL and CA at all temperatures examined . These differences are interpreted in terms of changes in the order of the bilayer and/or changes in the orientation of the phosphate moiety of PL . The significance of these results to the thermophily of the bacterium is discussed. Nucleic Acids Res, 1987 Oct 12, 15(19), 7735 - 47 Sequences implicated in the processing of Thermus thermophilus HB8 23S rRNA; Hartmann RK et al.; Nuclease S1 mapping analyses were performed in order to detect processing intermediates of pre-23S rRNA from Thermus thermophilus HB8 . Two processing sites were identified downstream the start of transcription and several consecutive cleavage sites are associated with the mature 5'-end . In the 3'-flanking region one "primary" site and two cleavages which generate short-living intermediates were detected . A series of successive intermediates in the region of the mature 3'-end implies the existence of--in analogy to Escherichia coli--a 3'-exonucleolytic activity . The data were correlated with potential secondary structures within the pre-23S rRNA, which exhibit various repeated sequence elements . M13 sequencing data support the existence of one secondary structural element associated with the strong "primary" cleavage site in the 3'-flanking region . In T . thermophilus we can exclude the formation of an extended base-paired and precursor-specific stem enclosing the 23S rRNA which is inferred to mediate recognition by RNase III in E . coli. Epidemiol Infect, 1987 Oct, 99(2), 275 - 82 Serogroups of thermophilic campylobacters from humans and from non-human sources, Israel 1982-1985; Rogol M et al.; The distribution of serogroups of thermophilic campylobacters isolated in Israel from human patients (2421 isolates), chicken (942), turkeys (158), cattle (398), wild birds (234) and other sources, was studied . Among the human isolates, 74 ROG-serogroups were identified . The six most commonly isolated of these (1, 18; 11; 12; 8,23; 4 and 5,39) were found frequently in chickens . Only four common serogroups in man were also common in cattle, three in turkeys and two in wild birds . Two common serogroups in man (1,18 and 5,39) were prevalent all over the country, while others were regionally distributed . When the prevalence of different serogroups in Israel was compared to that in Canada, some groups were common to both countries and others were common in only one or the other . Campylobacter jejuni accounted for 86.7% to 92.1% of the isolates from man, chickens, turkeys, cattle and most of the wild birds . C . coli was found in 34.4% of isolates from cattle egrets and in 76.5% of those from pigs. Epidemiol Infect, 1987 Oct, 99(2), 265 - 74 Survival of thermophilic campylobacters on fingertips and their elimination by washing and disinfection; Coates D et al.; A simple impression-plate technique has been used to investigate the survival of four thermophilic campylobacter strains applied to fingertips . Campylobacters suspended in 0.1% peptone water and dried on the fingertips survived for different periods of time ranging from less than 1 to greater than or equal to 4 min . However, campylobacters suspended in chicken liquor or blood survived for much longer periods . The most resilient organism was Campylobacter jejuni NCTC 11392 which, when suspended in 50% horse blood, survived for an hour . Suspensions containing 10(6)-10(7) organisms prepared in 50% blood and dried on to fingertips were removed by thorough hand washing with either soap and water or water alone followed by drying on paper towels, but persisted on wet hands . The organisms were also eliminated by wiping the hands with a tissue saturated with 70% isopropyl alcohol for 15 sec. Appl Environ Microbiol, 1987 Oct, 53(10), 2414 - 9 Difference in sporogenous bacterial populations in thermophilic (55 degrees C) and mesophilic (35 degrees C) anaerobic sewage digestion; Chen M; Spores, sporeforming vegetative cells, and asporogenous populations were enumerated in two semicontinuous anaerobic fermentors digesting municipal primary sludge at 35 and 55 degrees C for more than 87 days . In the 35 degrees C fermentor, the anaerobic total population was 312.5 X 10(6)/ml, with 25.0 X 10(6)/ml being sporogenous . The populations that digest casein, starch, pectin, and cellulose were 23.1 X 10(6), 59.2 X 10(6), 26.2 X 10(6), and 7.3 X 10(6)/ml, respectively, with 2.8 X 10(6), 6.7 X 10(6), 3.4 X 10(6), and 1.5 X 10(6)/ml being sporogenous, respectively . The sporeformers accounted for 8.0 to 20.0% of each of the respective populations . In the 55 degrees C fermentor, the anaerobic total population was 512.5 X 10(6)/ml, with 336.6 X 10(6)/ml being sporogenous . The populations that digest casein, starch, pectin, and cellulose were 97.7 X 10(6), 190.7 X 10(6), 75.8 X 10(6), and 11.2 X 10(6)/ml, respectively, with 47.8 X 10(6), 110.6 X 10(6), 43.3 X 10(6), and 5.1 X 10(6)/ml, respectively, being sporogenous . The sporeformers represented 45.5 to 65.7% of each of the respective populations . The numbers of thermophilic sporeforming vegetative cells in the 55 degrees C fermentor were 9.0 to 19.8 times higher than their counterparts in the 35 degrees C fermentor . Most sporeformers were in the vegetative state in the 35 and 55 degrees C fermentors . After 18 days of fermentation at 55 degrees C, sporeformers carried out most of the digestion; however, the digestion was shared by both sporeformers and asporogenous bacteria after 87 days of fermentation . In the 35 degrees C fermentor, asporogenous bacteria digested most of the sludge . During the 18- and 87-day experimental periods, sporeformers were never predominant. Bioorg Khim, 1987 Oct, 13(10), 1344 - 50 {Interaction of s4U8 region of tRNA Phe with tRNA-(adenine-1-)-methyltransferase from Thermus thermophilus}; Venkstern TV et al.; Photoaffinity labelling of tRNA (adenine-1-)-methyltransferase with an E . coli tRNA(Phe) derivative bearing 4-azidophenylmercuro group attached to s4U residue as well as direct photocross-linking of the native tRNA(Phe) with the enzyme via s4U residue has been studied . Both techniques labelling gave similar results, leading to covalent attachment of tRNA(Phe) to the enzyme within a specific complex . The data obtained indicate unambigously that s4U residue contacts with tRNA (adenine-1-)-methyltransferase within the corresponding specific complex. Biochimie, 1987 Oct, 69(10), 1097 - 104 An unusual rRNA operon constellation: in Thermus thermophilus HB8 the 23S/5S rRNA operon is a separate entity from the 16S rRNA operon; Hartmann RK et al.; We succeeded in identifying a promoter element within 200 base pairs upstream a transcriptional unit comprising only a 23S rRNA, 5S rRNA and a tRNA(gly) gene in Thermus thermophilus HB8 {1, 2} . This element shows a high degree of homology to the -35 and -10 consensus sequences for promoters described for Escherichia coli {3, 4} . The promoter activity was measured by the induction of the synthesis of functional chloramphenicol acetyltransferase in Escherichia coli . A region located at the transcriptional start, rich in guanosines and cytidines, is very similar in sequence to the one believed to be under stringent control in stable RNA and ribosomal protein genes of Escherichia coli {5} . Employing nuclease S1 protection we were able to determine the in vivo start of transcription, which was identical with the in vitro start using Escherichia coli RNA-polymerase . Furthermore we identified sequences in the region following the origin of transcription, which are homologous to sections in Escherichia coli rrn promoter-leader regions responsible for antitermination . Our finding of a promoter immediately preceding a 23S/5S rRNA operon proves a transcriptional decoupling of the 16S rRNA genes, a situation so far unprecedented among prokaryotes. Biol Chem Hoppe Seyler, 1987 Oct, 368(10), 1391 - 9 Structure and function of L-lactate dehydrogenases from thermophilic and mesophilic bacteria . VII . Nucleotide sequence of the lactate dehydrogenase gene from the mesophilic bacterium Bacillus megaterium . Preparation and properties of a hybrid lactate dehydrogenase comprising moieties of the B . megaterium and B . stearothermophilus enzymes; Waldvogel S et al.; The lactate dehydrogenase (LDH) gene of a mesophilic bacterium, Bacillus megaterium (DSM 090), was cloned in E . coli HB 101 using a pEMBL vector and synthetic oligonucleotide probes . The gene was strongly expressed in the vector used if the orientation of the insert allowed the LDH promoter and the vector's lac promoter to direct transcription in the same direction . The gene and its 5' and 3' flanking regions have been sequenced . Codon usage patterns of LDH genes from mesophilic and thermophilic bacilli were compared and found to be characteristically different . A hybrid gene was constructed from fragments of the LDH genes from B . stearothermophilus (coding for aa 15-100) and B . megaterium (coding for aa 101-331) . The hybrid LDH, named S100M, was more thermostable than B . megaterium LDH, less thermostabile than B . stearothermophilus LDH and unlike the two wildtype enzymes, it could not be activated by Fru-P2. J Bacteriol, 1987 Oct, 169(10), 4857 - 60 Structure and organization of the hisA gene of the thermophilic archaebacterium Methanococcus thermolithotrophicus; Weil CF et al.; A restriction fragment of Methanococcus thermolithotrophicus genomic DNA was cloned into pUC8 to produce plasmid pET9301, which complements mutations in the hisA gene of Escherichia coli . Sequencing the DNA (2,155 base pairs) cloned from this thermophilic methanogen demonstrated that the M . thermolithotrophicus hisA gene is located within a cluster of open reading frames (ORFs) and is 68 and 69% homologous at the nucleotide level to the hisA genes of the mesophilic methanococci M . voltae and M . vannielii, respectively . The ORF (ORF 206) immediately 5' to the hisA gene of M . thermolithotrophicus is partially deleted in the genomes of the two mesophilic species, whereas ORF 114, which is 5' to ORF 206, is conserved in all three species. J Biochem (Tokyo), 1987 Oct, 102(4), 875 - 83 Single-site catalysis of F1-ATPase from thermophilic bacterium PS3 and its dominance in steady-state catalysis at low ATP concentration; Yohda M et al.; Single-site catalysis by F1-ATPase from a thermophilic bacterium PS3 (TF1) was examined by incubating the enzyme with a submolar amount of radioactive ATP . The profile of single-site catalysis by TF1 at 23 degrees C was different from that of beef heart mitochondrial F1-ATPase (MF1) . ATP hydrolysis on the enzyme and release of the products was rapid, and subsequent addition of non-radioactive ATP (cold chase) did not promote the hydrolysis of radioactive ATP, indicating that the rate-limiting step was not the step of product release but the step of ATP binding to the enzyme . Thus, the characteristic features of so-called uni-site catalysis were not observed . At 60 degrees C, whether in the presence or absence of phosphate ion, a small amount of bound {alpha, gamma-32P}ATP and cold chase promotion were observed . However, since bound 32P1 was not detected by centrifugal gel filtration, it is not yet certain whether TF1 has typical uni-site characteristics . Based on the hydrolytic turnover rate for single-site catalysis and analysis of the kinetics of steady-state catalysis, it is proposed that single-site catalysis is dominant even in steady-state catalysis at ATP concentrations of less than about 20 microM. Biochimie, 1987 Oct, 69(10), 1105 - 12 Covalent cross-linking of AcVal-tRNA to Tetrahymena thermophila cytoplasmic ribosomes and two of its 17S rRNA mutants; Nurse K et al.; Tetrahymena thermophila 80S ribosomes have been cross-linked to non-enzymatically bound AcVal-tRNA, presumably at the ribosomal P-site . Like the ribosomes from Escherichia coli, yeast, and Artemia salina, cross-linking is exclusively to C-1609, the equivalent of the E . coli C-1400 residue . Mutation of the RNA from G-1707 to A or from U-1711 to C which results in resistance to paromomycin or hygromycin, respectively, failed to affect the rate, yield, or site of cross-linking . The presence of the antibiotics during cross-linking also was without effect . It is concluded that at these two positions the base changes made do not interfere with the tertiary structure of the decoding site. Biochim Biophys Acta, 1987 Sep 24, 915(2), 225 - 37 Structural and spectroscopic comparison of manganese-containing superoxide dismutases; Bjerrum MJ; Predicted secondary structures and optical properties of four manganese-containing superoxide dismutases isolated from Saccharomyces cerevisiae, Bacillus stearothermophilus, Escherichia coli and human liver are compared . The structural predictions are further compared with the known crystal structure of the manganese-containing superoxide dismutase from Thermus thermophilus HB8 . The secondary structures of the four dismutases are predicted by the methods of Chou and Fasman (Adv . Enzymol . 47 (1978) 45-148), Garnier et al . (J . Mol . Biol . 120 (1978) 97-120) and Lim (J . Mol . Biol . 88 (1974) 873-894) . The three models show satisfactory agreement and predict that the enzymes have a mixed alpha-helix and beta-sheet structure, and that they have homologous structures . The former conclusion is also reached from an analysis of the hydrophobic character of the amino-acid sequences of the four proteins according to Kyte and Doolittle (J . Mol . Biol . 157 (1982) 105-132) . The calculation of the secondary structure based on the 185-260 nm circular dichroism spectrum of manganese-containing superoxide dismutase from S . cerevisiae reveals that the enzyme consists of 61% alpha-helix, 13% beta-sheet, 11% turn and 8% random coil conformations, which is in good accordance with the prediction based on the amino-acid sequences . Comparison of the 400-700 nm circular dichroism spectra of manganese-containing superoxide dismutase from S . cerevisiae, E . coli and T . thermophilus demonstrates that manganese atoms have homologous coordination in the three enzymes . This investigation based on primary structures and spectral properties indicates that the four dismutases have the same overall structure . Since the structural predictions are in good agreement with the structure found for the manganese-containing superoxide dismutase from T . thermophilus HB8, it can be concluded that this structure is representative for the four enzymes and probably for manganese-containing superoxide dismutases in general. Eur J Biochem, 1987 Sep 15, 167(3), 595 - 600 The photochemical reaction center of Chloroflexus aurantiacus is composed of two structurally similar polypeptides; Shiozawa JA et al.; A method has been devised which allowed the isolation of highly purified reaction center from the thermophilic green bacterium, Chloroflexus aurantiacus . The procedure consisted of three chromatography steps . The final step was fast protein liquid chromatography on Mono Q in the presence of nonanoyl-N-methylglucamide (Mega-9) . The purified reaction center complex was photochemically active and had an A280/A813 of 1.4 or less . Under non-denaturing conditions, a pigmented protein band having a Mr of 52,000-55,000 was observed in sodium dodecyl sulfate gels . When the isolated complex was heat-dissociated in the presence of sodium dodecyl sulfate, just two polypeptides having very similar Mr (24,000 and 24,500) were observed . Two protein bands were also observed in two-dimensional isoelectric focusing/sodium-dodecyl-sulfate polyacrylamide gel electrophoresis; the PI values of the two polypeptides were 6.5 and 6.7 . Partial peptide mapping of the two isolated subunits, using both enzymatic and chemical cleavage techniques, yielded almost identical patterns which indicated a high degree of sequence homology between the two polypeptides . The N-terminal amino acid sequences of the two polypeptides were identical and did not exhibit any homology to reaction center subunits of purple sulfur bacteria . The Chloroflexus reaction center is believed to be composed of one molecule of each polypeptide, the photoactive bacteriochlorophyll a dimer and, as accessory pigments, an additional bacteriochlorophyll a and three bacteriopheophytins . Hence, it appears to be the smallest photochemically active reaction center isolated to date. Eur J Biochem, 1987 Sep 15, 167(3), 475 - 9 A novel archaebacterial NAD+-dependent alcohol dehydrogenase . Purification and properties; Rella R et al.; An NAD+-dependent alcohol dehydrogenase (alcohol: NAD+ oxidoreductase, EC 1.1.1.1) was detected in cellular extracts of the extreme thermophilic archaebacterium Sulfolobus solfataricus . The enzyme was purified to homogeneity and shown to be a dimer with a native molecular mass of 71 kDa by sucrose gradient centrifugation and SDS electrophoresis . The enzyme has a broad substrate specificity that includes linear and branched primary alcohols, linear and cyclic secondary alcohols, linear and cyclic ketones and anisaldehyde . The enzyme has an extraordinary thermophilicity and a remarkable thermostability, and appears to have some properties and a structure different from those previously described for thermophilic alcohol dehydrogenases. Cell, 1987 Sep 11, 50(6), 951 - 61 Selection of circularization sites in a group I IVS RNA requires multiple alignments of an internal template-like sequence; Been MD et al.; Circularization and reverse circularization of the Tetrahymena thermophila rRNA intervening sequence resemble the first and second steps in splicing, respectively . However, site-specific base substitutions show that different nucleotides are involved in selection of the 5' splice site and the circularization sites . Furthermore, a substitution at the major circularization site that prevents circularization can be suppressed by second substitutions at two different nucleotide positions . A model is proposed in which adjacent and overlapping sequences can function as a binding site, forming a short duplex with the sequence at the circularization site and thus directing circularization and reverse circularization . Because the 5' exon-binding site and three potential circularization binding sites fall within a contiguous eight nucleotide region, this sequence may translocate relative to the catalytic core of the ribozyme in a template-like manner. J Biol Chem, 1987 Sep 5, 262(25), 11979 - 81 A new naturally occurring polyamine containing a quaternary ammonium nitrogen; Oshima T et al.; A new polyamine, tetrakis(3-aminopropyl)ammonium, N+ (CH2CH2CH2NH2)4, was identified in cells of an extreme thermophile, Thermus thermophilus . This compound was chemically synthesized and its chemical properties were coincident with those of the amine isolated from the thermophile. Biochim Biophys Acta, 1987 Sep 4, 921(1), 7 - 12 Pathway of taurolipid B formation from exogenous taurolipid A by Tetrahymena thermophila; Kaya K et al.; When Tetrahymena thermophila was incubated with taurolipid A isolated from T . pyriformis NT-1, the exogenously added taurolipid A was deacylated rapidly, and taurolipid B content in the cells was increased . The deacylated taurolipid A (lysotaurolipid A) content reached a maximum early on during incubation, and then declined . Taurolipid B and lysotaurolipid B contents in the cells were increased continuously during the incubation . These observations suggest that lysotaurolipid A was an intermediate for taurolipid B formation . When cells were incubated with lysotaurolipid A, newly formed lysotaurolipid B and taurolipid B were observed . Furthermore, when cells were incubated with lysotaurolipid B, only taurolipid B was newly formed . In contrast, newly formed lysotaurolipid B was observed when cells were incubated with exogenous taurolipid B . From the results, we have postulated the biosynthetic pathway of taurolipid B from exogenous taurolipid A in cells of Thermophila. Equine Vet J, 1987 Sep, 19(5), 442 - 7 Technique for assessing respiratory health hazards from hay and other source materials; Clarke AF et al.; This paper describes and compares three techniques of categorisation of hay, straw and other feeds and beddings collected from stables . A hand-held sampler was used to categorise samples according to the presence of plant material, fungal spores and dust mites . An Andersen sampler was used to categorise samples according to the thermotolerances of fungi and actinomycetes . An aerodynamic particle sizer was used to categorise samples according to respirable particle release rates . The highest burden of respirable particles was associated with the presence of thermophilic and thermotolerant actinomycetes and fungi . The portable slit sampler proved to be an accurate, quick and simple semiquantitative method of assessing the mould contamination of source materials . This latter technique requires only a microscope and the sampler, and is thus ideal for veterinary practices and small diagnostic laboratories. Biol Chem Hoppe Seyler, 1987 Sep, 368(9), 1167 - 77 Structure and function of L-lactate dehydrogenases from thermophilic and mesophilic bacteria, VI . Nucleotide sequences of lactate dehydrogenase genes from the thermophilic bacteria Bacillus stearothermophilus, B . caldolyticus and B . caldotenax; Zulli F et al.; Based on the previously determined amino-acid sequence of lactate dehydrogenase from B . stearothermophilus, an oligonucleotide probe was synthesized and used to clone the structural genes for lactate dehydrogenase from B . stearothermophilus, B . caldolyticus and B . caldotenax . The nucleotide sequences of the entire LDH genes from these three thermophilic bacilli were determined by the method of Maxam and Gilbert . The nucleotide sequence of the LDH gene from B . stearothermophilus is exactly identical to the one published recently; it agrees with the experimentally determined amino-acid sequence except at three positions . The amino-acid homologies among these thermophilic enzymes are 90% or more . The LDH genes are efficiently expressed in E . coli. Appl Environ Microbiol, 1987 Sep, 53(9), 2111 - 8 Enzymatic methylation of sulfide, selenide, and organic thiols by Tetrahymena thermophila; Drotar A et al.; Cell extracts from the ciliate Tetrahymena thermophila catalyzed the S-adenosylmethionine-dependent methylation of sulfide . The product of the reaction, methanethiol, was detected by a radiometric assay and by a gas-chromatographic assay coupled to a sulfur-selective chemiluminescence detector . Extracts also catalyzed the methylation of selenide, and the product was shown by gas chromatography-mass spectrometry to be methaneselenol . The sulfide and selenide methyltransferase activities copurified with the aromatic thiol methyltransferase previously characterized from this organism (A.-M . Drotar and R . Fall, Pestic . Biochem . Physiol . 25:396-406, 1986), but heat inactivation experiments suggested the involvement of distinct sulfide and selenide methyltransferases . Short-term toxicity tests were carried out for sulfide, selenide, and their methylated derivatives; the monomethylated forms were somewhat more toxic than the nonmethylated or dimethylated compounds . Cell suspensions of T . thermophila exposed to sulfide, methanethiol, or their selenium analogs emitted methylated derivatives into the headspace . These results suggest that this freshwater protozoan is capable of the stepwise methylation of sulfide and selenide, leading to the release of volatile methylated sulfur or selenium gases. Genetics, 1987 Sep, 117(1), 13 - 23 Nucleo-cytoplasmic interaction during macronuclear differentiation in ciliate protists: genetic basis for cytoplasmic control of SerH expression during macronuclear development in Tetrahymena thermophila; Doerder FP et al.; A novel class of mutations affecting the developmental expression of SerH cell surface antigen genes of Tetrahymena thermophila is described . Unlike previous categories of mutation, the four independently isolated mutations of this class act through the cytoplasm to affect SerH genes during macronuclear development . That is, macronuclei which develop under the influence of mutant cytoplasm do not subsequently express H, most likely because the developmental processing of SerH genes is affected . The cytoplasmic effect is specific for the SerH locus and is independent of which SerH allele is present . In place of H, hitherto unknown antigens are expressed . Expression of SerH can be rescued during development either by wild-type cytoplasm exchanged between conjugants or by the homozygous wild-type genotype . The mutations segregate independently of the SerH genes and identify one, possibly two, bistable genes . Possible models to explain these results are discussed. Acta Paediatr Scand, 1987 Sep, 76(5), 754 - 62 Acute gastroenteritis in children attending day-care centres with special reference to rotavirus infections . I . Aetiology and epidemiologic aspects; Hjelt K et al.; Acute gastroenteritis (GE) among 214 children (aged 6 months-7 years) attending day-care centres (DDCs) in the Copenhagen County was studied during a 12-month period . A total of 197 cases of GE was observed in 109 children (i.e . 51% of the participants) . The aetiology was as follows: rotavirus (n = 48) (24%), pathogenic bacteria (n = 11) (6%), Giardia lamblia (n = 3) (2%), while the aetiology of 68% remains unknown . The pathogenic bacteria included Yersinia enterocolitica, thermophilic Campylobacter, Clostridium difficile (+/- toxin) and enteropathogenic E . coli . In 4% of the GE the infections were multiple and Cryptosporidium was seen in one of these cases . The rate of GE declined with age from 1.35 GE per child per year (age group 1.0- less than 2.0 years) to 0.36 (6.0- less than 8.0 years) . Serum sampled at the start of the study period showed that the frequency of detectable rotavirus IgG increased with age from 48% in the 6 months- less than 1.0 year group to 96% in the 4.0- less than 7.0 year group . The highest rates of rotavirus GE occurred from January to April (i.e . the rotavirus season) . Moreover, rotavirus GE was almost absent after the age of 4 . Hence, the rates of rotavirus GE per rotavirus season per child were 0.80 (age group 6 months-less than 1.0 year), 0.32 (1.0-less than 2.0), 0.14 (2.0-less than 3.0), 0.16 (3.0-less than 4.0), 0.06 (4.0-less than 5.0) and 0.04 (5.0-less than 6.0) . Only 2 out of the 48 rotavirus GE were reinfections.(ABSTRACT TRUNCATED AT 250 WORDS) J Clin Microbiol, 1987 Sep, 25(9), 1747 - 52 Prevalence and characterization of hippurate-negative Campylobacter jejuni in King County, Washington; Totten PA et al.; A total of 593 strains of thermophilic Campylobacter species were isolated either from humans with diarrhea or from poultry in King County, Washington . Of these strains, 98 (52 hippurate-positive strains and all 46 of the hippurate-negative strains) were selected for further phenotypic characterization and genetic classification . Hippurate hydrolysis, the test typically used to differentiate Campylobacter jejuni and C . coli, did not always correlate with the genetic classification . All hippurate-positive strains were classified as C . jejuni . Of the hippurate-negative strains, 20% were C . jejuni, 78% were C . coli, and 2% were C . laridis . Assuming that the remaining hippurate-positive strains were all C . jejuni, then hippurate-negative C . jejuni represented a small percentage (9 of 556 or 1.6%) of C . jejuni strains but a significant percentage (9 of 46 or 20%) of hippurate-negative strains . This finding suggests that hippurate hydrolysis should not be used as the sole criterion for differentiating thermophilic Campylobacter species, particularly when describing the disease states associated with these organisms. Biochimie, 1987 Sep, 69(9), 975 - 82 5S RNA-protein complexes released by EDTA treatment of 60S ribosomal subunits of Tetrahymena thermophila; Hayes F et al.; Treatment of large (60S) subunit of the cytoplasmic ribosome of the protozoa Tetrahymena thermophila with EDTA causes quantitative release of 5S rRNA associated with variable non quantitative amounts of one or more of 60S proteins L4, L15, L24, L31 and L41 . The composition of the group of proteins released with 5S rRNA depends on both the molar ratio of EDTA and 60S subunits and the concentration of 60S subunits, in treatment mixtures. Biol Chem Hoppe Seyler, 1987 Sep, 368(9), 1157 - 66 Structure and function of L-lactate dehydrogenases from thermophilic and mesophilic bacteria, V . The complete amino-acid sequence of the mesophilic L-lactate dehydrogenase from Bacillus megaterium; Stangl D et al.; The complete amino-acid sequence of L-lactate-dehydrogenase from the mesophilic Bacillus megaterium was determined . 92% of the 318 amino acids were established by sequence analysis of the N-terminus, of four CNBr fragments and of one fragment obtained by cleavage with BNPS-skatole . The primary structure was completed by sequencing overlapping fragments obtained by further cleavage of suitable CNBr fragments and BNPS fragments with either trypsin, endoproteinase Lys-C, o-iodosobenzoic acid or hydroxylamine . The C-terminal amino acids were determined by degradation with carboxypeptidase A . The sequence homology between lactate dehydrogenases from B . megaterium and those from other Bacilli is 59-61% and 35-37% to those from higher organisms . The high sequence homology among lactate dehydrogenases from Bacilli, adapted to different temperatures, allows comparative studies of the structural basis of protein thermostability. Appl Environ Microbiol, 1987 Sep, 53(9), 2165 - 70 Antimicrobial activity of lysozyme against bacteria involved in food spoilage and food-borne disease; Hughey VL et al.; Egg white lysozyme was demonstrated to have antibacterial activity against organisms of concern in food safety, including Listeria monocytogenes and certain strains of Clostridium botulinum . We also found that the food spoilage thermophile Clostridium thermosaccharolyticum was highly susceptible to lysozyme and confirmed that the spoilage organisms Bacillus stearothermophilus and Clostridium tyrobutyricum were also extremely sensitive . Several gram-positive and gram-negative pathogens isolated from food poisoning outbreaks, including Bacillus cereus, Clostridium perfringens, Staphylococcus aureus, Campylobacter jejuni, Escherichia coli O157:H7, Salmonella typhimurium, and Yersinia enterocolitica, were all resistant . The results of this study suggest that lysozyme may have selected applications in food preservation, especially when thermophilic sporeformers are problems, and as a safeguard against food poisoning caused by C . botulinum and L . monocytogenes. J Bacteriol, 1987 Sep, 169(9), 4302 - 7 Cloning of the debranching-enzyme gene from Thermoanaerobium brockii into Escherichia coli and Bacillus subtilis; Coleman RD et al.; The gene for an enzyme with single or dual specificity on complex carbohydrates has been transferred from its native host (Thermoanaerobium brockii), a thermophilic anaerobe, into Escherichia coli and Bacillus subtilis . Most of the gene coding region is in a 2.2-kilobase PstI fragment that is common to the E . coli and B . subtilis chimeric vectors pCPC902 and pCPC903, respectively . Although the T . brockii debranching enzyme secreted from B . subtilis was unglycosylated and had less thermostability, more enzyme was secreted from B . subtilis (0.80 to 1.0 U/ml) than from T . brockii (0.23 U/ml) . E . coli did not export any measurable enzyme . From the fermentation broth of B . subtilis containing pCPC903, three active species of the debranching enzyme were separated; two species are possibly protease digestion products of the larger protein (105,000 molecular weight) . Whereas the enzyme can cleave all of the alpha-1----6 glucosidic linkages (and none of the alpha-1----4 bonds) in pullulan, it hydrolyzed mostly alpha-1----4 and very few of the alpha-1----6 linkages in starch . Upon hydrolysis of pullulan by the enzyme, only maltotriose was produced, while starch was digested to various-sized oligomers. Appl Environ Microbiol, 1987 Sep, 53(9), 2077 - 81 Inactivation of animal viruses during sewage sludge treatment; Spillmann SK et al.; Using a previously developed filter adsorption technique, the inactivation of a human rotavirus, a coxsackievirus B5, and a bovine parvovirus was monitored during sludge treatment processes . During conventional anaerobic mesophilic digestion at 35 to 36 degrees C, only minor inactivation of all three viruses occurred . The k' values measured were 0.314 log10 unit/day for rotavirus, 0.475 log10 unit/day for coxsackievirus B5, and 0.944 log10 unit/day for parvovirus . However, anaerobic thermophilic digestion at 54 to 56 degrees C led to rapid inactivation of rotavirus (k' greater than 8.5 log10 units/h) and of coxsackievirus B5 (k' greater than 0.93 log10 unit/min) . Similarly, aerobic thermophilic fermentation at 60 to 61 degrees C rapidly inactivated rotavirus (k' = 0.75 log10 unit/min) and coxsackievirus B5 (k' greater than 1.67 log10 units/min) . Infectivity of parvovirus, however, was only reduced by 0.213 log10 unit/h during anaerobic thermophilic digestion and by 0.353 log10 unit/h during aerobic thermophilic fermentation . Furthermore, pasteurization at 70 degrees C for 30 min inactivated the parvovirus by 0.72 log10 unit/30 min . In all experiments the contribution of temperature to the total inactivation was determined separately and was found to be predominant at process temperatures above 54 degrees C . In conclusion, the most favorable treatment to render sludge hygienically safe from the virological point of view would be a thermal treatment (60 degrees C) to inactivate thermolabile viruses, followed by an anaerobic mesophilic digestion to eliminate thermostable viruses that are more sensitive to chemical and microbial inactivations. J Biol Chem, 1987 Aug 15, 262(23), 11088 - 96 31P and 13C NMR analyses of the energy metabolism of the thermophilic anaerobe Clostridium thermocellum; Tolman CJ et al.; The energy metabolism of an anaerobic obligate thermophile, Clostridium thermocellum, has been examined as a function of incubation temperature using 31P NMR spectroscopy . Specifically investigated were the generation and availability of ATP as a function of temperature, activation energies for key processes in energy metabolism including formation of a pH gradient across the cell membrane, transport of key nutrients, and initial steps in glycolysis, and the existence of a membrane phase transition in the intact organism . Cells generate ATP via glycolysis at all temperatures examined; hence, limitation of the energy supply is not directly responsible for the lack of growth of this organism at low temperatures . Estimations of activation energies show a distinct hierarchy in the ATP-utilizing reactions examined . Conservation of ATP hydrolysis energy as delta pH has the lowest activation energy (less than or equal to 4 kcal/mol), two transport processes exhibit 10 kcal/mol activation energies, and early phosphorylation steps in glycolysis have significantly higher activation energies (approximately 25 kcal/mol) . Neither the membrane-bound ATPase responsible for formation of the pH gradient nor the permease involved in phosphate transport shows evidence of a change in behavior around the phase transition temperature determined for extracted lipids of C . thermocellum . Line widths of inorganic phosphate do show a break in behavior around 35-40 degrees C . Possible explanations for this behavior are discussed. J Mol Biol, 1987 Aug 5, 196(3), 517 - 24 Identification of a vegetative promoter in Myxococcus xanthus . A protein that has homology to histones; Komano T et al.; A physical map of 330 x 10(3) base-pairs near the replication origin of Myxococcus xanthus chromosome has been established already . Using DNA fragments from this region, Northern blot hybridization analysis was carried out in order to identify the genes expressed during vegetative growth . One of the genes, tentatively designated as vegA, was cloned and its entire DNA sequence was determined . The amino acid sequence of the gene product deduced from the DNA sequence reveals that the VegA protein is a very basic protein with a molecular weight of 18,700 . The gene was expressed in Escherichia coli using an expression vector, and its gene product was identified using SDS/polyacrylamide gel electrophoresis . From the results of S1 nuclease mapping, the vegA promoter was found to contain the sequence TAGACA at the -35 region and the sequence AAGGGT at the -10 region . These two regions are separated by 18 nucleotides . Genetic analysis suggests that the vegA gene may be essential for the growth of M . xanthus . From a computer-aided search for homologies to know protein structures, it was found that the VegA protein has homologies to histone H4 of Tetrahymena thermophila and histone H2B of sea urchin. J Cell Sci, 1987 Aug, 88 ( Pt 1), 47 - 55 Isolation and characterization of a mutant of Tetrahymena thermophila blocked in secretion of lysosomal enzymes; Hunseler P et al.; The development of a sensitive screening procedure for mutants of Tetrahymena thermophila blocked in secretion of lysosomal enzymes is described . By means of this procedure a mutant blocked in secretion of lysosomal enzymes has been isolated . This sec- mutant, MS-1, is constitutively blocked in release of at least six lysosomal enzymes, under both nutrient and non-nutrient conditions . MS-1 possesses, bound within the cell, the same amount of active lysosomal enzymes as the wild type . During starvation in media of low ionic strength MS-1 develops a highly vacuolated phenotype . This phenotype is caused by the sec- allele . It is reversed to a normal cell shape when the mutant is transferred to isotonic medium . The sec- mutant MS-1 contains mucocysts and is capable of inducing exocytosis of these secretory organelles, suggesting that Tetrahymena possesses at least two independent protein-secreting organelles. Zentralbl Bakteriol Mikrobiol Hyg {A}, 1987 Aug, 266(1-2), 94 - 103 Occurrence, serotypes and biotypes of thermophilic Campylobacters isolated in Vienna; Hirschl AM et al.; During the 1982-1986 period of all bacterial pathogens found to have caused diarrhoea, 35% belonged to the genus Campylobacter (C) . Approximately 70% of the strains were isolated from persons under the age of 30 years, with a distinct peak of occurrence in the autumn . Biotyping and serotyping according to Lior yielded the following results: C . jejuni biotype I: 32.9%, C . jejuni biotype II: 48.6%, C . coli biotype I: 10.3%, C . coli biotype II: 8.2% . From the 121 strains serotyped, 118 (97.5%) were typable . The serotypes most frequently encountered were type 1 (15.7%), 4 (9.9%), 2 and 11 (7.4% each) . There were 2 familial outbreaks of Campylobacter enteritis which could be completely elucidated by biotyping and serotyping . One outbreak was caused by C . jejuni biotype I serotype 11, the other by C . jejuni biotype II serotype 6 . Considering the frequent occurrence of Campylobacter infections, isolates should be routinely typed . The existing typing methods and schemes are highly developed. J Bacteriol, 1987 Aug, 169(8), 3792 - 800 Specialized cell surface structures in cellulolytic bacteria; Lamed R et al.; The cell surface topology of various gram-negative and -positive, anaerobic and aerobic, mesophilic and thermophilic, cellulolytic and noncellulolytic bacteria was investigated by scanning electron microscopic visualization using cationized ferritin . Characteristic protuberant structures were observed on cells of all cellulolytic strains . These structures appeared to be directly related to the previously described exocellular cellulase-containing polycellulosomes of Clostridium thermocellum YS (E . A . Bayer and R . Lamed, J . Bacteriol . 167:828-836, 1986) . Immunochemical evidence and lectin-binding studies suggested a further correlation on the molecular level among cellulolytic bacteria . The results indicate that such cell surface cellulase-containing structures may be of general consequence to the bacterial interaction with and degradation of cellulose. Cell, 1987 Jul 31, 50(3), 477 - 83 Dynamics of telomere length variation in Tetrahymena thermophila; Larson DD et al.; We have analyzed the mechanism and dynamics of telomere length variation in the macronucleus of Tetrahymena thermophila . In a newly differentiated macronucleus, the average length of the telomeric repeated sequence, (C4A2 X T2G4)n, is closely regulated . In contrast, in vegetatively dividing cells in log phase, all macronuclear telomeric sequences lengthen coordinately by 3-10 bp per generation until up to 1000 bp are added . In both elongated and short telomeres, characteristic single-stranded breaks on both strands are distally located . Reduction of elongated telomeres to their original length involves either the appearance of a novel type of variant cell, incapable of net telomere elongation, or, under stationary phase conditions, a reversible removal of telomeric sequences . The demonstration that telomeres are dynamic structures provides evidence for a model of telomere length regulation by activities that add and remove telomeric repeats. Biochem Biophys Res Commun, 1987 Jul 31, 146(2), 705 - 10 In vitro mutated beta subunits from the F1-ATPase of the thermophilic bacterium, PS3, containing glutamine in place of glutamic acid in positions 190 or 201 assembles with the alpha and gamma subunits to produce inactive complexes; Ohtsubo M et al.; Using site-directed mutagenesis, Glu-190 or Glu-201 of the beta subunit of the F1-ATPase from the thermophilic bacterium PS3 were replaced with glutamine . It was possible to reconstitute complexes of the mutated beta subunits with alpha and gamma subunits, but the complexes did not have ATPase activity . It is concluded that carboxylic acid side chains of Glu-190 and Glu-201 of the beta subunit are essential for catalytic activity of F1-ATPase. Nucleic Acids Res, 1987 Jul 24, 15(14), 5681 - 97 Characterization of two types of histone H2B genes from macronuclei of Tetrahymena thermophila; Nomoto M et al.; Two histone H2B gene clones were isolated from macronuclei of Tetrahymena thermophila . Nucleotide sequences of the two clones were highly homologous within the coding region but not in the noncoding region . Comparison of the deduced amino acid sequences between the two clones showed three differences in a total of 121 amino acids . Each of the two clones contained a TAA triplet within the coding region, which appeared to code for a glutamine residue . To demonstrate the existence of histone mRNA containing UAA triplet, nuclease P1 protection mapping using total cellular RNA and nucleotide sequencing of primer extension products were carried out . The results clearly indicated that two cloned histone H2B genes were transcribed, giving rise to the major histone H2B mRNAs with a UAA triplet sequence in frame . The tentative 5'- and 3'-ends of histone H2B mRNAs were determined. J Mol Biol, 1987 Jul 20, 196(2), 441 - 2 Preliminary X-ray data for a D-amino acid amino-transferase from a novel thermophilic Bacillus; Stoddard B et al.; Crystals of the D-amino acid aminotransferase (D-ATA) from a novel thermophilic Bacillus species (Escherichia coli pICT113 cloned gene product) have been examined by X-ray analysis . The crystals grow as hexagonal prisms, with the symmetry of space group P61 or P65 (indistinguishable crystallographically) . The cell dimensions are a = b = 135 A, c = 53 A, alpha = beta = 90 degrees, and gamma = 120 degrees . The unit cell has a volume of 850,000 A3 with six asymmetric units per unit cell . There is one dimer of molecular weight 62,000 per asymmetric unit, and the crystals diffract to 2.7 A. J Mol Biol, 1987 Jul 5, 196(1), 217 - 21 Origin of the phosphate at the ligation junction produced by self-splicing of Tetrahymena thermophila pre-ribosomal RNA; Price JV; The exons of the self-splicing pre-ribosomal RNA of Tetrahymena thermophila are joined accurately in vitro, even when only 33 nucleotides of the natural 5' exon and 38 nucleotides of the natural 3' exon remain . RNA fingerprint analysis was used to identify the unique ribonuclease T1 oligonucleotide generated by exon ligation . Secondary digests of the ligation junction oligonucleotide with ribonuclease A confirmed the identity of the fragment and demonstrated that the phosphate group that forms the phosphodiester bond at the ligation junction is derived from the 5' position of a uridine nucleotide in the RNA . This observation supports the prediction that the splice junction phosphate is derived from the 3' splice site . These results emphasize the mechanistic similarities of RNA splicing reactions of the group I introns, group II introns and nuclear pre-mRNA introns. J Mol Biol, 1987 Jul 5, 196(1), 49 - 60 5' exon requirement for self-splicing of the Tetrahymena thermophila pre-ribosomal RNA and identification of a cryptic 5' splice site in the 3' exon; Price JV et al.; The intervening sequence (IVS) of the Tetrahymena thermophila ribosomal RNA precursor undergoes accurate self-splicing in vitro . The work presented here examines the requirement for Tetrahymena rRNA sequences in the 5' exon for the accuracy and efficiency of splicing . Three plasmids were constructed with nine, four and two nucleotides of the natural 5' exon sequence, followed by the IVS and 26 nucleotides of the Tetrahymena 3' exon . RNA was transcribed from these plasmids in vitro and tested for self-splicing activity . The efficiency of splicing, as measured by the production of ligated exons, is reduced as the natural 5' exon sequence is replaced with plasmid sequences . Accurate splicing persists even when only four nucleotides of the natural 5' exon sequence remain . When only two nucleotides of the natural exon remain, no ligated exons are observed . As the efficiency of the normal reaction diminishes, novel RNA species are produced in increasing amounts . The novel RNA species were examined and found to be products of aberrant reactions of the precursor RNA . Two of these aberrant reactions involve auto-addition of GTP to sites six nucleotides and 52 nucleotides downstream from the 3' splice site . The former site occurs just after the sequence GGU, and may indicate the existence of a GGU-binding site within the IVS RNA . The latter site follows the sequence CUCU, which is identical with the four nucleotides preceding the 5' splice site . This observation led to a model where where the CUCU sequence in the 3' exon acts as a cryptic 5' splice site . The model predicted the existence of a circular RNA containing the first 52 nucleotides of the 3' exon . A small circular RNA was isolated and partially sequenced and found to support the model . So, a cryptic 5' splice site can function even if it is located downstream from the 3' splice site . Precursor RNA labeled at its 5' end, presumably by a GTP exchange reaction mediated by the IVS, is also described. Mol Biol Evol, 1987 Jul, 4(4), 381 - 94 The evolution of prokaryotic ferredoxins--with a general method correcting for unobserved substitutions in less branched lineages; Fitch WM et al.; Thirty-one bacterial type ferredoxins were examined by means of the parsimony method for their phylogenetic implications . The results show reasonable relationships in that photosynthetic, thermophilic, and desulfovibrio groups are identifiable; but a number of interesting anomalies occur . These include a methanogen sequence that clusters among the desulfovibrios . There are several differences from the phylogeny of Woese . At least two duplications producing paralogous genes are demonstrated, plus the probable existence of two more . The partial internal gene duplication that doubled the length of ferredoxin is confirmed by showing that the probability of the two ancestrally reconstructed halves possessing that much similarity by chance is 10(-7) . Howard and co-workers proposed that the two halves of the Azotobacter vinelandii are reversed relative to most other sequences . A phylogeny, drawn with the halves of the azotobacter sequence (and its relatives) reversed produced a tree that had only three less nucleotide substitutions than did the tree without their halves reversed . This plus other evidence suggests that the significantly greater similarity observed across rather than within the halves is more likely the result of convergence. Mol Gen Genet, 1987 Jul, 208(3), 537 - 41 Synthesis and secretion of a heat-stable carboxymethylcellulose from Clostridium thermocellum in Bacillus subtilis and Bacillus stearothermophilus; Soutschek-Bauer E et al.; The cellulase gene celA of Clostridium thermocellum coding for the thermostable endoglucanase A was transferred from Escherichia coli to Bacillus subtilis 168 and B . stearothermophilus CU21 using plasmids derived from the Bacillus vector pUB110 . When the structural part of the gene was joined to a pUB110 promoter the recombinant plasmids (pSE102, pSE105) were stably maintained and expressed carboxymethylcellulose (CMCase) activity . In B . stearothermophilus CU21 (pSE105) the clostridial CMCase was produced over a wide temperature range up to the maximal growth temperature (68 degrees C) . In contrast to E . coli, all of the CMCase synthesized in bacilli was released into the culture medium . About 50% of the extracellular protein secreted by B . subtilis 168 (pSE102) carrying the celA gene consisted of endoglucanase A . These findings demonstrate the feasibility of producing cellulolytic enzymes from thermophilic anaerobes in bacilli. Anal Biochem, 1987 Jul, 164(1), 72 - 7 Activity staining of cellulases in polyacrylamide gels containing mixed linkage beta-glucans; Schwarz WH et al.; Endoglucanase and cellobiohydrolase components of thermophilic cellulases can be detected in situ after gel electrophoresis in the presence of sodium dodecyl sulfate by incorporating a mixed linkage beta-glucan (barley beta-glucan, lichenan) in the separation gel . Zymograms are prepared after a renaturation treatment and incubation by staining the gel with Congo red . This method is suitable for the detection of beta-glucanases with different substrate specificities cleaving beta-1,4-, beta-1,4-1,3-, or beta-1,3-glucans . Cellobiohydrolase activities can be detected by adding 4-methylumbelliferyl-beta-D-cellobioside to the incubation buffer . The gels are subsequently stained with Coomassie blue to establish identical molecular weights of beta-glucanase and protein bands . Applications of this technique for the comparison of cellulases and for the identification of cellulase components expressed from recombinant clones are presented. Nucleic Acids Res, 1987 Jun 25, 15(12), 4821 - 35 Gene organization, transcription signals and processing of the single ribosomal RNA operon of the archaebacterium Thermoproteus tenax; Kjems J et al.; The single ribosomal RNA (rRNA) operon from the extreme thermophile and archaebacterium Thermoproteus tenax was sequenced . Sites of transcriptional initiation and termination were established and the processing sites on the primary transcript were mapped with nuclease S1 . The operon contained genes coding for 16S and 23S RNAs but lacked those coding for tRNA and 5S RNA . Transcription initiates 175 bp upstream from the start of the 16S RNA gene (Wich et al., EMBO J . 6, 523-528, 1987) and terminates 49 bp downstream from the 23S RNA gene within a long pyrimidine sequence . An open reading frame downstream from the rRNA operon is transcribed . The sequences bordering both 16S and 23S RNA genes can form putative processing stems in the primary transcript that involve the whole of the 16S-23S RNA spacer . The stems contain irregular features that constitute processing signals and are conserved in other archaebacteria . The 16S RNA stem is cut prior to that of the 23S RNA and RNA maturation follows . An unusual 14 bp helix can form between the extremities of the transcript such that the whole transcript is highly structured and a fork-like structure is formed together with the processing stems . The 23S RNA sequence was aligned with other available 23S-like RNA sequences (Leffers et al., J . Mol . Biol . 195, in press): a putative secondary structure exhibiting archaebacterial-specific features was deduced using comparative sequence analyses . A rooted phylogenetic tree was also derived for the archaebacteria that confirms their division into three major subgroups. Biochemistry, 1987 Jun 16, 26(12), 3330 - 40 Guanosine binding required for cyclization of the self-splicing intervening sequence ribonucleic acid from Tetrahymena thermophila; Tanner NK et al.; We have converted the intramolecular cyclization reaction of the self-splicing intervening sequence (IVS) ribonucleic acid (RNA) from Tetrahymena thermophila into an intermolecular guanosine addition reaction . This was accomplished by selectively removing the 3'-terminal nucleotide by oxidation and beta-elimination; the beta-eliminated IVS thereby is no longer capable of reacting with itself . However, under cyclization conditions, a free guanosine molecule can make a nucleophilic attack at the normal cyclization site . We have used this guanosine addition reaction as a model system for a Michaelis-Menten kinetic analysis of the guanosine binding site involved in cyclization . The results indicate that functional groups on the guanine that are used in a G-C Watson-Crick base pair are important for the cyclization reaction . This is the same result that was obtained for the guanosine binding site involved in splicing {Bass, B . L., & Cech, T . R . (1984) Nature (London) 308, 820-826} . Unlike splicing, however, certain additional nucleotides 5' to the guanosine moiety make significant binding contributions . We conclude that the guanosine binding site in cyclization is similar to, but not identical with, the guanosine binding site in splicing . The same binding interactions used in cyclization could help align the 3' splice site of the rRNA precursor for exon ligation . We also report that the phosphodiester bond at the cyclization site is susceptible to a pH-dependent hydrolysis reaction; the phosphodiester bond is somehow activated toward attack by the 3'hydroxyl of a guanosine molecule or by a hydroxyl ion. Nucleic Acids Res, 1987 Jun 11, 15(11), 4583 - 91 The sequence of the 6S RNA gene of Pseudomonas aeruginosa; Vogel DW et al.; From the gram-negative eubacterium Pseudomonas aeruginosa we have isolated a stable 6S RNA, approximately 180 nucleotides in length . The RNA was partially sequenced and identified by comparison with the known Escherichia coli 6S RNA sequence . Southern hybridizations revealed a single copy gene coding for the 6S RNA . DNA from other prokaryotes, i.e . E . coli, Thermus thermophilus, Bacillus subtilis, Bacillus stearothermophilus and Halobacterium maris mortui, did not give detectable hybridization signals . The 6S RNA gene was cloned in E . coli and its complete primary structure was determined . Although the 6S RNA sequences from P . aeruginosa and E . coli share only a 60.4% homology, we are able to propose a common secondary structural model. J Biol Chem, 1987 Jun 5, 262(16), 7523 - 7 Nucleotide sequence of the 18-25 S ribosomal RNA intergenic region from a thermophile, Thermomyces lanuginosus; Nazar RN et al.; To identify important structural features in the intergenic sequences of ribosomal DNAs, the nucleotide sequence of the 18-25 S rRNA intergene region was determined in a thermophilic fungus, Thermomyces lanuginosus, and compared to that of common yeasts . While a striking sequence homology was observed in the mature ribosomal sequences, the two internal transcribed spacer regions were found to be unusually G + C rich and significantly shorter than any other organism . An extensive secondary structure could be predicted but estimates of the secondary structure did not reveal a minimum structure which is common to other species; rather, in all rDNA transcripts the intergenic regions appear organized into a secondary structure in which all of the mature rRNA junctions are localized within one domain . This suggests that a role of the internal transcribed spacer in ribosome maturation may be that of a "biological spring" which maintains processed sites in close proximity. Biochem J, 1987 Jun 1, 244(2), 331 - 5 Unusual ciliate-specific codons in Tetrahymena mRNAs are translated correctly in a rabbit reticulocyte lysate supplemented with a subcellular fraction from Tetrahymena; Andreasen PH et al.; The codon usage of Tetrahymena thermophila and other ciliates deviates from the 'universal genetic code' in that UAA and probably UAG are not translational termination signals but code for glutamine . Therefore, translation in vitro of mRNA from Tetrahymena in a reticulocyte lysate is prematurely terminated if a UAA or UAG triplet is present in the reading frame of the mRNA . We show that the addition of a subcellular fraction from Tetrahymena thermophila enables a rabbit reticulocyte lysate to translate Tetrahymena mRNAs into full-sized proteins . The activity of the subcellular fraction is shown to depend on the combined function of a protein component(s) and a tRNA(s) . The subcellular fraction is easily prepared and its usefulness for the identification of isolated mRNAs from Tetrahymena by their translation products in vitro is demonstrated. J Cell Biol, 1987 Jun, 104(6), 1485 - 94 Tetrahymena contain two distinct and unusual high mobility group (HMG)-like proteins; Schulman IG et al.; Previous studies have described the existence of high mobility group (HMG)-like proteins in macronuclei of the ciliated protozoan, Tetrahymena thermophila (Hamana, K., and K . Iwai, 1979, J . Biochem . {Tokyo}, 69:1097-1111; Levy-Wilson, B., M . S . Denker, and E . Ito, 1983, Biochemistry, 22:1715-1721) . In this report, two of these proteins, LG-1 and LG-2, have been further characterized . Polyclonal antibodies raised against LG-1 and LG-2 fail to cross react with each other or any other macronuclear polypeptide in immunoblotting analyses . As well, LG-1 and LG-2 antibodies do not react with calf thymus, chicken, or yeast HMG proteins . Consistent with these results, a 47 amino-terminal sequence of LG-1 has been determined that shows limited homology to both calf thymus HMGs 1 and 2 and HMGs 14 and 17 . Two internal sequences of V8 protease-generated peptides from LG-2 have been determined, and these do not share any homology to the LG-1 sequence or any other sequenced HMG proteins . Comparison of the partial sequences of LG-1 and LG-2 with the complete amino acid sequence of the Tetrahymena histone H1 (Wu, M., C . D . Allis, R . Richman, R . G . Cook, and M . A . Gorovsky, 1986, Proc . Natl . Acad . Sci . USA, 83:8674-8678) rules out the possibility that LG-1 and LG-2 are proteolytically derived from H1, the other major macronuclear perchloric acid-soluble protein . Interestingly, however, both LG-1 and LG-2 are efficiently extracted from macronuclei by elutive intercalation (Schroter, H., G . Maier, H . Ponsting, and A . Nordheim, 1985, Embo (Eur . Mol . Biol . Organ.) J., 4:3867-3872), suggesting that both may share yet undetermined properties with HMGs 14 and 17 of higher eukaryotes . Examination of the pattern of LG-1 and LG-2 synthesis during the sexual phase of the life cycle, conjugation, demonstrates that the synthesis of LG-1 and LG-2 is coordinately increased from basal levels during the differentiation of new macronuclei (7-13 h), suggesting that both of these proteins play a role in determining a macronuclear phenotype . However, a specific induction of LG-2 synthesis is detected in early stages of conjugation (meiotic prophase, 1-4 h), leading to maximal synthesis of LG-2 at 3 h . Interestingly, the early induction of LG-2 synthesis closely parallels the hyperphosphorylation of histone H1 . Taken together, these data suggest that LG-1 and LG-2 are not strongly related to each other or to higher eukaryotic HMG proteins.(ABSTRACT TRUNCATED AT 400 WORDS) J Bacteriol, 1987 Jun, 169(6), 2380 - 4 2-Methylthio-1,4-naphthoquinone, a unique sulfur-containing quinone from a thermophilic hydrogen-oxidizing bacterium, Hydrogenobacter thermophilus; Ishii M et al.; A quinone was extracted and purified from the cells of an extremely thermophilic hydrogen bacterium, Hydrogenobacter thermophilus TK-6 (IAM 12695) . Its chemical structure was determined as 2-methylthio-3-VI, VII-tetrahydromultiprenyl-1,4-naphthoquinone by elemental analysis, 1H nuclear magnetic resonance spectroscopy, mass spectroscopy, and infrared spectroscopy of the quinone and by gas-liquid chromatography and gas chromatography-mass spectroscopy analysis of the ozonolysis products of the quinone . It was shown that the other five strains of H . thermophilus have the same quinone system . We named the quinone with the 2-methylthio-1,4-naphthoquinone nucleus "methionaquinone." The abbreviation of MTK is recommended for this class of quinone. Biochem J, 1987 May 15, 244(1), 243 - 6 The interaction of spermine and pentamines with DNA; Basu HS et al.; We studied the effects of spermine, two naturally-occurring pentamines isolated from the thermophile Thermus thermophilus and one synthetic pentamine on the aggregation and 'melting' temperature of calf-thymus DNA and on the B-to-Z transition of poly(dG-me5dC) . All pentamines caused aggregation of DNA at much lower concentrations than that of spermine . Concentrations that increased the melting temperature of DNA and induced the B-to-Z transition in poly(dG-me5dC) were different for each pentamine, but were comparable with the concentration of spermine needed to cause these effects . Our results suggest that both the total charge and the distance separating the charge, which is a function of the length of the carbon chains between amino groups, are important for the induction of conformational changes in DNA . The biological role of pentamines in T . thermophilus appears to be related to their ability to promote DNA condensation at high temperature. Eur J Biochem, 1987 May 15, 165(1), 147 - 55 Purification and characterization of D-glyceraldehyde-3-phosphate dehydrogenase from the thermophilic archaebacterium Methanothermus fervidus; Fabry S et al.; The D-glyceraldehyde-3-phosphate dehydrogenase from the extremely thermophilic archaebacterium Methanothermus fervidus was purified and crystallized . The enzyme is a homomeric tetramer (molecular mass of subunits 45 kDa) . Partial sequence analysis shows homology to the enzymes from eubacteria and from the cytoplasm of eukaryotes . Unlike these enzymes, the D-glyceraldehyde-3-phosphate dehydrogenase from Methanothermus fervidus reacts with both NAD+ and NADP+ and is not inhibited by pentalenolactone . The enzyme is intrinsically stable up to 75 degrees C . It is stabilized by the coenzyme NADP+ and at high ionic strength up to about 90 degrees C . Breaks in the Arrhenius and Van't Hoff plots indicate conformational changes of the enzyme at around 52 degrees C. J Biol Chem, 1987 May 5, 262(13), 6334 - 8 Structural and functional relationship of ATP synthases (F1F0) from Escherichia coli and the thermophilic bacterium PS3; Steffens K et al.; Functional compatibility between the F1 and F0 parts of ATP synthases from Escherichia coli (EF1F0) and the thermophilic bacterium PS3 (TF1F0) was analyzed . F1-stripped everted membrane vesicles from both organisms bound the homologous or heterologous F1 part to the same extent . Titration of the reconstituted membrane vesicles with dicyclohexylcarbodiimide revealed a similar sensitivity of the homologous and hybrid F1F0 complexes towards the inhibitor . Furthermore, the heterologous enzymes exhibited ATP-dependent H+ translocation comparable to that of homologous F1F0 . Antisera raised against EF1 or subunits a, b, and c of EF0 were analyzed for cross-reactivity with TF1 and TF0 . Common antigenic sites have been detected with immunoblot analysis for subunit beta and subunit c of EF1F0 and the corresponding subunits from TF1F0 . A weak binding of the anti-a and anti-b antisera with the TF0 part has been observed in an enzyme-linked immunosorbent assay . Based on these findings the structural and functional relationship between the mesophilic and thermophilic ATP synthase complexes is discussed. Mol Cell Biol, 1987 May, 7(5), 1725 - 30 Nonintegrative transformation in the filamentous fungus Podospora anserina: stabilization of a linear vector by the chromosomal ends of Tetrahymena thermophila; Perrot M et al.; The effect of the chromosomal ends of Tetrahymena thermophila on the stability of linear transforming molecules in the filamentous fungus Podospora anserina was tested . A derivative of an integrative vector for this fungus has been constructed, so that after linearization, the ends of the plasmid are the telomeric sequences of T . thermophila . After transformation, this linear molecule was maintained as an extrachromosomal plasmid with no integrated copies in about 50% of the transformants . Under selective conditions, there was approximately one linear molecule per 5 to 10 nuclei, and these extrachromosomal molecules were rapidly lost under nonselective conditions . The circular plasmid carrying an inverted repeat of T . thermophila telomeres could be linearized and processed in vivo. Development, 1987 May, 100(1), 23 - 30 The assembly and positioning of cytoskeletal elements in Tetrahymena; Williams NE et al.; The oral skeleton of Tetrahymena is a precisely arranged assemblage of basal bodies, microtubule bundles and connecting filaments found associated with the feeding structure in this cell type . Tubulin and filament proteins have been isolated but no actin has been recovered . The conditional mutant NP1 of Tetrahymena thermophila forms a normal oral skeleton at the permissive temperature (28 degrees C), but forms an abnormal one at the restrictive temperature (37 degrees C) . Antibodies against tubulin and oral filament protein OF1 were used to visualize the abnormal oral skeleton and stages in its development, and ultrastructural comparisons of abnormal and wild-type oral skeletons were made . It was found that the overall pattern of organization was altered in the mutant, whereas the substructure appeared everywhere to be normal . Studies of cells in which the mutant phenotype was coming to expression revealed that normal basal bodies in the oral skeleton failed to move from the disordered state characteristic of early stages of development into the correct pattern of four organized clusters characteristic of later stages . Together, these results suggest that the lesion in NP1 does not affect cytoskeleton assembly per se, but instead affects a discrete mechanism responsible for the positioning of cytoskeletal elements with respect to each other after they have been formed (meta-assembly) . Reasons for suspecting the involvement of the membrane skeleton are presented. Biochem J, 1987 May 1, 243(3), 779 - 87 Purification and properties of a stable beta-glucosidase from an extremely thermophilic anaerobic bacterium; Patchett ML et al.; A beta-glucosidase (EC 3.2.1.21) was purified to homogeneity from cell-free extracts of an extremely thermophilic anaerobic bacterium . The enzyme has an Mr of 43,000 as determined by molecular-exclusion chromatography, has a pI of 4.55 and shows optimum activity at pH 6.2 . The enzyme is active against a wide range of aryl beta-glycosides and beta-linked disaccharides, with beta-galactosidase activity only slightly less than beta-glucosidase activity, and significant beta-xylosidase activity . Lineweaver-Burk plots for p-nitrophenyl beta-glucoside, o-nitrophenyl beta-glucoside and cellobiose substrates are biphasic concave-downwards . Inhibition of the beta-glucosidase by substrates and glucose is negligible . Thermal inactivation follows first-order kinetics, with t1/2 (65 degrees C) 45 h, t1/2 (75 degrees C) 47 min and t1/2 (85 degrees C) 1.4 min and a deactivation energy of 380 kJ/mol at pH 6.2 . At pH 7.0, which is the optimum pH for thermostability, t1/2 (75 degrees C) is 130 min . At 75 degrees C, at pH 6.2, the thermostability is enhanced about 8-fold by 10% (w/v) glycerol, about 6-fold by 0.2 M-cellobiose and about 3-fold by 5 mM-dithiothreitol and 5 mM-2-mercaptoethanol. J Dairy Sci, 1987 May, 70(5), 919 - 26 Enhancement of immune response in mice fed with Streptococcus thermophilus and Lactobacillus acidophilus; Perdigon G et al.; Swiss mice, fed for 8 consecutive d with 50 micrograms/d of viable cultures of Lactobacillus acidophilus and Streptococcus thermophilus, showed significant variation in their immune system . In order to study this phenomenon assays for macrophage and lymphocyte function were carried out . Both lactic acid bacteria enhanced significantly the enzymatic and phagocytic activity of peritoneal macrophages as checked against the controls and also accelerated the phagocytic function of the reticuloendothelial system as revealed by the carbon clearance test . On the 2nd d (100 micrograms), L . acidophilus reached a peak of K = .271, which remained high . Streptococcus thermophilus was effective only on the 2nd d and then decreased . The lymphocytic activity studied by immunoglobulin secreting cells was assayed by Jerne's method of plaque-forming cells (PFC) . This activity also was increased by the two microorganisms . Streptococcus thermophilus proved more effective than L . acidophilus . Lactobacillus acidophilus and S . thermophilus activated macrophages and lymphocytes and produced the same increase in the immune response of mice whether administered orally or intraperitoneally. Biochim Biophys Acta, 1987 Apr 8, 912(2), 178 - 84 Amino-acid sequence of a tetrameric, manganese superoxide dismutase from Thermus thermophilus HB8; Sato S et al.; The amino-acid sequence of a tetrameric manganese superoxide dismutase from Thermus thermophilus HB8 has been determined . The protein was cleaved with cyanogen bromide (BrCN) into four peptides and their alignment was deduced through the fragment of partial cleavage with BrCN and the peptides were produced by cleavage of the protein with o-iodosobenzoic acid . Most of the peptides were sequenced by solid phase Edman degradation . Some of the peptides were sequenced by the Edman dansyl method after sub-fragmentation by proteinase digestion . The amino-acid sequence consists of 203 residues corresponding to a subunit molecular weight of 23,144. Bioorg Khim, 1987 Apr, 13(4), 546 - 9 {Isolation and crystallization of phenylalanyl-tRNA-synthetase from Thermus thermophilus HB8}; Reshetnikova LS et al.; Method of isolation of phenylalanyl-tRNA synthetase from Thermus thermophilus HB8 is described, including chromatography on DEAE-sepharose, ammonium sulfate fractionation, hydrofobic chromatography on Toyopearl, gel filtration on ultrogel AcA-34, chromatography on phenylalanylaminohexyl-sepharose and heparine-sepharose . Yield of the purified enzyme was 10 mg from 1 kg of T . thermophilus cells . The enzyme is found to consist of two types of subunits with molecular masses 92 and 36 kDa and is likely to be a tetramer protein with molecular mass 250 kDa . Crystals of phenylalanyl-tRNA synthetase suitable for X-ray structural studies have been obtained. J Gen Microbiol, 1987 Apr, 133 ( Pt 4), 1089 - 97 Molecular cloning and nucleotide sequence of the 3-isopropylmalate dehydrogenase gene of Candida utilis; Hamasawa K et al.; A 3-isopropylmalate dehydrogenase (3-IMDH, EC 1.1.1.85) gene was cloned from a gene library of Candida utilis . One of the plasmids, pYKL30, could complement Escherichia coli leuB and Saccharomyces cerevisiae leu2 auxotrophs; a 2.2 kb HindIII fragment subcloned in pBR322 could still complement the leuB mutation . Southern hybridization confirmed that this fragment was derived from C . utilis . An open reading frame of 1089 bp that corresponded to a polypeptide of 363 amino acids, one residue shorter than the 3-IMDH of S . cerevisiae, was found in the cloned fragment . The homology between the 3-IMDHs of C . utilis and S . cerevisiae was 76.2% in nucleotides and 85.4% in amino acids . In contrast, the homology between the 3-IMDHs of C . utilis and Thermus thermophilus was much smaller and was restricted to some regions of the gene. Biol Chem Hoppe Seyler, 1987 Apr, 368(4), 353 - 67 Allophycocyanin complexes of the phycobilisome from Mastigocladus laminosus . Influence of the linker polypeptide L8.9C on the spectral properties of the phycobiliprotein subunits; Fuglistaller P et al.; The following phycobiliproteins and complexes of the allophycocyanin core were isolated from phycobilisomes of the thermophilic cyanobacterium Mastigocladus laminosus: alpha AP, beta AP, (alpha AP beta AP), (alpha AP beta AP)3, (alpha AP beta AP)3L8.9C, (alpha APB alpha AP2 beta AP3)L8.9C . The six proteins and complexes were characterised spectroscopically with respect to absorption, oscillator strength, extinction coefficient, fluorescence emission, relative quantum yield, fluorescence emission polarisation and fluorescence excitation polarisation . The interpretation of the spectral data was based on the three-dimensional structure model of (alpha PC beta PC)3 (Schirmer et al . (1985) J . Mol . Biol . 184, 257-277), which is related to the allophycocyanin trimer . The absorption and CD spectra of the complexes (alpha AP beta AP)3, (alpha AP beta AP)3L8.9C and (alpha APB alpha AP2 beta AP3)L8.9C could be deconvoluted into the spectra of the phycobiliprotein subunits . The assumptions made for the deconvolution could be checked by the synthesis of the spectra of (alpha APB beta AP)3 . The synthesised spectra are in good agreement with the corresponding measured spectra published by other authors . Considering the deconvoluted spectra the following influences on the chromophores could be ascribed to L8.9C: L8.9C neither influences the alpha AP nor the alpha APB chromophores . L8.9C shifts the absorption maximum of the beta AP chromophore to longer wavelength than the absorption maximum of the alpha AP chromophore in trimeric complexes . L8.9C increases the oszillator strength of the beta AP chromophores to about the value of the alpha AP chromophores in trimeric complexes . L8.9C turns the beta AP chromophores from sensitizing into weak fluorescing chromophores . By means of the hydropathy plot and the predicted secondary structure, a postulated three-fold symmetry in the tertiary structure of L8.9C could be confirmed. J Dairy Sci, 1987 Apr, 70(4), 738 - 45 Purification and characterization of X-prolyl-dipeptidyl-aminopeptidase from Lactobacillus lactis and from Streptococcus thermophilus; Meyer J et al.; X-Prolyl-dipeptidyl-aminopeptidase recently was found in several lactic acid bacteria . This article describes the purification of the enzymes from Lactobacillus lactis and Streptococcus thermophilus and compares their characteristics . Enzymes from both strains are serine-peptidases . They both have a molecular weight of about 165,000 daltons, an isoelectric point near 4.5, and are constituted of two subunits . The pH optimum of the enzyme isolated from L . lactis is 7.0, whereas the enzyme from S . thermophilus possesses a broad pH optimum between 6.5 and 8.2 with glycyl-L-prolyl-aminomethylcoumarin as substrate . Below pH 5, both enzymes are unstable; however, that from S . thermophilus is more rapidly denatured . The enzyme from S . thermophilus is more sensitive to heat than the corresponding enzyme from L . lactis . Enzymes from the both strains have different specificities towards various substrates and are differently effected by metals, chelators, and other inhibitors . The importance of this enzyme for the metabolism of lactic acid bacteria is discussed. J Mol Biol, 1987 Mar 20, 194(2), 181 - 92 Tetrahymena actin . Cloning and sequencing of the Tetrahymena actin gene and identification of its gene product; Hirono M et al.; Actin is ubiquitous in eukaryotes, nevertheless its existence has not yet been clearly proven in Tetrahymena . Here we report the cloning and sequencing of an actin gene from the genomic library of Tetrahymena pyriformis using a Dictyostelium actin gene as a probe . The Tetrahymena actin gene has no intron . The predicted actin is composed of 375 amino acids like other actins and its molecular weight is estimated as 41,906 . Both T . pyriformis and T . thermophila possess a single species of actin genes which differ in their restriction patterns . Northern hybridization analysis revealed that the actin gene was actively transcribed in vivo . To detect the gene product, we synthesized an N-terminal peptide of the deduced sequence and prepared its antibody . Using an immunoblotting technique, we identified Tetrahymena actin on a two-dimensional gel electrophoretic plate . The actin spot migrated near an added spot of rabbit skeletal muscle actin, but clearly differed from the latter in its isoelectric point and apparent molecular weight . The primary structure of Tetrahymena actin shares about 75% homology equally with those of other representative actins . This value is extremely low as a homology rate between known actins . Tetrahymena actin diverges not only in relatively variable regions of other actins, but also in relatively constant regions . The hydrophilicity levels of two regions (residues 190 to 200 and residues 225 to 235) are also quite different between the Tetrahymena actin and skeletal muscle actin . Thus, we conclude that actin is present in Tetrahymena, but it is one of the most unique actins among the actins known hereto. Nucleic Acids Res, 1987 Mar 11, 15(5), 1905 - 20 Characterization of an authentic intermediate in the self-splicing process of ribosomal precursor RNA in macronuclei of Tetrahymena thermophila; Kister KP et al.; We have characterized a 1.5 kb RNA species in T . thermophila macronuclei previously found in vivo and including intron sequences linked to the 3' exon . This IVS-3' exon RNA could be detected in gels as a discrete molecule only after denaturation of nuclear RNA . After addition of 32P-GTP, as splicing cofactor in a nuclear in vitro system, the IVS-3' exon RNA was labeled at its 5' terminus, as was the by-product of splicing, the excised IVS RNA . The time course of labeling indicates that the IVS-3' exon RNA acts like a reaction intermediate and specifically a kinetic precursor to IVS RNA . Partial nuclease digestions showed that the IVS-3' exon RNA and the IVS RNA have the same 5' terminal sequence . In addition the IVS-3' exon RNA can release the 15-mer oligonucleotide cleaved off during circularization of IVS RNA under conditions of high temperature . Taken together, the structural, functional, and kinetic properties of the IVS-3' exon RNA strongly suggest that it represents a previously postulated in vivo intermediate in the splicing pathway. J Bacteriol, 1987 Mar, 169(3), 1328 - 30 Effect of growth temperature on the long-chain diols and fatty acids of Thermomicrobium roseum; Pond JL et al.; Long-chain 1,2-diols constitute the hydrophobic backbone of membrane lipids (replacing glycerolipids) in the thermophilic eubacterium Thermomicrobium roseum . The effects of incubation temperature on chain length and chain branching of diols and fatty acids were investigated . The percentage of branched chains decreased, and chain length increased slightly, with increased growth temperatures. Exp Cell Res, 1987 Mar, 169(1), 63 - 73 Unidirectional co-stimulation by a non-mating strain of Tetrahymena thermophila; Katz M et al.; We report that a fatty acid auxotroph of Tetrahymena thermophila (RH179E1) fails to mate, yet retains the ability to co-stimulate normal cells unidirectionally . Thus, co-stimulation can be analyzed experimentally in the absence of pair formation . We show that the co-stimulation of normal cells of one mating type is sufficient to shorten the waiting period for pair formation of those cells with initiated cells . This is the first evidence that co-stimulation causes a hyperinduction of mating reactivity in T . thermophila, generating in turn a positive feedback mechanism for (presumably) gamone production . Co-stimulation by the variant strain is at a maximum after 3-4 h of exposure when the variant and wild-type cells are at a ratio of 1:1 . When mixed with wild-type cells, RH179E1 induces the formation of progeny (at low frequency) which inherit exclusively genetic material of the wild-type cells. J Appl Bacteriol, 1987 Mar, 62(3), 209 - 16 Conventional taxonomy of lactobacilli surviving radurization of meat; Hastings JW et al.; All of the 113 catalase-negative, Gram-positive, rod-shaped strains isolated from radurized minced beef (5 kGy) were homofermentative, non-thermophilic, and belonged to the sub-genus Streptobacterium . The majority of the strains (100) were identified as Lactobacillus sake . These were divided into four sub-groups based on their sugar fermentation pattern: group IA1 (melibiose (+), maltose (-), amygdalin (-), 76 strains); group IA2 (melibiose (+), maltose (-), amygdalin (+), 14 strains); group IB1 (melibiose (+), maltose (+), amygdalin (+), four strains); group IB2 (melibiose (+), maltose (+), amygdalin (-), six strains) . Of the remaining strains, two produced L(+)-lactic acid and were identified as L . farciminis, three were identified as L . curvatus and eight showed characteristics of both L . sake and L . curvatus and were designated 'L . sake/curvatus.' With one exception, all strains were aciduric and relatively insensitive to the chemical preservatives tested . Most L . sake strains produced significant amounts of H2O2 . Electron microscopy confirmed a possible relationship between the thickness of the cells and radiation resistance . The problems and limitations of this type of taxonomic study and possible reasons for the predominance of L . sake species in radurized meat are discussed. J Dairy Sci, 1987 Mar, 70(3), 514 - 23 Oxygen uptake activity and aerobic metabolism of Streptococcus thermophilus STH450; Teraguchi S et al.; Streptococcus thermophilus STH450 had a very high oxygen uptake . This strain was then compared with aerobic metabolism to S . thermophilus ATCC 19258, a reference strain for aerobic metabolism . Molecular oxygen, which was absorbed by S . thermophilus STH450 during aerobic glycolytic metabolism, was involved in the oxidation of NADH by the catalytic activity of NADH oxidase . The portion of pyruvate that corresponded to the oxidized NADH was committed to form alpha-acetolactate, acetoin, and diacetyl . Both strains were deficient in peroxidase and pyruvate oxidase activities; therefore, NADH oxidase was probably the terminal oxidase in aerobic glycolytic metabolism . Oxygen uptake and NADH oxidase activities were significantly higher in S . thermophilus STH450 than in S . thermophilus ATCC 19258 . alpha-Acetolactate, acetoin, and diacetyl also accumulated during aerobic glycolytic metabolism of S . thermophilus STH450 . However, when both strains were grown in the presence of pyruvate, these metabolites were equivalent . Hence, less oxygen might be needed for pyruvate metabolism. J Biochem (Tokyo), 1987 Mar, 101(3), 633 - 42 Evidence for two activated forms of pyruvate kinase from Bacillus stearothermophilus in the presence of ribose 5-phosphate; Sakai H et al.; Allosteric activation of pyruvate kinase from a thermophilic bacterium, Bacillus stearothermophilus, by ribose 5-phosphate (R5P) was kinetically examined . Two activated forms of this enzyme could be distinguished, depending on the R5P concentration . One form (Form I) was observed at about 10(-5) M R5P . It showed a slightly negative cooperativity for phosphoenolpyruvate (PEP) . The other form (Form II) was observed at more than 10(-3) M R5P and showed Michaelis-Menten kinetics for PEP . The PEP and ADP concentrations that yield half-maximal velocity were essentially identical for the two forms (about 0.1 and about 0.5 mM, respectively), but Form I had a larger Vmax value than Form II . In the absence of R5P, the enzyme showed a homotropic positive cooperativity for PEP; the concentration required for the half-maximal velocity was about 2 mM and that of ADP was about 1.6 mM . The enzyme was more susceptible to protease digestion in the presence of R5P than in the absence of it . The concentration of R5P required for the enzyme to be susceptible to protease digestion was approximately identical with that required to generate Form I . With more than 10(-3) M R5P, the thermostability of the enzyme was greatly increased . The concentration of R5P required for the enzyme to be thermostable was in good agreement with that required to generate Form II . These results indicate that the two activated forms distinguished kinetically differ in their conformations, too . The saturating level of PEP did not cause such a change in the thermostability or the susceptibility to protease. Biochim Biophys Acta, 1987 Feb 27, 908(2), 150 - 7 Purification and characterization of an inducible mitochondrial DNA polymerase from Tetrahymena thermophila; Ostergaard E et al.; Treatment of the eukaryotic organism Tetrahymena with various types of DNA-damaging agents has been reported to cause a 35-fold induction of a mitochondrial DNA polymerase . We here report that the enzyme can be induced in large-scale cultures by exposure of the cells to thymine starvation and/or intercalating agents . The induced DNA polymerase has been purified to near homogeneity, with a specific activity of approx . 300,000 units/mg protein . The relative molecular mass of the active form of the enzyme is approx . 100,000, as determined by glycerol gradient sedimentation . The subunit structure has been analysed by SDS polyacrylamide gel electrophoresis of the highly purified preparation and by immunoprecipitation with a monoclonal antibody directed to the DNA polymerase . A polypeptide of Mr 47,000 has been observed to be a subunit of the enzyme . This corresponds to the size of the subunits suggested for mitochondrial DNA polymerase from chicken embryos and mouse myeloma cells. Nucleic Acids Res, 1987 Feb 11, 15(3), 921 - 32 Syntheses of rRNA, 5.8S, 5S and tRNA are inhibited equally by 8-methoxypsoralen phototreatment of Tetrahymena thermophila; Nielsen PE; Treatment of the ciliated protozoan Tetrahymena thermophila with 8-methoxypsoralen combined with long wavelength ultraviolet irradiation (UVA, lambda approximately 360 nm) resulted in a dose dependent equal inhibition of the synthesis of rRNA, 5.8S, 5S and tRNA . Similar results were obtained with 3-carbethoxy-8-methoxypsoralen which predominantly forms DNA mono-adducts . In contrast the synthesis of tRNA in T . thermophila was much less sensitive than that of rRNA, 5.8S and 5S RNA to treatment with short wavelength ultraviolet irradiation (UVB, lambda approximately 254 nm) . These results are interpreted in favor of a mechanism by which psoralen-DNA adducts (crosslinks much greater than monoadducts) inhibit RNA transcription initiation (in contrast to UVB which causes premature chain termination) . Furthermore it is argued that RNA synthesis is regulated in equally sized domains regardless of the gene-size. J Appl Toxicol, 1987 Feb, 7(1), 35 - 41 Discovery of multiple organofluorophosphate hydrolyzing activities in the protozoan Tetrahymena thermophila; Landis WG et al.; Recently it has been found that homogenates of Tetrahymena thermophila can hydrolyze the potent acetylcholinesterase inhibitors O,O-diisopropylphosphofluoridate (DFP) and O-1,2,2-trimethylpropylmethylphosphonofluoridate (soman) . Upon purification of the DFP hydrolyzing activity 10-fold it had been noted that the soman hydrolyzing activity increased only 2-3 fold . Treatment with manganous ion and comparison of the soman and DFP hydrolysis rates of the homogenate indicated that a mixture of the squid-type and Mazur-type DFPases may be present . Subsequent purification of the enzymatic activities within the Tetrahymena-homogenate demonstrated that there are at least five functioning proteins of molecular weights 67,000 to 96,000 . None are directly homologous to the DFPases found in hog kidney or squid . The enzymatic activities are designated DFPase-1 through DFPase-5 . A hypothesis is presented that the functions of DFPases are in the normal metabolism of organophosphates naturally synthesized by T . thermophila. J Appl Bacteriol, 1987 Feb, 62(2), 167 - 76 A study of thermophilic campylobacters in a river system; Bolton FJ et al.; Fifteen kilometres of a river system traversing rural and urban areas and subject to sewage works effluent discharge was studied during a 12 1/2 month period . A total of 312 samples was collected from 12 sites at 14 d intervals and tested by a glass microfibre filtration method and a most probable number (MPN) method . Campylobacters were found in 43% of samples by the filtration method and 21% by the MPN method . The lowest frequency of isolation and lowest counts (less than 10 campylobacters/100 ml) were associated with samples collected from rural sites and fast-flowing stretches of river . The greatest frequency of isolation and highest counts (greater than 10-230 campylobacters/100 ml) were associated with sites adjacent to or downstream of sewage works . There was an obvious seasonal trend; most isolations and highest counts were obtained in late autumn and winter, and fewest isolations and lowest counts in spring and summer . Surface water run-off from adjacent farmland following heavy rainfall also increased the counts of campylobacters in the river system . Biotyping of isolates demonstrated that the most prevalent Campylobacter sp . was Campylobacter jejuni but C . coli, C . laridis and a previously unrecognized group of campylobacters were also isolated . Serotyping differentiated 14 serotypes of C . jejuni, 11 of C . coli and two of C . laridis . Furthermore, serotypes of C . jejuni commonly isolated from enteritis in man were frequently found in river water tested during this study. Biol Chem Hoppe Seyler, 1987 Feb, 368(2), 121 - 30 Ribosomal proteins and DNA-binding protein II from the extreme thermophile Bacillus caldolyticus; Beck A et al.; Crystallographic studies, presently on ribosomal and DNA-binding proteins from the moderate thermophile Bacillus stearothermophilus, can be expected to benefit from the use of even more stable proteins from extreme thermophiles . Bacillus caldolyticus, which is able to grow in the temperature range of 70-80 degrees C, appears to be a suitable candidate . We have compared the two bacilli using two criteria: the two-dimensional gel patterns of ribosomal proteins and the properties of DNA-binding protein II . The latter protein is ubiquitous in the eubacterial kingdom and can be purified in large quantities . B . caldolyticus can be grown at 75 degrees C in continuous culture with a generation time of 45-60 min . The yield of ribosomes compares favorably with that of B . stearothermophilus . The gel patterns of the ribosomal proteins are very similar but several differences, in particular among the 50S proteins, are observed . The N-terminal amino-acid sequence of the DNA-binding protein differs in 3 positions (out of 39) from B . stearothermophilus and the protein shows an increased resistance to thermal denaturation . Tetragonal and monoclinic crystals of DNA-binding protein II have been obtained which are suitable for X-ray studies and the diffraction patterns of the two crystal forms are shown. Proc Natl Acad Sci U S A, 1987 Feb, 84(3), 675 - 9 Control of oligomeric enzyme thermostability by protein engineering; Ahern TJ et al.; The ability to control the resistance of an enzyme to inactivation due to exposure to elevated temperatures is essential for the understanding of thermophilic behavior and for developing rational approaches to enzyme stabilization . By means of site-directed mutagenesis, point mutations have been engineered in the dimeric enzyme yeast triosephosphate isomerase that improve its thermostability . Cumulative replacement of asparagine residues at the subunit interface by residues resistant to heat-induced deterioration and approximating the geometry of asparagine (Asn-14----Thr-14 and Asn-78----Ile-78) nearly doubled the half-life of the enzyme at 100 degrees C, pH 6 . Moreover, in an attempt to model the deleterious effects of deamidation, we show that replacement of interfacial Asn-78 by an aspartic acid residue increases the rate constant of irreversible thermal inactivation, drastically decreases the reversible transition temperature, and reduces the stability against dilution-induced dissociation. Nucleic Acids Res, 1987 Jan 26, 15(2), 683 - 93 Structure determination of a new fluorescent tricyclic nucleoside from archaebacterial tRNA; McCloskey JA et al.; A highly fluorescent nucleoside was detected in enzymatic digests of the extremely thermophilic archaebacterium Sulfolobus solfataricus by combined liquid chromatography-mass spectrometry (LC/MS) . Following isolation, the structure was determined primarily by mass spectrometry, to be 3-(beta-D-ribofuranosyl)-4,9-dihydro-4,6,7-trimethyl-9-oxoimidazo{ 1, 2-a}purine (mimG), a new derivative of the Y (wye) nucleoside . The structural assignment was verified by comparison of the base released by acid hydrolysis with the corresponding synthetic base, using mass spectrometry, chromatography, and UV absorption and fluorescence properties . Nucleoside mimG was also detected by LC/MS in hydrolysates of the thermophiles Thermoproteus neutrophilus and Pyrodictium occultum . These results constitute the first finding of a member of the hypermodified Y family of nucleosides in archaebacteria. J Biol Chem, 1987 Jan 15, 262(2), 558 - 63 Functions of isolated domains of methionyl-tRNA synthetase from an extreme thermophile, Thermus thermophilus HB8; Kohda D et al.; Methionyl-tRNA synthetase (MetRS, 2 X 75 kDa) was purified to homogeneity from an extreme thermophile, Thermus thermophilus HB8 . The polypeptide chain of MetRS was cleaved by limited digestion with trypsin into four domains: T1 (29 kDa), T2 (23 kDa), T3 (14.5 kDa), and T4 (7.5 kDa), which were aligned in that order . MetRS was also cleaved into similar fragments with a variety of other proteases . Domains T1, T2, T3, and T4 were isolated by column chromatography . "Tandem domain" T1-T2 (56 kDa) is fully active in the aminoacylation of tRNA and is further cleaved with trypsin into domains T1 and T2 . Domain T1 is the smallest aminoacylation unit so far reported . Domain T2 (enzymatically inactive) interacts with tRNAMetf, as found by UV-induced cross-linking . Isolated domain T3 forms a dimer and is responsible for the dimer assembly of two protomers in MetRS . Domain T4 is a flexible tail of MetRS . These domains, in particular T1 and T2, will be important for detailed structure analyses in relation to aminoacylation activity. Nucleic Acids Res, 1987 Jan 12, 15(1), 141 - 60 Unusual features of transcribed and translated regions of the histone H4 gene family of Tetrahymena thermophila; Horowitz S et al.; The complete DNA sequence is presented of H4-II, the second of the pair of histone H4 genes of the ciliated protozoan, Tetrahymena thermophila . Both H4 genes code for the same protein . Codon usage in these and other Tetrahymena genes is severely restricted and is similar to that in yeast . Flanking regions are AT-rich (greater than or equal to 75%), relative to coding sequences (approximately 45% GC) . Except for small, similarly positioned homologies, flanking sequences of the two genes are different . Canonical sequences in higher eukaryotic promoters are not obvious in these genes . Instead, short, localized, base composition eccentricities characterize the 5' flanking sequences of all Tetrahymena genes analyzed . The consensus, P yP u(A)3-4 ATGG initiates translation in these and all other known Tetrahymena genes . Nuclear transcripts and messages of both growing and starved cells begin at multiple sites, mainly at the first or second A residue following a pyrimidine . The palindrome typical of histone message 3' termini in higher organisms is not present . Downstream of both genes are sequences similar to the processing/polyadenylation signal of higher eukaryotes, although the unique 3' ends are not those predicted by the location of the signals. Biochim Biophys Acta, 1987 Jan 5, 911(1), 81 - 94 Spectroscopic studies of the seven-iron-containing ferredoxins from Azotobacter vinelandii and Thermus thermophilus; Johnson MK et al.; The seven-iron-containing ferredoxins from Azotobacter vinelandii and Thermus thermophilus have been investigated by low-temperature magnetic circular dichroism (MCD) and electron paramagnetic resonance (EPR) spectroscopies and room temperature ultraviolet-visible absorption spectroscopy . The results confirm the presence of one trinuclear and one tetranuclear iron-sulfur cluster in both ferredoxins and facilitate comparison of the electronic and magnetic properties of the oxidized and reduced {3Fe-xS} clusters . MCD magnetization data are consistent with an S = 2 ground state for both reduced {3Fe-xS} clusters, but indicate differences in the rhombicity of the zero-field splittings . The data permit rationalization of the absence of a delta M = 4 EPR transition for the reduced {3Fe-xS} cluster in A . vinelandii ferredoxin I . Spectroscopic studies of anaerobically isolated A . vinelandii ferredoxin I do not support the hypothesis that the {3Fe-xS} cluster arises as a result of aerial oxidative damage to a {4Fe-4S} cluster during isolation . The possibility that two distinct forms of {3Fe-xS} clusters can exist in A . vinelandii ferredoxin I was investigated by spectroscopic studies as a function of pH . The results reveal two distinct and interconvertible forms of the reduced {3Fe-xS} cluster, but do not permit rationalization of the inconsistencies in the structural data that have been reported for the oxidized clusters. Nucleic Acids Symp Ser, 1987, (18), 277 - 80 Self-splicing of Tetrahymena rRNA can proceed with phosphorothioate substitution at the splice sites; Deeney CM et al.; The self-excision of a 413-base intervening sequence of the 26S rRNA of Tetrahymena thermophila has been investigated using phosphorothioate-substituted RNA . Transcripts containing this intron were prepared by T7 RNA polymerase-catalyzed polymerisation using a M13 mICE10 vector in the presence of various nucleoside alpha-thiotriphosphate analogues . Wild-type transcripts incorporating phosphorothioates 5' to adenosine or uridine were inactive, whereas incorporation 5' to cytidine or guanosine allowed splicing . The first two substitutions place phosphorothioates inter alia at the 5' and 3' splice sites respectively . Mutagenesis at either site allowed phosphorothioate substitution 5' to guanosine at each splice site . This did not block splicing, suggesting that substitution at internal sites within the intron has more effect. Folia Microbiol (Praha), 1987, 32(4), 354 - 9 Bioconversion and binding of sterols by thermophilic moulds; Satyanarayana T et al.; None of the fourteen thermophilic moulds was able to break down the aliphatic side chain of sterols, viz . cholesterol, lanosterol, sitosterol, and stigmasterol so as to yield 4-androstene-3,17-dione, 1,4-androstadiene-3,17-dione and progesterone . In Acremonium alabamensis and Talaromyces emersonii, cholestenone was detected as a product of fermentation of cholesterol whereas the former yielded stigmastadienone from stigmasterol and sitosterol . Lanosterol appeared to be resistant to fungal bioconversion . All the thermophilic moulds exhibited avidity for binding sterols to the mycelium, but the ability to bind sterol seemed to depend upon the nature of the organism and the sterol. Antonie Van Leeuwenhoek, 1987, 53(2), 85 - 91 Lipid composition of thermophilic moulds Acremonium alabamensis and Thermomucor indicae-seudaticae; Satyanarayana T et al.; The total lipid content of Acremonium alabamensis and Thermomucor indicae-seudaticae ranged 2.6-7.3 and 8.5-13.0% of dry mycelium, respectively during development . Neutral lipid fraction increased during growth while polar and phospholipids declined . Both moulds contained palmitic, oleic, linoleic and palmitoleic acids as major fatty acid components in lipids . Degree of unsaturation of lipids of A . alabamensis was greater than that of T . indicae-seudaticae . Neutral lipids were more unsaturated than the polar lipids . The ratio of unsaturation index of polar lipids to neutral lipids was either one or less than one . The principal phospholipids of these moulds were phosphatidyl choline, phosphatidyl ethanolamine and phosphatidic acid . However, phosphatidic acid was not found in very high amounts as observed in Humicola grisea var . thermoidea. Development, 1987 Jan, 99(1), 51 - 68 Positional reorganization in compound janus cells of Tetrahymena thermophila; Frankel J et al.; The janus mutations of Tetrahymena thermophila convert the large-scale organization of the dorsal surface of the cell into