- Volume 108, Issue 2, 1978
Volume 108, Issue 2, 1978
- Biochemistry
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Inhibition by Mercurial Reagents and Role of SH Groups of the Adenosine Triphosphatase from Escherichia coli 414 Membranes
More LessMercurials selectively inhibited Escherichia coli 414 ATPase (EC 3.6.1.3). Inhibition of the soluble ATPase (BF1) was greater than for the membrane-bound enzyme. The titration of 4 SH groups per mol BF1 with 0.05 mm-p-chloromercuri[14C]benzoate showed a good dose-response curve for the inhibition of basal ATPase but not for the trypsin-stimulated activity. Accessible SH groups did not seem to be related to the active site of the enzyme. Mercurials appeared to affect E. coli ATPase by inducing a molecular change in the holo-enzyme, followed by dissociation. One to two SH groups with different degrees of accessibility were located in the α and γ subunits of ATPase (BF1) but only one was located in the β subunit, irrespective of the concentration of p-chloromercuribenzoic acid, suggesting a structural role for SH groups in BF1.
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Nature of the Nets Produced by Protoplasts of Schizosaccharomyces pombe During the First Stage of Wall Regeneration in Liquid Media
More LessThe microfibrillar nets formed during the first stage of wall regeneration by protoplasts of Schizosaccharomyces pombe in a liquid medium contain, in contrast to normal walls, (1→3)-β-linked glucan of high crystallinity, like the nets produced by protoplasts of Saccharomyces cerevisiae under the same conditions. During wall regeneration by protoplasts of Schizosaccharomyces, β-glucan synthesis precedes the formation of (1→3)-α-glucan.
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- Development And Structure
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The Surface Properties of Neisseria gonorrhoeae
More LessGonococci were labelled with 125I using the lactoperoxidase system. The amount of label incorporated was similar with all strains including those which appeared capsulated. Electro-phoresis on sodium dodecyl sulphate–polyacrylamide gels revealed that the major proteins labelled were those found in outer membrane preparations. Comparison of variants of one strain showed that the major outer membrane protein (protein I) was always present and heavily labelled. The second major protein (protein II) was present in variable amounts but labelling was proportional to the amount present. A third protein (III) was only present in outer membranes from a freshly isolated variant but was present in whole cells of each strain. Protein III was not labelled in whole cells but was labelled in outer membrane preparations suggesting that many membranes have their inner surface exposed. The labelling of a strain adapted to growth in guinea-pig chambers failed to reveal any new major surface proteins. The results demonstrate the variation in surface topography possible with variants of one strain of gonococcus but show that one major protein antigen is always expressed on the surface.
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An Electron Microscopic Study of the Location of Peptidoglycan in Group A and C Streptococcal Cell Walls
More LessThe morphological appearance of deproteinized Group A and C streptococcal walls after treatment by different procedures extracting teichoic acids and polysaccharides (formamide, hydrochloric acid, nitrous acid, trichloroacetic acid, sulphuric acid, sodium hydroxide and sodium deoxycholate) was compared with the content of teichoic acids and polysaccharides remaining in the treated walls. All procedures extracted teichoic acids almost completely, but polysaccharides were extracted to various degrees. The ultrastructural appearance of walls after these extractions still exhibited the triple-layered wall profile; only a reduction of thickness of the wall and of electron density of the layers occurred. There was no direct correlation between the reduction of rhamnose content and thickness of walls.
The ultrastructural localization of peptidoglycan in the streptococcal walls was explored by means of the indirect immunoferritin technique using anti-peptidoglycan antibodies isolated from anti-Group A-variant antisera. Ferritin particles were bound predominantly to filamentous structures which protruded from both surfaces of peptidoglycan fragments and isolated walls. Peptidoglycan was also detected on the filamentous protrusions of whole cocci. These results contradict models of the streptococcal wall in which peptidoglycan forms the innermost layer and support a mosaic structure in which peptidoglycan forms a network of the peptidoglycan-polysaccharide complex.
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- Genetics And Molecular Biology
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Bacterial Donor Mutants Affecting the Efficiency of Generalized Transduction by Salmonella Phage P22
More LessP22 lysates of some mutants (LD mutants) of Salmonella typhimurium gave lower transduction frequencies than phages grown on the wild type. The lower frequency was not due to the production of reduced numbers of transducing particles, or superinfection exclusion, or decreased incidence of integration. The defect was in a host-controlled modification-restriction system. LD mutants possess m – r – characteristics. The degree of expression of transducing particles varied with the phenotype selected, although some gene products, including the numbers of plaque-forming phages of P22, were not affected. However, several other Salmonella phages, some of them related to P22, exhibited high sensitivities. The LD system appeared to be identical to Colson’s SA system ( Colson & van Pel, 1974 ).
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Mutants with Increased Sensitivity to Canavanine in the Fungus Coniochaeta velutina
More LessMutants of Coniochaeta velutina with increased sensitivity to canavanine were derived from wild-type, pyr-3 (presumably defective in carbamoyl-phosphate synthase for pyrimidine synthesis), arg (blocked in steps prior to arginine formation) and pro (blocked in steps prior to glutamate semialdehyde formation) strains, and were classified into eight distinct groups by genetic mapping. Four canavanine-sensitive genes (cns) were allelic or very tightly linked to known arg loci. The pairs of genes involved were cns A and arg-3 (arginine requirer), cnsB and arg-7 (citrulline requirer), cnsC and arg-2 (citrulline requirer) and cnsG and arg-1 (ornithine requirer). Another (cnsF) was mapped in linkage group III.
A relationship was shown between the level of intracellular arginine and the extent of canavanine sensitivity; cns strains contain less intracellular arginine than the less sensitive wild-type. In most cases, this was shown to result from mutations affecting the enzyme(s) involved in arginine metabolism. cnsA and cnsG were thought to be affected in arginine- and ornithine-synthesizing enzymes, respectively. cnsB and cnsC exhibited a requirement for citrulline when the two genes were combined and were suggested to be loci specifying the two polypeptide chains of carbamoyl-phosphate synthase for arginine synthesis. cnsH was the locus controlling ornithine carbamoyltransferase. The metabolic lesions of the other cns mutants (cnsD, cnsE and cnsF) were unclear.
Of the eight cns mutants, six could suppress pyr-3M and/or pro mutations: cnsG was able to suppress pyr-3M; cnsB, cnsE, cnsD and cnsF were effective in suppressing pro; cnsH suppressed both pyr and pro. The mechanism of the pyr and pro suppressions was explained on the basis of metabolic cross-feeding.
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Genetical and Biochemical Studies of Glucosephosphate Isomerase Deficient Mutants in Saccharomyces cerevisiae
More LessA number of glucose-negative mutants of Saccharomyces cerevisiae were isolated and shown to contain very low activities of glucosephosphate isomerase. Mutants almost totally lacking this enzyme (less than 1% of wild-type activity) grew on fructose if provided with a small quantity of glucose. Larger amounts of glucose led to the accumulation of glucose 6-phosphate and growth inhibition. These mutants did not grow on galactose. Other mutants with low enzyme activities (about 1% of wild type) grew on fructose alone and also on galactose. The mutant characters were determined in both cases by single gene mutations which were mapped on chromosome II and, presumably, identify a structural gene locus for the enzyme.
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- Medical Microbiology
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The Surface Properties of Neisseria gonorrhoeae
More LessMonovalent rabbit antisera were prepared to highly purified gonococcal lipopolysaccharide (LPS), to pili and to two major purified outer envelope proteins. All these antisera were free from significant specific IgM antibody and were standardized to 4 μg specific IgG antibody per test, permitting accurate comparisons between the different gonococcal surface antigens as triggers of the complement-dependent bactericidal reaction. LPS was the most effective antigen at inducing a bactericidal response to homologous and heterologous gonococci, followed by the two individual outer envelope proteins. Pili were relatively ineffective. Strain p9 gonococci grown in vivo or which possessed a ‘capsule’ in vitro were more resistant to serum killing than the non-capsulated parent strain. One highly susceptible strain, f62, which was killed by complement in the absence of any LPS antibody, was able to directly activate complement by the alternative pathway.
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- Physiology And Growth
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Bud Formation and the Inducibility of Pseudo-mycelium Outgrowth During Release from Stationary Phase in Candida albicans
More LessThe kinetics of bud formation at 25 °C and germ tube formation at 37 °C were examined in populations of Candida albicans released from stationary phase by dilution into fresh nutrient medium. Evidence is presented for four separate isolates and three different defined media that: (i) cells do not deplete the media of growth-limiting nutrients when they enter stationary phase; (ii) cells accumulate as singlets when they enter stationary phase, presumably at a point early in the cell cycle; (iii) daughter cells do not separate from mother cells during the first three to four cell divisions following release from stationary phase at 25 °C; and (iv) released cells cannot be induced to form germ tubes by an increase in temperature once they have formed their first bud at 25 °C. The relationships between the lack of cell separation, the inducibility of tube formation and stationary phase are discussed.
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The Relationship Between DNA Replication and the Induction of Sporulation in Bacillus subtilis
More Less6-(p-Hydroxyphenylazo)uracil (HPUra), which is thought to be a specific inhibitor of DNA replication in many Gram-positive bacteria, has been used to investigate the relationship between DNA replication and the induction of sporulation in Bacillus subtilis. Gene frequency analysis of samples removed from sporulating cultures indicates that sporulating cells terminate their final round of DNA replication at the same time as they escape from the inhibitory effect of HPUra. If the rate of DNA replication during starvation is slowed down by the use of sub-inhibitory concentrations of HPUra, the onset of this escape is delayed. When cells containing only completed chromosomes are transferred from a rich growth medium to a poor sporulation medium they initiate new rounds of DNA replication in the sporulation medium and then subsequently sporulate. The results are consistent with the hypothesis of Mandelstam & Higgs (1974) that induction of sporulation occurs 15 to 20 min after the initiation of a round of DNA replication.
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Oscillations of Respiration and Adenine Nucleotides in Synchronous Cultures of Acanthamoeba castellanii
More LessSynchronous cultures of the soil amoeba Acanthamoeba castellanii, prepared by a size selection procedure involving minimum metabolic perturbation, divided with a high degree of synchrony and showed a discrete S-phase of DNA synthesis. Oxygen uptake rates doubled overall during one cell cycle time of 7 to 8 h but rose to seven distinct maxima during this period. Control experiments showed that the selection procedure did not give rise to the oscillations. The maxima of respiration were more sensitive to inhibition by cyanide than were the minima of respiration; the effect of carbonyl cyanide p-trifluoro-methoxyphenylhydrazone (an uncoupler of oxidative phosphorylation) was, however, greater at respiratory minima, than at the maxima. Pool levels of ATP, ADP and AMP also oscillated during the cell cycle with maximum amplitudes (peak-trough, % minimal values) of 108, 194 and 520, respectively. Adenylate charge values varied between 0·63 and 0·88. Respiratory maxima were in phase with maximum ADP levels but out of phase with maxima of ATP/ADP ratios. These results suggest that the overall changes in respiration rates during the cell cycle of A. castellanii are the result of in vivo respiratory control.
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Cyanide Production and Degradation During Growth of Chromobacterium violaceum
More LessCyanogenesis by growing cultures of Chromobacterium violaceum was stimulated by the inclusion of glycine and methionine in the growth medium. Increases in the ferrous ion and phosphate concentrations of the growth medium stimulated cyanide production. Chromobacterium violaceum possesses a number of cyanide-utilizing enzymes: β-cyanoalanine synthase, γ-cyano-α-aminobutyric acid synthase and rhodanese. Studies on the activities of these enzymes in cell-free extracts of cultures growing under both high and low cyanide-evolving conditions are presented. Addition of chloramphenicol to high and low cyanide-evolving cultures towards the end of exponential growth had a profound effect on the medium cyanide concentrations. These observations are shown to have been caused by chloramphenicol blocking the induction of the cyanide-utilizing enzymes.
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The Effects of Cooperativity and Growth Yield Variation on the Kinetics of Nitrogen or Phosphate Limited Growth of Chlorella in a Chemostat Culture
More LessAccounting for a cooperativity effect in substrate uptake by a microbial culture leads to a Hill type of equation for the specific rate of substrate uptake (Q), that is,
where S is substrate concentration, n is the Hill number and K sh is a constant. For substrates such as N or P, which are conserved in the biomass, the substrate content of the biomass (α) will vary according to the relation
where α 0 is the minimum substrate content of the biomass, occurring when S → 0, and α m is the maximum substrate content of the biomass, occurring when the biomass is saturated with substrate. The specific growth rate is given by
where L is a constant given by α 0/α m. If the substrate is conserved, α 0 = 1/Y 0 and α m = 1/Y m where Y 0 and Y m are the maximum and minimum yields from the substrate, respectively.
Experimental tests of these relations applied to N (urea)-limited growth of Chlorella vulgaris in chemostat cultures showed satisfactory agreement between the results and the theory. For N uptake the growth constants were n = 5·5, L = 0·47, K s = (LK sh)1/n = 82 μg N l–1, = K sh 1/n = 94 μg N l–1. The apparent departure of L from unity in N-limited growth can be accounted for by starch storage in the biomass.
For P (phosphate)-limited growth, n = 1·3, L = 0·15 and K s was about 28 μg P l–1. A discrepancy was found between the K sh value for P uptake obtained from the Q data and that from the α data. This discrepancy may be attributed to phosphate storage in the biomass, which is not allowed for in the model.
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- Short Communications
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- Taxonomy
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Genetic Relationships among Bacterial Endosymbionts of Paramecium aurelia
More LessGenetic relationships among bacterial endosymbionts of Paramecium assigned to the genus Caedobacter Preer et al. 1974 were determined using DNA-DNA hybridization techniques. It was shown that mu, nu and pi endosymbionts are closely related to each other, but not to any of the strains of kappa endosymbionts tested, and that kappa consists of at least three distinct and genetically diverse groups. Therefore, it is proposed that Caedobacter be split into two genera, Caedobacter and Pseudocaedobacter, depending upon the ability or lack of ability, respectively, to produce cells containing R bodies. Strains were screened for covalently closed, circular DNA by ethidium bromide/CsCl centrifugation, but only pi and hump-killer kappa particles were found to contain this form of DNA.
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Some Characteristics of Ureaplasma urealyticum. Urease Activity in a Simple Buffer
More LessUrealytic activity of the cytoplasmic fraction of Ureaplasma urealyticum prepared by digitonin lysis was assayed in a simple buffer system (HEPES plus EDTA) by measuring the release of 14CO2 from [14C]urea. The K m of this preparation agreed with our previous observations of the same activity measured in a more complex reaction mixture. The substrate concentration at which maximum velocity occurred was approximately 20 mm. The activity was sensitive to heavy metals and inhibitors which react with sulphydryl groups such as N-ethylmaleimide and p-chloromercuribenzoate. It was not inhibited by Ca2+ or Mg2+ or by the reaction products, ammonia and carbon dioxide.
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Characterization of Serratia marcescens, S. liquefaciens, S. plymuthica and S. marinorubra by the Electrophoretic Patterns of their Esterases
More LessEsterases of 62 Serratia marcescens, S. liquefaciens, S. plymuthica and S. marinorubra strains were analysed by horizontal slab electrophoresis in polyacrylamide-agarose gel. Five principal bands hydrolysed β-naphthyl acetate but differed in their range of activity towards other substrates and in their sensitivity to di-isofluoropropyl phosphate. Additional bands were detected in some strains of S. marcescens and S. marinorubra. The comparative distribution of bands showed that the four Serratia species were characterized by distinct electrophoretic patterns of their esterases. Serratia marcescens, S. liquefaciens and S. plymuthica appeared to be more closely related to one another than to S. marinorubra. In the case of S. plymuthica and S. marinorubra, congruence was found between biovars and esterase patterns. The numerous electrophoretypes observed within Serratia species might provide useful epidemiological markers.
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- Corrigendum
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Volumes and issues
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Volume 170 (2024)
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