- Volume 86, Issue 1, 1975
Volume 86, Issue 1, 1975
- Biochemistry
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Chemistry and Ultrastructure of the Hyphal Walls of Pythium Acanthicum
More LessSUMMARY: Combined methylation, Smith degradation and enzymic studies show that the hyphal wall of Pythium consists predominantly of a (1,3)-linked β-d-glucan main chain substituted by a short chain at the 6-position of each third unit. Electron micrographs show that the hyphal wall has two different textures; the inner surface is distinctly microfibrillar while the outer surface is non-fibrillar. The microfibrils are covered with amorphous matrix material, partly soluble in aqueous KOH and almost completely removable by treatment with exo-laminaranase. This matrix material consists also of a branched β-d-glucan containing (1,3)- and (1,6)-linkages. The microfibrils remaining after (1,3)-glucanase treatment are composed of a β-glucan with (1,4)- and (1,3)-linked units that are occasionally substituted in the 6-positions.
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Ammonia Assimilation by Rhizobium Cultures and Bacteroids
More LessSUMMARY: The enzymes involved in the assimilation of ammonia by free-living cultures of Rhizobium spp. are glutamine synthetase (EC. 6.3.1.2), glutamate synthase (l-glutamine: 2-oxoglutarate amino transferase) and glutamate dehydrogenase (EC 1.4.1.4). Under conditions of ammonia or nitrate limitation in a chemostat the assimilation of ammonia by cultures of R. leguminosarum, R. trifolii and R. japonicum proceeded via glutamine synthetase and glutamate synthase. Under glucose limitation and with an excess of inorganic nitrogen, ammonia was assimilated via glutamate dehydrogenase, neither glutamine synthetase nor glutamate synthase activities being detected in extracts. The coenzyme specificity of glutamate synthase varied according to species, being linked to NADP for the fast-growing R. leguminosarum, R. melitoti, R. phaseoli and R. trifolii but to NAD for the slow-growing R. japonicum and R. lupini.
Glutamine synthetase, glutamate synthase and glutamate dehydrogenase activities were assayed in sonicated bacteroid preparations and in the nodule supernatants of Glycine max, Vicia faba, Pisum sativum, Lupinus luteus, Medicago sativa, Phaseolus coccineus and P. vulgaris nodules. All bacteroid preparations, except those from M. sativa and P. coccineus, contained glutamate synthase but substantial activities were found only in Glycine max and Lupinus luteus. The glutamine synthetase activities of bacteroids were low, although high activities were found in all the nodule supernatants. Glutamate dehydrogenase activity was present in all bacteroid samples examined. There was no evidence for the operation of the glutamine synthetase/glutamate synthase system in ammonia assimilation in root nodules, suggesting that ammonia produced by nitrogen fixation in the bacteroid is assimilated by enzymes of the plant system.
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Oligomeric Structure of Cholera Toxin: Characteristics of the H and L Subunits
More LessSUMMARY: Structural analysis of cholera toxin by sodium dodecylsulphate polyacrylamide electrophoresis demonstrated two types of non-covalently linked subunits, heavy (H) and light (L), with respective molecular weights of 28000 and 8000 to 9000. The H:L protein ratio was 1:2, indicating that the toxin of molecular weight 84000 consists of 1 H and 6 or 7 L subunits, linked into an aggregate with non-covalent bonds. Choleragenoid toxoid, a natural toxin derivative, contained only the L subunits of the toxin. Reduction and alkylation cleaved the H but not the L sub-unit. The specific cleavage of the H subunit by reduction appeared to yield identical half-molecules; the smaller peptide seemed to originate from non-specific degradation. The H subunit also differed from L subunits by having a higher affinity for labelling with radioactive iodine and by precipitating below pH 3·5.
In immunodiffusion studies the toxin possessed antigenic determinants shared with the toxoid as well as toxin-specific determinants. Comparative analyses with purified subunit preparations revealed that the toxoid-shared determinants reside in the L-type of subunit and the toxin-specific ones in the H subunit.
By precipitation-in-gel, binding to ganglioside-coated tubes, and sodium dodecyl sulphate polyacrylamide electrophoresis it was demonstrated that the ability of toxin to attach to the apparent receptor ganglioside, G m1, is similar to that of choleragenoid toxoid, and is due to the G m1 -binding ability of the L subunits. The toxin H subunit did not react with the G m1 ganglioside.
The results support our previous structural model for cholera toxin, and explain the antigenic and receptor-binding properties of the toxin in terms of component subunits.
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Biosynthesis of Lysine in Rhodotorula Glutinis: Role of Pipecolic Acid
More LessGlutamate-α-ketoadipate transaminase, saccharopine reductase, and saccharopine dehydrogenase activities were demonstrated in extracts of Rhodotorula glutinis but α-aminoadipate reductase activity could not be measured in whole cells or in extracts. Lysine auxotroph lys1 grew in the presence of l-lysine or dl-α-aminoadipate and incorporated radioactivity from dl-α-amino-[14C]adipate into lysine during growth. Growing wild-type cells converted l-[U-14C]lysine into α-amino-[14C]adipate, suggesting both biosynthetic and degradative roles for α-aminoadipate. Lysine auxotrophs lys1, lys2 and lys3 of R. glutinis, unlike lysine auxotrophs of Saccharomyces cerevisiae, satisfied their growth requirement with l-pipecolate. Moreover, extracts of wild-type R. glutinis catalysed the conversion of l-pipecolate to α-aminoadipate-δ-semialdehyde. These results suggest a bio-synthetic role for l-pipecolate in R. glutinis but not in S. cerevisiae.
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Isolation and Chemical Characterization of Plasma Membranes from the Yeast and Mycelial Forms of Candida Albicans
More LessSUMMARY: It has been possible to induce the yeast-mycelium transformation in Candida albicans by growth of the organism under completely defined conditions in batch culture. Protoplasts have been obtained from the two forms by using a lytic enzyme preparation from Streptomyces violaceus. A plasma membrane fraction was prepared by osmotic lysis of these protoplasts and fractionated by using a combination of differential and discontinuous sucrose density-gradient flotation centrifugation. The purity of this fraction was determined by radioactive dansylation and iodination of plasma membranes of intact protoplasts followed by localization of the radioactivity upon fractionation. This procedure demonstrated less than 4% contamination of the plasma membrane fraction with other cell membranes. Chemical analysis of this fraction revealed that the major components were protein and lipid. Membranes from the yeast form contained (w/w): 50% protein, 45% lipid, 9% carbohydrate and 0·3% nucleic acid. Plasma membranes from the mycelial form contained significantly more carbohydrate and were found to be composed of (w/w): 43% protein, 31% lipid, 25% carbohydrate and 0·5% nucleic acid. Marked differences were also observed between the phospholipid, free and esterified sterols, and total fatty acids of membranes from the two forms of the organism.
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- Development And Structure
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Changes in Morphology and Cell Wall Structure that Occur during Growth of Vibrio sp. NCTC4716 in Batch Culture
More LessSUMMARY: When grown in batch culture in various media Vibrio sp. nctc4716 displayed a distinct sequence of morphological forms. Organisms occurred as stout, almost straight rods in exponential phase, curved rods (characteristic of Vibrio spp.) in stationary phase, and predominantly as spheres in decline phase. The spheres were formed after growth had ceased due to the depletion of the carbon/energy source. They were not viable, survival of the culture depending on the few rod forms that remained during the decline phase. The spheres seemingly arose from degradation, but not complete removal, of the peptidoglycan present in the walls. Though spheres contained less nucleic acid and low molecular weight cytoplasmic constituents than did rods, many still possessed an intact cytoplasmic membrane.
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Ultrastructure of an Extremely Thermophilic Acidophilic Micro-organism
More LessA thermoacidophilic micro-organism, isolated from volcanic hot springs near Naples, was cultivated in vitro, and examined by electron microscopy in sections and after negative staining. The cells were almost spherical, with a diameter of about 0·7 to 1·0 μm. Their morphology was very primitive: the protoplasm was composed only of ground cytoplasm, ribosomes, and randomly distributed DNA strands. They were surrounded by a plasma membrane and by an extracellular coat about 20 nm thick which displayed a regular hexagonal pattern. Cell replication occurred by binary fission with median constriction during which a bipolar localization of nuclear material was observable. The morphology is compared with that of other known micro-organisms living in similar habitats.
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- Genetics And Molecular Biology
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R Factors from Serratia Marcescens
More LessSUMMARY: R factors transmissible to Escherichia coli from naturally occurring strains of Serratia marcescens belonged either to compatibility groups (S and L) not represented amongst plasmids reported in other genera or to groups (C, FII, P and M) notable for their wide host range.
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Thermosensitive Production of Their Transfer Systems by Group S Plasmids
More LessTransfer of plasmids of group S is much more efficient at low temperatures (e.g. 22 °C) than at 37 °C. This is due to failure of the donor strain to produce the transfer system during growth at the higher temperature.
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Chromosome Transfer in Proteus Mirabilis Mediated by a Hybrid Plasmid
More LessSUMMARY: A previously-described fused plasmid, P-lacR1drd19, was found to mediate chromosomal transfer between cells of Proteus mirabilis strain pm5006; pm5006-(P-lacR1drd19) was usually the donor and various auxotrophs of pm5006 resistant to nalidixic acid and/or streptomycin were recipients. The donor was usually counterselected with nalidixic acid and/or high concentrations of streptomycin. Recombination experiments with single markers indicated a 40-fold variation in recombination frequencies for different markers. Mapping double-auxotrophic markers by their gradient of transmission confirmed this variation and placed each of two independent isolates of eight markers in a linkage group his-ser-ura-pyrB-trp-cys-ade-ilv. Some donor markers did not register. Despite low recombination frequencies, interrupted mating experiments showed a polarity of early marker transfer. The segregation of unselected markers confirmed the order of some markers and showed that genetic material passed from the presumptive donor to the recipient. Recipients with two auxotrophic markers which could not be co-transduced by phage 5006M were converted to prototrophy by conjugation. The plasmid transferred to recipients at high frequency and all recombinants carried it. Recombinants could act as donors in further matings. Recombinants were fully susceptible to phage 5006M, unlike transductants of pm5006 by this phage. Direct involvement of the plasmid was indicated by drastically diminished recombination frequencies in crosses with recipients carrying P-lac as resident. P-lac had previously been shown to reduce the frequency of transfer of the hybrid plasmid to cells harbouring it. The histidine region was the first to register in recipients and recombined at the highest frequency of 5 × 10−6/donor cell. Some temporary association of plasmid and perhaps only the histidine region of the chromosome is favoured as the mechanism of chromosomal transfer. This could explain why not all donor markers could be mapped. Transduction and transformation were excluded as the cause of results.
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Mutations which Affect Amino Acid Transport in Aspergillus Nidulans
More LessMutants deficient in amino acid transport (aau), and unable to utilize l-glutamate as a sole carbon and nitrogen source, have been isolated. There are four unlinked genes involved: aauA, aauB, aauC and aauD. The transport levels of certain amino acids, and the growth characteristics on certain nitrogen and carbon sources and toxic amino acid analogues, indicate that: aauAI has defective transport of acidic amino acids; aauBI has defective transport of acidic and neutral amino acids; aauCI and aauDI have defective transport of acid, neutral and aromatic amino acids. aauAI and aauBI are recessive for all three characteristics in the heterozygous diploid; aauCI and aauDI are dominant.
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- Physiology And Growth
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Sub-cellular Location of Mercury in Yeast Grown in the Presence of Mercuric Chloride
More LessSUMMARY: The distribution of 203Hg in Saccharomyces cerevisiae grown in the presence of mercuric chloride has been examined by physical and chemical fractionation procedures and autoradiography. The major fraction of the bound mercury is tightly bound to the wall. A significant quantity of mercury penetrates to the cytoplasm but only a minor fraction is present as low molecular weight components. The wall-associated mercury is not readily released by extraction with sodium hydroxide or ethylenediamine but a major fraction is solubilized by Pronase and Helicase treatment. Isolated walls are capable of binding their own weight of mercury to high-affinity adsorption sites. The major role of the cell envelope in the in vivo binding of mercury and the penetration to the cytoplasm of mercury was confirmed by autoradiography.
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Glucose Repression of Enterotoxins A, B and C and Other Extracellular Proteins in Staphylococci in Batch and Continuous Culture
More LessSUMMARY: The production of enterotoxins, lipase and total extracellular protein by four strains of Staphylococcus aureus grown in batch culture at a controlled pH of 6·5 in a completely defined medium was markedly reduced by glucose or glycerol constantly maintained at 0·1m. A concomitant increase in the production of deoxy-ribonuclease, up to 13-fold, showed however that not all extracellular proteins are under the same control mechanism. The presence of glucose and glycerol in the medium also resulted in a rapid increase in the specific growth rate. However, growth of S. aureus s6 in Mg++-limited continuous culture showed that glucose repression of enterotoxin B when the growth rate was held constant was more than twice that in batch culture. Therefore glucose repression can occur independently of an increase in growth rate. The specific rate of production of enterotoxin B, lipase, deoxyribonuclease, β-haemolysin and total extracellular protein by S. aureus s6 increased as the growth rate increased from 0·07 to 0·24 h−1. Non-replicating cells grown in the absence of glucose produced considerable amounts of enterotoxin, and production was not repressed by the presence of glucose in the re-suspension medium. In contrast, no enterotoxin B or C was obtained from non-replicating cells grown in the presence of glucose. Chloramphenicol completely inhibited enterotoxin production by non-replicating cells, indicating that synthesis of new protein was required.
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The Effect of Coenzyme Leakage and Replacement on the Growth and Metabolism of Two Fusobacteria
More LessSUMMARY:Suspensions in water of two species of Fusobacterium leaked several coenzymes when incubated at normal growth temperatures. Chromatography of filtrates from these suspensions revealed the presence of NAD, NADP, FMN, tetrahydrofolic acid and, in one of the two, pyridoxal phosphate. Analyses of some enzymic activities in whole organisms demonstrated deficiencies in coenzymes: glutamate dehydro-genase was virtually inactive in the absence of added NAD; tryptophanase activities were diminished by washing but the extent differed between strains; histidase activity was not decreased by washing or suspension in water or saline. Both lag phase and doubling time increased markedly in severely washed organisms inoculated into fresh medium. Addition of appropriate coenzymes shortened the lag phase for both strains and shortened the doubling time in one.
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- Short Communications
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- Taxonomy
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Torulopsis Bacarum, Torulopsis Pustula and Torulopsis Multis-Gemmis sp. nov., Three New Yeasts from Soft Fruit
More LessThree new species of yeasts isolated from fresh soft fruit are described. Torulopsis pustula was isolated from blackcurrants, Torulopsis bacarum from blackcurrants, strawberries and raspberries, and Torulopsis multis-gemmis from raspberries. All differ from known species in the test substrates they utilize. In addition, cells of Torulopsis pustula often form a bulbous projection near their apices from which budding takes place. These buds are frequently pear- or heart-shaped.
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A Maximal Predictive Classification of Klebsielleae and of the Yeasts
More LessThe concepts of the numerical method of maximal predictive classification are illustrated with classifications of 13 species of enterobacteria and of 434 species of yeast. The method seeks to classify into a specified number of classes (k) such that more correct statements can be made about the constituent members than with any other classification. The best choice of k relates to the separation of the classes as measured by the average number of correct statements made for an individual assigned to a class to which it does not belong. The maximal predictive classifications are compared with previous classifications of the two groups, which seem to be poor predictively (in terms of the characters considered in this study). The results suggest that taxonomists may be more concerned with maximizing class separation rather than with prediction, but many more groups of organisms would need similar study before this view could be held with confidence.
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Extremely Thermophilic Acidophilic Bacteria Convergent with Sulfolobus Acidocaldarius
More LessSUMMARY: A series of extremely thermophilic acidophilic bacteria has been characterized as closely resembling the species Sulfolobus acidocaldarius except for a totally different guanosine-cytosine content in the DNA; some conceptual consequences of this situation are discussed. Both organisms also share special features, including a very characteristic type of ether lipid, with other extreme acidophilic thermophiles.
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- Corrigendum
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Volumes and issues
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Volume 170 (2024)
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