- Volume 100, Issue 1, 1977

SUMMARY: Choline, a component of the wall teichoic acid of Streptococcus pneumoniae, was converted to cytidine diphosphocholine via choline phosphate by enzymes which were identified in cell-free extracts of the pneumococcus. The first enzyme, choline kinase, was investigated in some detail. It appeared to have a pH optimum of 7·3 to 7·4 and was stimulated by Mg2+. Kinetic studies gave an apparent Michaelis constant (K m) for ATP of 1 mm, and for choline of 0·19 mm, with V max values of 3 nmol min−1 (mg protein)−1 and 0·5 nmol min−1 (mg protein)−1 respectively. The second enzyme, CDPcholine pyrophosphorylase was specific for CTP and had a requirement for Mg2+ with an optimum at 7 mm.

SUMMARY: A membrane-bound nitrate reductase from Thiobacillus denitrificans can utilize either sulphite or NADH as an electron donor. The sulphite-dependent nitrate reductase activity was released from the membrane by treatment with sodium deoxycholate. Cytochrome c and FAD were separated from the solubilized enzyme by heat treatment and subsequent chromatography on DEAE-cellulose. The bacterial cytochrome c and a preparation of horse-heart cytochrome c served as electron mediators to the solubilized sulphite-dependent nitrate reductase activity with apparent K m values of 1·5 and 1·3 μ m respectively. The NADH-linked enzymic activity, which was unstable during storage, was re-activated with reduced glutathione. It was also inactivated after treatment with deoxycholate but this effect was reversed by menadione. A possible scheme for electron transport for the sulphite- and NADH-dependent enzyme is proposed.

SUMMARY: Glucose-6-phosphate-dependent glycogen synthase from Dictyostelium discoideum was purified and characterized. Several nucleoside phosphates inhibited enzyme activity at their physiological concentrations (about 10−4 m). Glucose 6-phosphate and UDPglucose released these inhibitions at levels above their physiological concentrations. Unless compartmentation is invoked, experiments suggest that the glucose-6-phosphate-dependent enzyme is inactive in vivo.

SUMMARY: In Lactobacillus casei S-1, d-galactosamine and l-rhamnose comprised a phage receptor for phage J-1. A mixture of d-galactosamine and l-rhamnose effectively inactivated phage J-1, and a J-1-resistant mutant strain, L. casei S-1/J-1, lacked d-galactosamine in its surface component. The phage-inactivating effects of d-galactosamine and l-rhamnose were strongly dependent on the concentration of each substance and on temperature.

It is suggested that the receptor for phage J-1 involves both d-galactosamine in the cytoplasmic membrane and l-rhamnose in the wall of the host bacterium L. casei s-1, which lacks teichoic acid in its wall.

SUMMARY: A partial order for genes which specify meta-cleavage pathway enzymes has been derived from properties of a polarity mutant strain (PsU5) of Pseudomonas putida ncib10015 and a partial revertant, PsU5/R21. The polar mutation is within the 2-hydroxymuconic semialdehyde dehydrogenase gene and results in loss of detectable 2-hydroxymuconic semialdehyde hydrolase and 90 to 95 % reduction in the activities of 4-oxalocrotonate tautomerase, 4-oxalocrotonate decarboxylase, 2-oxopent-4-enoate hydratase and 4-hydroxy-2-oxovalerate aldolase. The partial revertant PsU5/R21 regained wild-type levels of all enzymes except the aldehyde dehydrogenase.

It is probable that all genes determining the dehydrogenase and subsequent enzymes are transcribed as a polycistronic message. This is the first report of mapping genes determining the enzymes of the meta-cleavage pathway.

SUMMARY: In Pseudomonas putida ncib10015 the gene determining phenol hydroxylase is in a separate operon from the genes determining catechol 2,3-oxygenase and subsequent enzymes of the meta-cleavage pathway.

In the wild-type strain, non-coordinate induction of phenol hydroxylase and catechol 2,3-oxygenase occurred when catechol, phenol or isomers of cresol were used as inducers. Phenol hydroxylase was repressed to a greater extent than catechol 2,3-oxygenase when induced by phenol in the presence of either benzoate or acetate. In a mutant deficient in catechol 2,3-oxygenase, 3- or 4-methylcatechol repressed formation of phenol hydroxylase but not of enzymes functioning later in the pathway.

Four mutants had an altered regulatory element that resulted in different effects on the expression of phenol hydroxylase and catechol 2,3-oxygenase genes when isomers of cresol were used as inducers.

Repression of enzyme induction by acetate, benzoate or 3- or 4-methylcatechol showed that catechol 2,3-oxygenase and subsequent meta-cleavage pathway genes form part of the same operon. Benzoate or 3- or 4-methylcatechol repressed phenol hydroxylase but none of the other enzymes in the pathway, while acetate coordinately repressed catechol 2,3-oxygenase and subsequent enzymes.

SUMMARY: Mutants of Pseudomonas putida ncib10015 were isolated which produced catechol 2,3-oxygenase and the subsequent enzymes of the meta-cleavage pathway constitutively, and were defective in phenol hydroxylase activity. All revertants of one mutant (C3), and most revertants of a second mutant (C1), regained wild-type characteristics with respect to inducibility of phenol hydroxylase and the other meta-cleavage enzymes. Their behaviour was consistent with them being regulatory mutants.

Other mutants deficient in phenol hydroxylase activity were not constitutive for catechol 2,3-oxygenase and other enzymes of the pathway. Their properties were consistent with mutations in a structural gene.

A model in which phenol hydroxylase is under positive control and catechol 2,3-oxygenase and subsequent enzymes are under negative control is proposed for the regulation of enzymes of the meta-cleavage pathway in P. putida.

SUMMARY: Arginine hydroxamate inhibits the growth of Bacillus subtilis. From a large number of mutants isolated as resistant to this arginine analogue, 29 were chosen for further investigation. Most of these shared diminished ability to utilize arginine, citrulline and/or ornithine as sole nitrogen source. All 29 had reduced levels of the catabolic enzymes arginase and ornithine aminotransferase under various conditions in which these enzymes are induced in the parent. In some circumstances, five of the mutants also showed elevated levels of the biosynthetic enzyme ornithine carbamoyltransferase. On the basis of these data, the 29 mutants were divided into six phenotypic classes; in four of these, control of ornithine carbamoyltransferase was the same as in the wild type, while in the other two it was altered. It is suggested that the isolates carry regulatory mutations, and that certain of these may affect simultaneously the formation of arginine catabolic and biosynthetic enzymes. The implication of the latter is that in B. subtilis, as in yeast, controls of the catabolic and biosynthetic pathways are connected.

Single representatives of five of the phenotypic classes carry mutations conferring arginine hydroxamate resistance linked to cysA by transduction with phage PBS1; this did not appear to be true for a representative of the sixth class.

SUMMARY: Non-conjugative Tc, Sm, SmSu, SmSpc, SmSpcSu and TcSmSpcSu determinants (Tc, Sm, Spc and Su denote resistance to tetracycline, streptomycin, spectinomycin and sulphonamides, respectively) in wild-type Escherichia coli strains were mobilized with transfer factors F, I and A2 and implanted in E. coli k12. F and I were transmitted at very high rates in all matings in which these E. coli k12 strains were used as donors; the rate of A2 transmission was not measured. When the Tc determinants, derived from 32 of the wild-type E. coli strains, were serially transferred between strains of E. coli k12 they were transmitted either at a very low rate in both first and second matings, at a very low rate in the first mating and at a very high rate in the second mating, or at a very high rate in both matings. In the donors that transmitted at the low rate, the Tc determinants were probably chromosomally located, a recombination event between them and the transfer factors being responsible for the formation of the donors that transmitted at the high rate; once they became extrachromosomally located, the Tc determinants continued to be transmitted at the high rate. Of the resistance determinants studied, Tc was the only one suspected of being chromosomally located.

F transmitted most of the four Sm and eight SmSpc determinants at high rates and the 14 SmSu determinants at much lower rates. I and A2 transmitted most of the three kinds of determinants at high rates but four of the E. coli k12 SmSpc+A2+ strains transmitted them at very low rates and two E. coli k12 SmSpc+I+ strains and one E. coli k12 SmSu+I+ strain did not transmit them at all. F and I did not establish linkage with Sm or SmSu but linkage was detected in the strains that transmitted at the high rate between them and SmSpc. The one TcSmSpcSu determinant studied was transmitted at two moderately low rates by F with which it was not linked. In the first matings following implantation in E. coli k12 by I or A2, it was transmitted at a very low rate, but in most subsequent matings its transmission rate was high; for the I+ strain this was certainly due to transfer factor linkage being established.

SUMMARY: Infection of subcutaneously implanted chambers in guinea pigs conferred immunity against homologous infection of other chambers in the same animals. However, attempts to immunize guinea pigs by subcutaneous injection of filtered fluid from infected chambers, or with small doses of formalin-killed, chamber gonococci were not successful. Thus, neither organisms grown in vivo nor their extracellular products appeared to be exceptionally immunogenic.

In immunizing tests with different isolates of gonococci adapted to growth in guinea-pig chambers, cross-immunity to chamber infection with low challenge doses was detected only between two of six isolates. The killing of gonococci in chambers of immunized animals, which occurred only after homologous challenge or with the heterologous strain showing cross-immunity, was not due primarily to humoral factors in the chamber fluid but probably to an enhanced effectiveness of phagocytosis. The serum of immunized animals was bactericidal for homologous strains and for the strain showing cross-immunity but not for strains showing no cross-immunity. Hence, serum bactericidal activity might be a useful indicator for investigating the specificity of immunity produced by different gonococcal strains.

SUMMARY: Carbon dioxide at a concentration of 1 to 2% (v/v) in air enhanced the growth rate and inhibited sclerotium formation in the fungus Sclerotium rolfsii Sacc. A CO2 concentration of 10% inhibited growth. Similar growth patterns were observed when the fungus was grown on a medium supplemented with the fungicide carboxin, which inhibits succinate dehydrogenase. A high CO2 concentration (1 to 10%) or growth on carboxin-supplemented medium caused a decrease in succinate dehydrogenase activity and significant increases in isocitrate lyase, isocitrate dehydrogenase, malate synthase and malate dehydrogenase activities. Mycelium of S. rolfsii grown at a high CO2 concentration contained less glyoxylate, lipids and glycogen than mycelium grown in air. It is suggested that sclerotium formation in S. rolfsii requires a balanced supply of carbohydrate intermediates and energy.

SUMMARY: The aerobic nocardioform actinomycete, found to be common in herbivore dung and aquatic habitats and previously given the trivial name Lspi, was studied along with representative strains of taxa within the ‘rhodochrous’ complex. Lspi strains were recovered as a homogeneous cluster which separated from the other reference strains at the 69% similarity level in a numerical taxonomic analysis. The specific name Rhodococcus coprophilus is proposed for this new species. We also consider the generic name Rhodococcus Zopf to be appropriate for most of the species currently accommodated within that rather ill-defined taxon known as the ‘rhodochrous’ complex and variously classified within the genera Nocardia, Mycobacterium, Jensenia, Corynebacterium or Gordona.