- Volume 97, Issue 2, 1976

Summary: Crude chitin synthetase preparations from the mycelial and yeast forms of Mucor rouxii behaved differently. The mycelial preparations, incubated at 28 °C, lost virtually all chitin synthetase activity in a few hours; by contrast, the activity of enzyme preparations from yeast cells increased several fold during similar incubations. These spontaneous changes were probably caused by endogenous protease(s). Seemingly, the chitin synthetase in yeast preparations was present mainly in a latent, ‘zymogenic’, form that was activated by proteases. In the mycelial preparations, chitin synthetase was present mainly in an active state and was rapidly degraded by endogenous proteolysis. Exogenous proteases accelerated activation and destruction of chitin synthetase; an acid protease from Rhizopus chinensis was the most effective activator. The activation of chitin synthetase was inhibited by a soluble protein in the cell-free extract. Treatment with the detergent Brij 36T stabilized the chitin synthetase of crude preparations against spontaneous changes. Stabilized preparations were rapidly activated by exogenous proteases. The different behaviour of chitin synthetases in crude extracts of mycelium and yeast cells is consistent with, and perhaps partially responsible for, the differences in wall construction between mycelial and yeast forms of M. rouxii.

Summary: During growth in a liquid culture containing a single soluble or an insoluble cellulosic carbon source, Rhizoctonia lamellifera released cellulolytic enzymes into the medium. These enzymes were separated by gel filtration and ion-exchange chromatography into seven components, three of high and four of low molecular weight. One of the components had the character of a C1 cellulase. When the components were combined they released more reducing sugars from cellulosic substrates than when used singly.

Summary: Escherichia coli k12 cannot grow on d-lyxose, but a mutant was isolated which can utilize d-lyxose as sole source of carbon and energy for growth. d-Lyxose is transported into the bacteria by the d-xylose permease. The mutant constitutively synthesizes a new isomerase which is not inducible in the parent strain under any of the conditions tested. This enzyme, whose native substrate appears to be d-man-nose, fortuitously converts d-lyxose into d-xylulose. Its structural gene is located at around 85 min on the E. coli genetic map, away from other known isomerase genes. d-Xylulose is subsequently catabolized by the enzymes of the normal d-xylose metabolic pathway. d-Mannose isomerase was partially purified and some of its properties were examined.

Summary: Carbon assimilation by Claviceps purpurea, growing as a parasite on cereals, has been investigated by supplying the host plant with 14CO2 in a closed system. The presence of the pathogen induced the plant to exude photosynthate which contained high levels of sucrose. During the period of 14CO2 supply, 14C was incorporated into the sucrose and so the path of carbon into the parasite could be traced. Hexoses, derived by the action of the fungal sucrase on sucrose, were assimilated by the pathogen and largely converted into polyols - mainly mannitol and, to a lesser extent, trehalose. The rate of carbohydrate metabolism decreased with maturation of the ergot, and also showed qualitative differences between the basal and apical regions of the ergot which were probably a function of nutrient supply.

Summary: Rhodanese (thiosulphate: cyanide sulphurtransferase EC. 2.8.1.1) was purified 25- to 30-fold from thiosulphate-grown Thiobacillus A2. It exhibited a pH optimum between pH 10·2 and 10·4 and apparent K M values of 0·36 mm-Na2S2O3 and 17 mm-KCN. Ultraviolet spectrophotometry and thin-layer chromatography showed that the enzyme catalysed the reaction of S2O3 2− with dihydrolipoic acid or dihydrolipoamide, producing α-lipoate or lipoamide, with the intermediate production of the persulphides of dihydrolipoate and dihydrolipoamide, which were demonstrated chromatographically. This is the first demonstration of catalysis by a thiobacillus rhodanese of reactions which are likely to be physiologically important in the oxidative dissimilation of thiosulphate by a central energy-conserving pathway.

Summary: Rhodanese from Thiobacillus a2 was shown by proton nuclear magnetic resonance (NMR) spectroscopy to use dihydrolipoate or dihydrolipoamide as acceptor of the sulphane moiety of thiosulphate with the formation of α-lipoate or lipoamide respectively. Correlation is shown between assays of the enzyme activity by NMR spectroscopy and by ultraviolet spectrophotometry.

Summary: Fluorescent antibodies (FA) prepared against the Mo-Fe and Fe proteins of nitrogenase from Klebsiella pneumoniae m5a1 were used to detect these protein components in toluene-treated whole cells that were actively reducing acetylene. The FA were highly specific, staining only nitrogenase component proteins originating from Klebsiella. Cross-reactions between the FA and purified nitrogenase proteins from other dinitrogen-fixing micro-organisms did not occur, except in the case of Bacillus polymyxa. The tests rapidly and accurately assayed the component proteins in Klebsiella mutants and derivatives to which Klebsiella nif genes had been transferred either by plasmid or by other means. Cross-reactions also indicated the degree of relatedness between nitrogenase proteins from dinitrogen-fixing micro-organisms of various origins.

Summary: Measurements of the specific activities of representative enzymes of the pentose phosphate cycle, Embden-Meyerhof-Parnas (EMP) pathway and the tricarboxylic acid (TCA) cycle in Coprinus cinereus sporophores at different stages of development indicate that glycolysis is the major route of carbohydrate catabolism throughout sporophore development. Enzymes of the pentose phosphate cycle were always at lower specific activities than the enzymes of the EMP pathway, and the activities of the pentose phosphate cycle enzymes declined drastically as development proceeded. This conflicts with the findings for Agaricus bisporus, but the changes in some enzymes were qualitatively similar to those occurring in the development of sporophores of Schizophyllum commune. A number of enzymes of the TCA cycle were detected, but there was no 2-oxoglutarate dehydrogenase activity nor increase of isocitrate lyase activity over the basal repressed level. However, glutamate decarboxylase and 4-aminobutyrate aminotransferase were detected, suggesting that the inoperative 2-oxoglutarate dehydrogenase is by-passed through the glutamate decarboxylation loop. The results are discussed in relation to the changes which also occur in the specific activities of the two glutamate dehydrogenase enzymes during development of the sporophore.

Summary: Ultrastructural changes in the plasma membrane of Saccharomyces cerevisiae during the exponential and stationary growth phases were studied by freeze-etching. In the exponential phase, plasma membrane-intercalated particles were distributed randomly. In the stationary phase, several areas of the plasma membrane showed a hexagonal arrangement of particles; these areas appeared to increase with the age of the culture. The polarity of the particles also changed partially: the E-face of the plasma membrane was only sparsely embedded with particles in exponential phase cells, but relatively densely embedded in stationary phase cells. Invaginations of the plasma membrane on the P-face were devoid of particles during both growth phases. Invaginations of the E-face were sparsely embedded with particles in exponential phase cells, but densely embedded with particles in stationary phase cells.

Summary: Arginine inhibited the competence of Bacillus subtilis growing in a chemostat at a dilution rate of 0·277 h−1. The biosynthesis of competence-stimulating activity was only partially repressed. The inhibitory effect may be due to an alteration in the cell’s capacity for being stimulated to competence and/or in its ability to take up DNA irreversibly. MnSO4 at 10−6 m restored competence immediately.

Summary: A study was made of growth, septation and branching in Geotrichum candidum, a mould which forms physiologically complete septa. A correlation was observed between septation and branch initiation; branches were almost invariably formed just behind septa. Primary branches and their parent intercalary compartments initially increased in length at an exponential rate before eventually attaining a constant rate of extension. The whole branching system (which eventually contained seven tips) produced by an intercalary compartment increased in length exponentially until it attained a total length of at least 1·5 mm.

The total length and the number of nuclei of undifferentiated mycelia increased exponentially at the same specific growth rate. The results suggest that nuclei divide just before or just after arthrospore formation.

Summary: Synchronous cultures of yeast and hyphal phases of Candida albicans showed exponential increases in RNA content and stepwise exponential increases in DNA content. The periods of DNA synthesis in the two phases coincided with one another and with the budding peaks of the yeast phase. Hyphae grown in synchronous cultures also showed an exponential increase in length. The hyphal phase was therefore normal.

Hyphal nuclear division occurred after hyphal DNA synthesis. Germination was a unique event for a hypha and unlike yeast bud formation, preceded the first period of DNA synthesis.

The exponential increase in RNA and DNA in asynchronous cultures of hyphae in serum paralleled the exponential increase in the numbers of cells in asynchronous cultures of yeasts in defined media. Thus there are no factors in serum which inhibit the normal exponential growth of C. albicans.

Summary: A chemically-defined medium was developed which supported growth of Streptococcus faecium and permitted synthesis of urease. This streptococcus cannot utilize ammonia and needs a complex medium, but its requirements are probably provided in the rumen. The specific activity of urease was inversely related to growth and in no medium was there high growth and high urease activity. Anaerobic culture and the presence of urea in the medium were essential for urease activity, but not for growth.

Summary: Azotobacter beijerinckii was grown in ammonia-free glucose/mineral salts media in chemostat culture under oxygen or nitrogen limitation. Selected enzymes of the tricarboxylic acid cycle and poly-β-hydroxybutyrate metabolism were monitored in relation to oxygen supply for both steady and transition states. Two dissolved oxygen concentrations were used for the nitrogen-limited steady state to investigate the possible effects of respiratory protection of nitrogenase on these enzymes. The levels of NADH oxidase, isocitrate dehydrogenase and 2-oxoglutarate dehydrogenase increased markedly on relaxation of oxygen limitation while pyruvate dehydrogenase and citrate synthase were relatively unaffected. β-Ketothiolase and acetoacetyl-CoA reductase levels decreased as oxygen limitation was relaxed. Respiratory activity, as measured by the Q o2 value, increased with oxygen supply rate. Imposition of oxygen limitation on a nitrogen-limited culture caused an immediate increase in the NADH/NAD ratio but this rapidly readjusted to its previous steady-state value. These changes are discussed in relation to respiratory protection of nitrogenase and poly-β-hydroxybutyrate metabolism in A. beijerinckii.