Addition of L-proline to the production medium for cultivation of strain No. 14-5, an isoleucine auxotrophic mutant of Brevibacterium flavum 2247, reduced L-proline accumulation by about 40%. The formation, but not the activity, of glutamate kinase, the first enzyme required for L-proline biosynthesis from L-glutamate, seems to be controlled by L-proline. No difference was observed in specific activities of this enzyme assayed in vitro in either strain No. 14-5 or the parent strain. During the growth of strain No. 14-5 (lacking in threonine dehydratase activity), the increase of intracellular glycine, methionine, aspartate, and threonine was considerably greater than in the parent strain. Intracellular valine, leucine, and lysine also increased. L-Proline produced was reduced to 42.9% without changing the level of intracellular glutamate by the addition of L-threonine and L-lysine to the medium before cultivation, but L- proline production was increased by nearly 20% when both amino acids were added near before the growth of this strain arrived at the stationary phase. Glutamate kinase formation was repressed by nearly 20% by the addition of both amino acids, whereas the activity was not inhibited. A cell homogenate of the parent strain, which was not able to produce L-proline in the medium, did produce L-proline from L-glutamate in the presence of ATP. From these findings, it was concluded that in strain No. 14-5 both the higher level of available ATP resulting from the inhibition of aspartate and homoserine kinase and the accumulation of intracellular L-glutamate promoted by biotin-rich condition contributed to the abundant production of L-proline through stimulation of the glutamate kinase reaction.
1.The presence or absence of GMP reductase had no effect on nucleoside production in adenine-requiring mutants of Bacillus subtilis K, which produced a large amount of inosine when cultured in an adenine-limited medium. 2. Mutants resistant to 8-azaxanthine were derived from strain 38-3, a GMP reductase-positive inosine producer, and from strain 30-12, a GMP reductase-negative inosine producer. About 60% of the resistant mutants from strain 38-3 were found to produce xanthosine as well as inosine, while 70% of the resistant mutants from strain 30-12 produced guanosine as well as inosine. Loss of the adenine requirement in these 8-azaxanthine-resistant mutants reduced the accumulation of xanthosine or guanosine and completely halted the accumulation of inosine. Many more mutants with an altered nucleoside accumulation pattern were obtained when 8-azaxanthine was used as a selective agent than when 8-azaguanine was used. 3.Guanine-requiring mutants, GU-16 and 308-18, were isolated respectively from an 8-azaxanthine-sensitive inosine producer and from an 8-azaxanthine-resistant xanthosine-inosine producer, and the effects of guanosine on xanthosine and inosine formation were compared in these two guanine auxotrophs; strain GU-16 produced a large amount of xanthosine only when grown in media containing limited amounts of guanosine, and the production was sharply reduced by higher amounts of guanosine. In strain 308-18, however, xanthosine formation was not so greatly affected by exogenous guanosine as in strain GU-16. There was no difference in the effect of guanosine on the total amount of purine nucleosides produced, including both xanthosine and inosine, in these two strains.
The activity of 6-phosphogluconate dehydrogenase (not decarboxylating) in the sonicate was found to be nearly equal to that of heterolactic fermentation in the washed cells of Leuconostoc mesenteroides B 07, showing a significant participation of the dehydrogenase in the metabolic pathway of the fermentation system in this organism. It was found that, at pH 5.9 (intracellular pH), activity of the dehydrogenase and the apparent equilibrium constant for this reaction are increased as much as 13- and 120-fold, respectively, at high concentrations of the enzyme protein with a change in quaternary structure.
The activity of 2-keto-6-phosphogluconate decarboxylase in the sonicate of washed cells of Leuconostoc mesenteroides B07 was estimated. The value calculated for intracellular activity of the decarboxylase showed good agreement with that for heterolactic fermenting activity of the washed cells. It was concluded that oxidative decarboxylation of 6-phosphogluconate in the organism depends quantitatively on the decarboxylase.
Plasmalogens, aldehyde-containing phospholipids, were found to occur widely in all four fractions of ruminal microbes obtained after differential centrifugation of sheep rumen contents. Plasmalogens were also detected in ruminal and soil anaerobic bacteria which were obtained by enrichment cultures with various energy sources. They occurred widely in all strictly anaerobic bacteria and Propionibacterium tested. The strains included Desulfovibrio sp., Selenomonas ruminantium, Bacteroides ruminicola, Veillonellagazogenes, Peptostreptococcus elsdenii, Propionibacterium freudenreichii, Propionibacteriumshermanii, Clostridium saccharoperbutylacetonicum, Clostridiumacetobutylicum, Clostridium perfrigens, Clostridium kaneboi, and Clostridiumkainantoi. Although the ubiquity of plasmalogens in these anaerobic bacteria was confirmed, the amount of plasmalogens per unit weight of cells varied considerably depending on bacterial species. Amount of plasmalogens in Selenomonas and Propionibacterium also varied by cultural conditions. However, plasmalogens were not detected in aerobic bacteria and in facultative anaerobic bacteria even when they were cultured under strictly anaerobic conditions.
Cells of Bacillus subtilis grown to the age of middle or late logarithmic phase were sensitive to saline and lost their colony-forming ability rapidly during the liquid holding. The sensitive cells turned to resistant cells when the growth progressed to stationary phase. The sensitive cells were generally competent, that is, able to take up DNA and perform transformation. Release of DNA and transformation-stimulating agents from the sensitive cells were observed and the saline-sensitive phenomenon in Bacillus subtilis was compared with the cold-sensitive phenomenon in Escherichia coli.
Revertants obtained from three threonine-requiring mutants of Bacillussubtilis were studied. The revertants from mutants 347 and 350 did not form normal colonies at 37° while they did at 50° (temperature-dependent), and the revertants from mutant 530 did not form colonies at 50° while they did at 37° (temperature-sensitive). By genetic analysis, one reversion observed in 350 was attributed to a mutation occurring in the same cistron, and one reversion observed in 530 was attributed to a suppressor mutation occurring at a map position of 0.64. Mechanism of reversion was discussed.
Both mycobacillin production and sporulation are initiated in the postlog phase of growth of B. subtilis B3, reaching their maximum by about the same time. Mycobacillin is released during enndotropic sporulation of vegetative cells though not during germination of spores in complex growth medium. Antisporogenic chemicals like glucose (in excess), diethylmalonate, acriflavin, fluoroacetic acid, sodium bisulphite, β-phenethyl alcohol, α-picolinic acid and m-tyrosine inhibit mycobacillin production. By acriflavin and actinomycin D treatment, two types of mutants, oligosporous and asporogenous, were obtained. Mycobacillin production is affected adversely in oligosporous mutants whereas asporogenous mutants do not produce the antibiotic at all.