Colonies of microorganisms capable of forming curdlan-type polysaccharide in glucose medium were found to stain blue with Aniline Blue, Brilliant Blue, or Trypan Blue, and red with Congo Red. Staining with Aniline Blue was highly specific for colonies forming curdlan-type polysaccharide. With the use of Aniline blue to detect curdlan-type polysaccharide and other β-1, 3-glucans, the distribution of these polysaccharides in microorganisms was investigated. Among 687 strains of bacteria, 5 strains of Bacillus stained with Aniline Blue, in addition to Alcaligenes faecalis var. myxogenes 10C3, some strains derived from it, and some strains of Agrobacterium. Sixteen of 60 strains of yeast also stained with this dye. No positive reaction was detected in 13 strains of fungi tested. This method was also useful for isolation of colonies forming only curdlan-type polysaccharide from organisms with the ability to form other water-soluble polysaccharides.
Changes in the metabolic pathways of hypoxanthine in a strain of Streptomyces sp. has been studied. Hypoxanthine was converted to inosine 5′-monophosphate by the cells of the early period of cultivation, in which a high activity of hypoxanthine phosphoribosyltransferase but none of xanthine dehydrogenase was found, and was oxidized to xanthine and 6, 8-dihydroxypurine by those of the middle and the late periods, in which both activities were observed. If the cells of the early growth period were starved of the nitrogen source, the activity of xanthine dehydrogenase increased and hypoxanthine stimulated the increase, whereas that of hypoxanthine phosphoribosyltransferase remained constant. Synthesis of the nucleotide but not the oxidation was inhibited under these conditions of nitrogen limitation. If the cells were then supplied with the nitrogen source, however, the purine base was metabolized to both the synthetic and oxidative directions at the same time. These results suggest that the nitrogen source in the medium plays an important role in the control of hypoxanthine metabolism; repressing the formation of xanthine dehydrogenase on the one hand and promoting the reaction of hypoxanthine phosphoribosyltransferase on the other.
The enzymatic transformation of cortisol with Bacillus cereus was examined in the presence of some redox agents and enzyme inhibitors. At the early phases of fermentation, the enzymic 1, 2-dehydrogenation reaction of cortisol was highly accelerated in the presence of oxidized glutathione. Thereafter, the enzymic reduction of the 20-keto group of cortisol and prednisolone was noticeably enhanced. In the presence of reduced glutathione, prednisolone was quantitatively converted to the 20β-hydroxy derivative. The bioconversion of cortisol to prednisolone, 20β-hydroxycortisol, and 20β-hydroxyprednisolone was also activated in the presence of azide and cyanide. With iodoacetate, the 1, 2-dehydrogenation reaction was promoted while the 20-keto reduction was markedly repressed. Pretreatment of the bacterial cells with various C19 and C21 steroids as well as with cholesterol affected the transformation pattern in different manners. The 1, 2-dehydrogenation reaction of cortisol was considerably induced when the cells were pretreated with Δ1-dehydroprogesterone, Δ1, 4-androstadienedione, and Δ1-dehydrotestosterone.
The activity of catalase (EC 18.104.22.168) in Escherichia coli was profoundly repressed by growth with such substrates as glucose, mannose, galactose, ribose, glycerol, and pyruvate. The low catalase activity of the cells grown with glucose increased about twice when the cells were incubated with phosphate buffer containing yeast extract for 2hr. This increase of catalase activity was enhanced by continuous addition of H2O2 during the incubation. The increase of catalase activity was completely inhibited by chloramphenicol and Puromycin. The synthesis of catalase induced by H2O2 was repressed by glucose, and the repression was not eliminated by cyclic-AMP.
A rotatory calorimeter of the conduction type was remodelled to fit the continuous study of microbiological processes. Provisions for aeration and agitation made it possible to be used for calorimetric studies of microorganisms aerobically growing in a 5-ml culture. The time constant of the calorimeter response was found to be 8min and the calorimeter could be used for most kinetic studies on microbial cultures. By using a separate culture apparatus, quantitative analyses can be made on samples taken from parallel culture and the results can be correlated with the thermogram obtained with the calorimeter. The performance of the instrument was demonstrated with the aerobic growth of Saccharomyces cerevisiae. Calorimetric study by the use of the present model may give a detailed information at the cellular level about microbial growth processes when combined with turbidity measurements.
The spore coat of Bacillus subtilis was isolated and solubilized by treatment with sodium dodecyl sulfate (SDS) plus dithiothreitol (DTT) solution. This treatment solubilized about 85% of the spore coat fraction and the soluble fraction mainly consisted of protein. The protein was purified in a buffer containing the detergent. The results of gel filtration, SDS-disc gel electrophoresis and NH2-terminal analysis showed the major component of the spore coat protein to be one kind with a molecular weight of 14, 000.