Hiroshi Nishira

Metabolism of Aniline by Rhodococcus erythropolis AN–13

The metabolic pathway of aniline was examined in Rhodococcus erythropolis AN-13 that was isolated from soil when aniline was provided as a sole source of carbon and nitrogen. cis, cis-Muconic acid and β-ketoadipic acid were detected by thin-layer chromatography in an incubation mixture containing aniline and resting cells of this strain. These two carboxylic acids were also formed from catechol, when the substrate was incubated with cell-free extract of aniline-grown cells, and characterized spectrally as crystalline samples. Ammonia was released from aniline by resting cells. The cell-free extract of aniline-grown cells had a strong catechol 1,2-dioxygenase activity. Catechol, once formed from aniline, was apparently converted so rapidly to cis, cis-muconic acid that it could not be isolated. These results suggest that R. erythropolis AN-13 converted aniline to catechol with the release of ammonia and then mineralized catechol ultimately to inorganic end products, H2O and CO2, through the β ketoadipic ac...

Isolation of Aniline-assimilating Bacteria and Physiological Characterization of Aniline Biodegradation in Rhodococcus erythropolis AN-13

Twenty bacterial strains were isolated from soil, when aniline was provided as a sole source of carbon and nitrogen. Rhodococcus erythropolis AN-13, one of the isolates, grew on aniline at concentrations from 0.65 to 2.6mg/ml and completely degraded it. Ammonia was released from aniline with cell growth and accumulated in the cultural broth. Aniline-grown cells of this bacterium oxidized aniline and catechol, but did not take up a significant amount of oxygen in the presence of o- or p-aminophenol. These facts suggest that R. erythropolis AN-13 metabolized aniline through catechol. Additional carbon and nitrogen sources did not cause any repression of aniline biodegradation in this strain, but promoted its breakdown with an increase of cell growth. The bacterium used aniline in preference to glucose in a medium containing these two substrates.

Production of Phenolic Compounds by Aspergillus S-4 in Sake Cake Medium and Identification of Terrein

High performance liquid chromatography (HPLC) analysis of the culture filtrate of an isolated mold, Aspergillus terreus S-4, showed a larger peak of a terrein-like substance than those of gallic acid and protocatechuic acid. The chemical structure of this substance was investigated and identified as terrein, 4,5-dihydroxy-3-propenyl- 2-cyclopenten-1-one. We also examined some culture conditions for the production of terrein using a sake cake medium for strain S-4.

Tannic acid staining and extraction of enzymes in polyacrylamide gel electrophoresis

Abstract A procedure was developed for a rapid staining of proteins in polyacrylamide gels with tannic acid and the extraction of enzymatic activity from the gels. Lysozyme and Taka-amylase A were stained with tannic acid and localized on pH 4.3, and 8.0 and 9.5 gels, respectively. After the gels were rinsed in buffer solutions, the activities of the enzymes were recovered in good yield from the gels. The use of these techniques is discussed.

Immunological studies on β-amylase of Bacillus cereus

SummaryAn antiserum against the β-amylase from Bacillus cereus BQ10-S1 Spo II was prepared using rabbits. The antiserum obtained was confirmed to form a specific immunoprecipitate with the purified β-amylase and showed a single band of protein with a molecular weight of 6.0x104 on the nitrocellulose sheet by the Western-Blotting method. The antiserum showed a precipitin line with the β-amylase from B. megaterium strain no. 32 by the Ouchterlony technique. However, the spur was formed on the Ouchterlony plate between the line of immunoprecipitin of the β-amylase from B. cereus BQ10-S1 Spo II and that from B. megaterium strain no. 32. On the other hand, no immune reaction occurred with the β-amylase from B. polymyxa no. 72 and those from higher plants such as soybean and barley. B. cereus BQ10-S1 Spo II was found to secret β-amylase mainly from the mid to the late logarithnic phase of cell growth. With the use of antiserum, the amount of the β-amylase secreted was estimated to be about 52 μg/109 cells, that of the parent strain (B. cereus BQ10-S1) about 14 μg/109 cells. These quantities of β-amylase corresponded in each case with enzyme productivity of the two strains (about 1,100 U/ml and 270 U/ml).

Lytic Enzyme toward Aniline-assimilating Rhodococcus erythropolis AN-13: Screening and Production

A bacterial strain, SH-548, that produces a lytic enzyme toward intact cells of aniline-assimilating Rhodococcus erythropolis AN-13, was isolated from soil. The isolated bacterium was identified as a Flavobacterium species. The growth conditions for the enzyme production by Flavobacterium sp. SH-548 were examined; organic nitrogen compounds, such as meat extract and Polypepton, were effective for its production. The lytic enzyme of this strain lysed intact cells of Rhodococcus, Bacillus, Nocardia, Corynebacterium, Brevibacterium, Streptococcus, Micrococcus, Cellulomonas and DAB (diaminobutyric acid)-type coryneform bacterial strains. However, it did not act on those of Staphylococcus aureus or gram-negative bacteria, Enterobacter, Escherichia, Klebsiella, Proteus or Pseudomonas strains. Bacterial strains having cell walls of the glycolyl type were readily lysed by this enzyme.