Isozymic nature of spore coat-associated alanine racemase of Bacillus subtilis.
Spore coat-associated alanine racemase of Bacillus subtilis, which converts L-alanine to D-alanine, that is, the germinant to the competitive inhibitor, to regulate spore germination for survival of the organism under unfavorable growth conditions, was examined. The dormant spores, L-alanine-initiated germination of which is inhibited by diphenylamine, were used to characterize the enzyme in the native form because of its unextractablility from dormant spores. The presence of isozymes, Enz-I and Enz-II with Km for L-alanine of about 20mM and 50mM and optimum activity at around 40 degrees C and 65 degrees C, respectively, was proposed. The enzymes were selectively used depending on the L-alanine concentration and the temperature. The pH profiles of the activity (optimun at pH 9.0) and the stability (stable between pH 6-11 at 60 degrees C) were similar, but Enz-II was more heat-stable than Enz-I and the denaturation curve demonstrated a two-domain structure for Enz-II. Sensitivity to D-penicillamine, hydroxylamine and HgCl2 was similar between Enz-I and Enz-II, while that to D-cycloserine, L- and D-aminoethylphosphonic acid, monoiodoacetate and N-ethylmaleimide was different; HgCl2 was the most effective inhibitor among these compounds.
Kanda-Nambu
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Department of Microbiology, Nagoya City University Medical School, Nagoya, Japan.
Yasuda
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Y
Tochikubo
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K
D000409:Alanine / Q000737:chemistry* / Q000378:metabolism*; D000411:Alanine Racemase / Q000037:antagonists & inhibitors / Q000737:chemistry* / Q000378:metabolism*; D001412:Bacillus subtilis / Q000201:enzymology*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004795:Enzyme Stability; D006863:Hydrogen-Ion Concentration; D019811:Hydroxylamine / Q000494:pharmacology; D007527:Isoenzymes; D007700:Kinetics; D013171:Spores, Bacterial / Q000737:chemistry / Q000201:enzymology; D013696:Temperature
D004791:Enzyme Inhibitors; D007527:Isoenzymes; D019811:Hydroxylamine; D000411:Alanine Racemase; D000409:Alanine