Histidine decarboxylase of lactobacillus 30a: inactivation and active-site labeling by L-histidine methyl ester

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Abstract
Histidine decarboxylase from Lactobacillus 30a is rapidly and irreversibly inactivated upon incubation with L-histidine methyl ester. The rate of inactivation is 1st-order with respect to remaining active enzyme and exhibits saturation kinetics with a kinact [inactivation rate constant] of 1.2 mM and an apparent 1st-order rate constant of 0.346 min-1 at pH 4.8 and 25.degree. C. On complete inactivation, 3 mol of [14C]histidine (from L-[14C]histidine methyl ester) and 2 mol of 14C (from L-histidine [14C]methyl ester) are bound in nondialyzable form/mol (190,000 g) of protein inactivated with a corresponding loss of 3 of the 5 DTNB[5,5''-dithiobis(2-nitrobenzoic acid)]-titratable-SH groups essential for activity of the native enzyme. Imidazole propionate, a competitive inhibitor of the enzyme, protects against inactivation, loss of -SH groups, and incorporation of radioactivity from the histidine and the methyl ester moieties of the labeled inhibitor, and kinetic evidence indicates that imidazole propionate and histidine methyl ester compete for binding at the active site of histidine decarboxylase in a mutually exclusive manner. Treatment of the labeled protein with alkali or hydroxylamine results in the quantitative release of radioactivity. Inactivation of histidine decarboxylase by L-histidine methyl ester results from 2 modes of interaction between the inhibitor and the active site of histidine decarboxylase; the major interaction involves an essential -SH group.