2-Bromoacetamidopyridine: A new chemical probe of the active site of histidine decarboxylase from Lactobacillus 30a

Abstract

2-Bromoacetamidopyridine was examined as a potential active site-directed alkylating agent for histidine decarboxylase from Lactobacillus 30a. At the optimum pH of the enzyme (pH 4.8), the reagent binds reversibly to the substrate binding site as evidenced by its ability to competitively inhibit L-histidine decarboxylation, to protect the enzyme against inactivation by the substrate analog L-histidine methyl ester and to inhibit Schiff-base binding of histamine to the pyruvoyl prosthetic group. Upon raising the pH of the reaction medium from 4.8 to 7.2, 2-bromoacetamidopyridine irreversibly inactivates the decarboxylase. Incorporation experiments with 2-bromo[1-¹⁴C]acetamidopyridine demonstrate that, under inactivating conditions, one reagent molecule is covalently bound per catalytic unit of enzyme inactivated.