Malate dehydrogenase, inhibition of pig heart supernatant enzyme by iodoacetamide.

Abstract

Pig heart supernatant malate dehydrogenase is alkylated by 250 mM iodoacetamide at 37 degrees C in pH 7.5 Tris/acetate buffer which is 0.05 M with respect to acetate to form enzyme with 1,3-dicarboxamidomethyl histidine, 3-carboxamidomethyl histidine, 1-carboxamidomethyl histidine, carboxamidomethyl cysteine, and carboxamidomethyl methionine. 1,3-Dicarboxamidomethyl histidine forms with a stoichiometry of 1/enzyme subunit at a rate equaling the rate (approximately 1.24 +/- 0.09 X 10(-2) min-1) of enzyme inactivation by iodoacetamide. All other amino acid derivatives form at a slower rate initially or continuously. The enzyme is protected against inactivation by iodoacetamide through NADH binding, suggesting that there is an "essential" histidine residue at or near the active site.