[Effect of anions on inhibition of lactate dehydrogenase by pyruvate]

The effect of anions Cl- and I- on structural and kinetic properties of LDH was investigated. It was shown that anions are specific inhibitors of LDH competing with pyruvate in the active ternary complex, LDHNADHpyq. The following dissociation constants for the anions were obtained from inhibition data: 0.4 +/- 0.02 and 0.07 +/- 0.01 M for Cl- and I-, respectively. The slope of Hill plot are near 1.0. The anions abolished the inhibition of LDH at high pyruvate concentrations. The following dissociation constants were obtained from these data: 0.1 and 0.015 M for Cl- and I- respectively. The inhibition by anions and the abolishing of substrate inhibition by anions were studied also for the lactate oxidation reaction. The dissociation constants for anions obtained from these data are in good correlation with the constants obtained for the pyruvate reduction reaction. It was concluded that anions do not interact with the group at the catalytic site with pK approximately 7.8, presumably His-195. The degree of pyruvate inhibition does not depend on the buffer system. The differences in the degree of inhibition obtained previously in phosphate, imidazole and tris-buffer systems can be explained by the presence of Cl- anions in the last two buffer. The rate constants of hydroxy leads to keto pyruvate transition was obtained in various buffer systems. It was shown that the hydroxy-form of pyruvate does not cause the inhibition of LDH.