Reversible Inhibition of NADPH-Cytochrome P450 Reductase by α-Lipoic Acid

Abstract

NADPH-cytochrome-P450 reductase both purified from rat hepatic microsomes and involved in microsomal fraction was inactivated by treatment with α-lipoic acid. Since α-lipoic acid contains disulfide bond, it reacts with SH-groups of the reductase via the reaction of thiol-disulfide exchange resulting in the loss of the enzyme reducing activity. NADP + completely protected reductase from the inactivation. The modification of reductase was reversible: the modified enzyme was partially reactivated with dithiothreitol and dihydrolipoic acid in the case when cytochrome c was used as a substrate of reductase. In the case when inorganic substrate, K 3Fe(CN) 6, was used for assay the activity of modified reductase no reactivation was observed. It was found that the order of the reaction of inactivation of membrane-bound microsomal reductase is equal to 1.2 ± 0.2, which is in an agreement with pseudo-first order kinetics, and the second-order-rate constant of 26 M −1min −1. The results have shown that well known therapeutic agent α-lipoic acid is an efficient inhibitor of both purified and microsomal reductase.