Oxidation of NADH by vanadium: Kinetics, effects of ligands and role of H 2O 2 or O 2

Abstract

The mechanism of oxidation of NADH by either vanadium(V) or vanadium(IV) was examined in the presence of reducing agents, complexing agents, and hydrogen peroxide. Reducing agents that stimulate the oxidation of NADH by V(V) include: a variety of cysteine analogues, glutathione, β-mercaptoethanol, dithiothreitol, and ascorbate. Complexing agents which stimulate NADH oxidation by V(V) include cystine, glutathione disulfide, and dehydroascorbate. Vanadium(IV)-dependent systems which oxidize NADH include combinations of V(IV) with cysteine or air alone. Combination of either V(V) or V(IV) with hydrogen peroxide leads to NADH oxidation. Based on kinetic analysis and the use of the diagnostic inhibitors—superoxide dismutase, catalase, albumin, mannitol, ethanol, and anaerobic conditions—we have assigned two major mechanisms of NADH oxidation. One is the previously reported mechanism which involves V(V)-superoxide as the NADH oxidant. This reaction is inhibited by superoxide dismutase and anaerobic conditions but not by catalase or ethanol. This reaction is observed for V(V) in the presence of reducing agents and complexing agents. The second reaction mechanism operates when V(IV) comes in contact with hydrogen peroxide and involves V(III)-superoxide as the NADH oxidant. This reaction is inhibited by catalase (if unligated hydrogen peroxide is an intermediate) and superoxide dismutase but not anaerobic conditions or ethanol. This mechanism is observed for reactions of V(IV) with air or hydrogen peroxide.