Inactivation of the NAD-dependent hydrogenase from the hydrogen-oxidizing bacterium Alcaligenes eutrophus Z1. Thermoinactivation mechanism

Abstract The thermoinactivation of the NAD-dependent hydrogenase (Hydrogen:NAD + oxidoreductase, EC 1.12.1.2) from the hydrogen-oxidizing bacterium Alcaligenes eutrophus Z1 under aerobic conditions was studied. The dissociative character of the thermoinactivation was demonstrated. The existence of inactivation intermediates of about M r 80 000 was established by HPLC and gradient polyacrylamide gel electrophoresis. Aggregates of M r 600 000–800 000 were the final products of thermoinactivation. A kinetic scheme for hydrogenase thermoinactivation taking into account the dissociation of the native enzyme tetramer into constituent heterodimers was postulated. All the relevant kinetic constants were calculated using a computer simulation procedure.