Role of arginine residues of bovine liver dihydrodiol dehydrogenase 2 in the binding of anionic substrates.

Bovine liver dihydrodiol dehydrogenase (DD2) was inactivated following pseudo-first order manner by the treatment of 5 mM 2,3-butanedione (BD) as functions of incubation-time and concentration. Anionic substrates or analog which have a carboxyl group, D-glucuronate, p-carboxybenzaldehyde and D-glycerate, protected DD2 efficiently. But, the other substrates or coenzymes and their analogs did not show any protection on the inactivation, i.e., D,L-glyceraldehyde, D-erythrose, NADP+, NAD+, 2',5'-ADP, 2'-AMP. Results of kinetic analyses suggest that the inactivated enzyme lost its binding ability to anionic substrates. The inactivated enzyme was reactivated very effectively by removing excess BD by gelfiltration for 30 min.