Pressure relaxation of the equilibrium of the pig heart lactate dehydrogenase system.
The relaxation behaviour of a system of reactants equilibrated in the presence of pig heart lactate dehydrogenase was studied after pressure perturbation. Two relaxations were observed when protein fluorescence was recorded, but only the slower relaxation was apparent in observations of A340. The faster relaxation therefore involves transfer between free and enzyme-bound NADH, whereas the slower relaxation represents the reduction of NAD+. Both relaxations were observed in Tris buffer, where there is little effect of pressure on pH, and in phosphate buffer, where pH changes are significant; however, the amplitudes depended on the buffer used. The slower reciprocal relaxation time increases with increasing total enzyme concentration and decreases slightly with increasing NAD+ concentration. Computer simulations, based on a proposed mechanism, were compared with the experimentally determined amplitudes and relaxation times as a test of the mechanism.
Pressure relaxation of the equilibrium of the pig heart lactate dehydrogenase system. The relaxation behaviour of a system of reactants equilibrated in the presence of pig heart lactate dehydrogenase was studied after pressure perturbation. Two relaxations were observed when protein fluorescence was recorded, but only the slower relaxation was apparent in observations of A340. The faster relaxation therefore involves transfer between free and enzyme-bound NADH, whereas the slower relaxation represents the reduction of NAD+. Both relaxations were observed in Tris buffer, where there is little effect of pressure on pH, and in phosphate buffer, where pH changes are significant; however, the amplitudes depended on the buffer used. The slower reciprocal relaxation time increases with increasing total enzyme concentration and decreases slightly with increasing NAD+ concentration. Computer simulations, based on a proposed mechanism, were compared with the experimentally determined amplitudes and relaxation times as a test of the mechanism.