Effects of D , O on the Association - Dissociation Equilibrium in Subunit Proteins

Heavy water has been used as a tool to study the role of water in the association-dissociation equilibrium of subunit proteins. In the case of lactate dehydrogenase, DzO was found to stabilize the enzyme at low ionic strength and inhibit subunit interchange between heartand muscle-type enzyme in solution. When frozen, however, subunit interchange occurred to a similar extent in either DzO or HzO. In the case of bovine liver glutamate dehydrogenase, DtO appears to facilitate the association of monomers to form polymers. The maximum weight average molecular weights of glutamate dehydrogenase were found to be approximately doubled in DzO compared with HzO, and the dissociation constant of the polymer appeared to be much smaller in D20. The weight average molecular weights were calculated from sedimentation velocity and diffusion rate measurements in the analytical ultracentrifuge. Because these measurements were made at 8”, at which the contribution of hydrophobic bonding would be minimal, it was concluded that the stabilizing action of D20 reflected the increased strength of deuterium bonds and deuterium water bridges. Alteration of the association-dissociation equilibrium in regulatory proteins by DzO is a possible mechanism to explain certain of the potent inhibitory effects of DzO on biological systems.