The l-amino acid oxidases of snake venom. V. Mechanism of the reversible inactivation

1. 1. The spontaneous, reversible inactivation of the l -amino acid oxidase of snake venoms has been compared with reversible processes which regulate the activity of other enzymes. 2. 2. Since no change in the size, shape, electrophoretic mobility or solubility of the oxidase has been detected upon extensive inactivation of the enzyme, the process has been interpreted as involving only a limited alteration of the configuration of the native enzyme rather than an extensive unfolding. 3. 3. The mechanism of this reversible process has been interpreted as follows. The active enzyme is in reversible equilibrium with an inactive form, which differs from the active form by the presence of a new acidic group not present in the native enzyme, or an acidic group whose dissociation constant changes by about two pH units as a result of structural alterations in the protein during the inactivation step. The probable nature of this ionizing group has been discussed and its possible identity with an imidazole nitrogen, located at the active center of the enzyme, has been pointed out. 4. 4. On the basis of the above hypothesis, a kinetic development has been presented to account for the fact that the rate of reactivation of the enzyme is a function of pH while the rate of inactivation is not. 5. 5. It has been suggested that the inactivation process may involve the reversible breakage of one or two strong covalent linkages or of several weaker hydrogen bonds and that at least one of these may be joined to an imidazole ring at the active center of the enzyme. 6. 6. The possible nature of the ionizing groups at the active center of the enzyme is discussed from considerations of the influence of pH on the activity and stability of the enzyme.