Studies on Succinate Dehydrogenase

Abstract

Mammalian succinate dehydrogenase exists in unactivated and activated states. Transition of the unactivated enzyme to the activated form appears to be a conformation change initiated by combination of the protein with substrates or competitive inhibitors and results in a many-fold increase in catalytic activity. This activation has now been shown to be reversible on removal of the bound activating agent by gel exclusion in the case of the soluble enzyme and by repeated washing and centrifugation or gel exclusion in the case of particles. While the activation has a very high energy of activation and does not proceed at observable rates at 15°, the deactivation is rapid even at temperatures near 0°. The ease of deactivation depends on the tightness of the binding of the activator. With tightly bound activators, such as malonate, replacement of the bound activator with succinate, followed by removal of the latter on Sephadex columns, has been found to be particularly effective. A modest degree of activation of succinate dehydrogenase occurs in particulate and soluble preparations also without added activating agents at 38°. This type of temperature activation may be due to the presence of traces of bound activator in the preparation. The activation-deactivation cycle has major effects on succinoxidase activity and on all types of succinate dehydrogenase assays, but the fumarate-FMNH2 reaction (fumarate reductase activity) is unchanged by these treatments. The spectral shifts accompanying the activation are not reversed by removal of the activator and consequent deactivation. It appears, however, that the colored enzyme-competitive inhibitor complexes form only with the fully activated enzyme, although the same competitive inhibitors combine with the unactivated form in a colorless complex.