INACTIVATION OF ALPHA-KETOGLUTARATE DEHYDROGENASE DURING ITS ENZYMATIC-REACTION

The activity of some muscle alpha-ketoglutarate dehydrogenase complexe s (KGDC) decreases during their enzymatic reaction as a result of inac tivation of the first component of the complex, namely, alpha-ketoglut arate dehydrogenase (KGD). This decrease is associated with transforma tion of the complex produced by KGD and alpha-keto substrate in the co urse of oxidative phosphorylation. Kinetics of KGD inactivation during the reaction depends on the keto substrate structure. When alpha-keto glutarate (KG) is used as a substrate, KGD inactivation occurs in two stages. The major (irreversible) decrease in its activity is observed during the first minutes of reaction. During reaction with a catalytic ally active KG analog, alpha-ketoadipate (KA), the enzyme is irreversi bly inactivated in one stage. This suggests that the reversible stage of inactivation is due to the high rate of catalysis, which is charact eristic of KG-utilizing reactions. Decrease in the rate of KG oxidatio n after treatment of the enzyme with sulfhydryl reagents eliminates th is reversible stage. Preincubation of KGD with KG phospho-derivatives (succinylphosphonate or its monomethyl ether) changes the properties o f KGD in a similar way to the reversible decrease of activity during c atalysis. The arginine residue of KGD, which is essential for enzymati c activity, becomes inaccessible for butanedione in the complexes of K GD with some phospho-derivatives. The data suggest that the reversible inactivation of KGD in the course of catalysis is due to an interacti on of the leaving substrate carboxyl with the essential arginine resid ue of the enzyme.