THE α-KETOGLUTARIC OXIDASE SYSTEM OF AZOTOBACTER

Abstract

A soluble alpha-ketoglutaric oxidase system obtained from A. vinelandii, strain O, has been purified 10-fold by a protamine precipitation, fractionation with ammonium sulfate, and adsorption and elution from Ca phosphate gel. Diphosphothiamine, diphosphopyridine nucleotide, and Mg++ are required for full activity when ferricyanide is used as the electron acceptor. Added phosphate, arsenate, alpha-lipoic acid, or coenzyme A is inactive in this system. The oxidase reduces diphosphopyridine nucleotide in the presence of large amts. of coenzyme A. Another enzyme fraction (deacylase) was obtained from Azotobacter extracts which, when coupled with the bacterial oxidase, reduces diphosphopyridine nucleotide in the presence of catalytic amts. of coenzyme A. The bacterial oxidase and deacylase couple with the succinyl-Coenzyme A deacylase and oxidase obtained from pig heart. This is evidence that succinyl-coenzyme A is an intermediate in the oxidation of alpha-ketoglutarate by diphosphopyridine nucleotide with extracts from Azotobacter.