Biosynthesis of α-Ketoglutarate by the Reductive Carboxylation of Succinate in Bacteroides ruminicola

Experiments with growing cells and with cell-free extracts of Bacteroides ruminicola indicate that this anaerobic bacterium can synthesize alpha-ketoglutarate by a reductive carboxylation of succinate. When the organism was grown in medium containing succinate-1,4-(14)C, most of the radioactivity in cells was in the protein fraction and most of the (14)C in protein was in the glutamic acid family of amino acids (glutamate, proline, and arginine). When unlabeled succinate was added to culture medium containing glucose-U-(14)C, incorporation of radioactivity into the glutamic acid family of amino acids was greatly reduced. This supports the concept that succinate is an intermediate in synthesis of alpha-ketoglutarate. Cell-free extracts of the organism incubated with succinate-1,4-(14)C incorporated (14)C into amino acids and most of this was found in glutamate. The cofactors which stimulate glutamate synthesis from succinate by extracts from these cells appear to be similar to the factors that have been demonstrated with extracts from photosynthetic bacteria. The position of label in glutamate synthesized from succinate-1,4-(14)C, the probable absence of isocitric dehydrogenase, and studies with labeled citrate and with inhibitors of citric acid cycle enzymes support the concept of a reductive carboxylation of succinate as the only, or at least a major, mechanism for synthesis of alpha-ketoglutarate in this organism. This appears to be the first evidence for a net synthesis of alpha-ketoglutarate by this reaction in a nonphotosynthetic heterotrophic organism.