The Fixation of Carbon Dioxide by Rat Liver Mitochondria and Its Relation to Gluconeogenesis

Abstract

SUMMARY The finding reported here, that isolated mitochondria can synthesize malate in good yield from pyruvate and COz, offers an alternative to the transmembrane movement of oxaloacetate. Thus, malate could be formed within the mitochondria by the action of malic dehydrogenase and NADH on oxaloacetate. The malate could pass through the mitochondrial membrane and then be re-oxidized to oxaloacetate by the malic dehydrogen- ase of the cell sap with subsequent conversion of this extramito- chondrial oxaloacetate to P-enolpyruvate. 2. A net synthesis of malate and citrate was demonstrated in mitochondria incubated under these conditions. 3. Essentially all of the radioactive compounds formed were released from the mitochondria during the incubation. 4. It is proposed that in rat liver the initial stages of gluco- neogenesis from pyruvate proceed as follows. In the mito- chondria, pyruvate is carboxylated to oxaloacetate. The oxalo- acetate, which does not react with acetyl coenzyme A to form citrate, is reduced to malate. Malate or fumarate or both cross the mitochondrial membrane into the cytoplasm of the cell. There malate is oxidized to oxaloacetate which is in turn con- verted to phosphoenolpyruvate. Fumarate could function in a similar manner, and it does not appear feasible at the present time to devise experiments to choose between malate and fumarate as the compound which moves across the membrane. Thus, in the experiments reported here, the mitochondria were twice washed, but there might well have been fumarase present outside the mitochondria which would have regenerated malate from fumarate, had only fuma- rate crossed the mitochondrial membrane. AcknowledgmentsWe are indebted to Miss Rosalind Levene of Dr. Cecil Cooper’s laboratory for assessing the respiratory con- trol of our mitochondrial preparation. The helpful criticism of Dr. M. F. Utter during the preparation of this manuscript is greatly appreciated. The technical assistance of Mr. Glenn DeBoer and Mr. Endre Solti is gratefully acknowledged.