Extraction, partial purification and functional reconstitution of two mitochondrial carriers transporting keto acids: 2‐oxoglutarate and pyruvate

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Abstract
Bovine heart submitochondrial particles were treated with a medium containing Triton X‐114 and cardiolipin. The extract was subjected to hydroxyapatite chromatography. Only a few major polypeptides of similar molecular masses were found in the eluate, as shown by electrophoresis in an SDS‐polyacrylamide gel stained with silver. The eluate was reconstituted into liposomes and was shown to catalyse two different transport activities: 2‐oxoglutarate‐2‐oxoglutarate exchange sensitive to phthalonate and phenylsuccinate and pyruvate‐pyruvate exchange sensitive to 2‐cyano‐4‐hydroxycinnamate. Since both activities were found to have characteristics similar to those described for intact mitochondria, it was concluded that at least two of the polypeptides found in the hydroxyapatite eluate correspond to the two mitochondrial carriers.