THE MITOCHONDRIAL OXOGLUTARATE CARRIER PROTEIN CONTAINS A DISULFIDE BRIDGE BETWEEN INTRAMEMBRANOUS CYSTEINE-221 AND CYSTEINE-224

The oxoglutarate carrier (OGC) purified from bovine heart mitochondria was treated, both in its active and in its SDS-denatured state, with the fluorescent N-(1-pyrenyl)maleimide and other SH reagents before an d after reduction with dithioerythritol or beta-mercaptoethanol, The n umber of SH groups per OGC polypeptide chain was found to be about 1 f or the oxidized carrier and 3 for the reduced carrier, The bovine oxog lutarate carrier contains three cysteines: Cys-184, Cys-221 and Cys-22 4. Sequencing of BrCN cleavage products of oxoglutarate carrier showed that N-(1-pyrenyl)maleimide binds to only Cys-184 of the oxidized pro tein and also to Cys-221 and Cys-224 after reduction of the protein, T hese results show the presence of a disulfide bridge between the latte r two cysteines of the purified carrier. The oxidized and the reduced forms of the oxoglutarate carrier exhibited different V-max but virtua lly the same K-m values for oxoglutarate.