PURIFICATION AND CHARACTERIZATION OF THE TRICARBOXYLATE CARRIER FROM EEL LIVER-MITOCHONDRIA

The tricarboxylate carrier from eel (Anguilla anguilla) liver mitochon dria was solubilized with Triton X-100 and purified by sequential chro matography on hydroxyapatite and Matrex Gel Blue B. On SDS-polyacrylam ide gel electrophoresis, the purified fraction showed a single polypep tide band with an apparent molecular mass of 30.4 kDa. When reconstitu ted into liposomes, the tricarboxylate transport protein catalyzed a v ery active 1,2,3-benzenetricarboxylate-sensitive citrate/citrate excha nge. It was purified 641-fold with a recovery of 13.3% and a protein y ield of 0.02% with respect to the mitochondrial extract. The propertie s of the reconstituted carrier, i.e., requirement for a counteranion, substrate specificity and inhibitor sensitivity, were similar to those of the tricarboxylate carrier purified from rat liver mitochondria. T hese studies provide the first information on the mitochondrial tricar boxylate transport protein of a fish. (C) 1996 Academic Press, Inc.