Purification and reconstitution of two anion carriers from rat liver mitochondria: the dicarboxylate and the 2-oxoglutarate carrier.
Two anion-transporting systems, i.e., the dicarboxylate carrier and the 2-oxoglutarate carrier, have been purified from rat liver mitochondria and functionally identified. The dicarboxylate carrier has been isolated in active form by hydroxyapatite chromatography after partial removal of the solubilizing detergent Triton X-114 from the mitochondrial extract. The SDS gel electrophoresis of this preparation consists mainly of one protein band with an apparent Mr of 28,000, identified as the dicarboxylate carrier. Complete purification of the 28 kDa protein in inactive form has been achieved by sequential chromatography on hydroxyapatite and Celite followed by SDS extraction of the retained protein. The 2-oxoglutarate carrier has been purified by hydroxyapatite chromatography after extensive removal of Triton X-114 from the detergent extract. SDS gel electrophoresis of the purified fraction shows a single band with an apparent Mr of 32,500. When reconstituted into liposomes, the functional properties of the two isolated carrier proteins resemble closely those of the dicarboxylate and the 2-oxoglutarate transport systems characterized in mitochondria.
Bisaccia
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Department of Pharmaco-Biology, University of Bari, Italy.
Indiveri
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C
Palmieri
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F
D000818:Animals; D002352:Carrier Proteins / Q000302:isolation & purification* / Q000378:metabolism; D029382:Dicarboxylic Acid Transporters; D003998:Dicarboxylic Acids / Q000378:metabolism*; D007656:Ketoglutaric Acids / Q000378:metabolism*; D007700:Kinetics; D008081:Liposomes; D026901:Membrane Transport Proteins; D008930:Mitochondria, Liver / Q000378:metabolism*; D008970:Molecular Weight; D051381:Rats
D002352:Carrier Proteins; D029382:Dicarboxylic Acid Transporters; D003998:Dicarboxylic Acids; D007656:Ketoglutaric Acids; D008081:Liposomes; D026901:Membrane Transport Proteins; C021723:oxoglutarate translocator