The mitochondrial tricarboxylate carrier of silver eel: dimeric structure and cytosolic exposure of both N- and C-termini.
The mitochondrial tricarboxylate carrier plays a fundamental role in the hepatic fatty acid synthesis. In this study, we investigated the transmembrane organization of this protein in the inner membrane of eel liver mitochondria using anti-N-terminal and anti-C-terminal antibodies. These antibodies recognized the N- and C-termini of the tricarboxylate carrier in intact mitoplasts, thus suggesting a cytosolic exposure of these regions in the membrane-bound protein. This structural arrangement of the tricarboxylate carrier was further confirmed by protease treatment of intact mitoplasts. Moreover, the oligomeric state of the native tricarboxylate carrier was investigated by blue native electrophoresis. A dimeric form of the carrier protein was found when eel liver mitochondria were solubilized with the mild detergent digitonin. These findings suggest an arrangement of the dimeric tricarboxylate carrier into an even number of membrane-spanning domains, with the N-terminal and C-terminal regions oriented toward the intermembrane space of fish mitochondria.
Capobianco
Loredana
L
Dipartimento di Scienze e Tecnologie Biologiche ed Ambientali, Università di Lecce, Via Prov.le Lecce-Monteroni, 1-73100 Lecce, Italy.
Ferramosca
Alessandra
A
Zara
Vincenzo
V
D000818:Animals; D015153:Blotting, Western; D002352:Carrier Proteins / Q000737:chemistry* / Q000378:metabolism*; D003600:Cytosol / Q000378:metabolism*; D004072:Digitonin / Q000494:pharmacology; D019281:Dimerization; D004305:Dose-Response Relationship, Drug; D004524:Eels; D004591:Electrophoresis, Polyacrylamide Gel; D004797:Enzyme-Linked Immunosorbent Assay; D007425:Intracellular Membranes / Q000378:metabolism; D008928:Mitochondria / Q000378:metabolism*; D017434:Protein Structure, Tertiary; D013997:Time Factors
D002352:Carrier Proteins; C020723:citrate-binding transport protein; D004072:Digitonin