Adenine nucleotide translocator isoforms 1 and 2 are differently distributed in the mitochondrial inner membrane and have distinct affinities to cyclophilin D

Different isoforms of the adenine nucleotide translocase (ANT) are expresse d in a tissue-specific manner. It was assumed that ANT-1 and ANT-2 co-exist in every single mitochondrion and might be differently distributed within the membrane structures that constitute the peripheral inner membrane or th e crista membrane. To discriminate between ANT originating from peripheral or from cristal inner membranes we made use of the fact that complexes betw een porin, the outer-membrane pore protein, and the ANT can be generated. S uch complexes between porin and the ANT in the peripheral inner membrane we re induced in rat heart mitochondria and isolated from rat brain and kidney . Using ANT-isotype-specific antibodies and sequence analysis of the N-term inal end, it was discovered that the peripheral inner membrane contained AN T-1 and ANT-2, whereas the cristal membrane contained exclusively ANT-2. Cy clophilin was co-purified with the porin-ANT complexes, whereas it was abse nt in the crista-derived ANT. This suggested that ANT-1 might have a higher affinity for cyclophilin. This specific intramitochondrial distribution of the two ANT isotypes and cyclophilin D suggests specific functions of the peripheral and crista-forming parts of the inner membrane and the two ANT i sotypes therein.