Mitochondrial permeability transition as induced by cross-linking of the adenine nucleotide translocase.

Abstract Mitochondrial permeability transition is caused by the opening of a transmembrane pore whose chemical nature has not been well established yet. The present work was aimed to further contribute to the knowledge of the membrane entity comprised in the formation of the non-specific channel. The increased permeability was established by analyzing the inability of rat kidney mitochondria to take up and accumulate Ca 2+ , as well as their failure to build up a transmembrane potential, after the cross-linking of membrane proteins by copper plus ortho -phenanthroline. To identify the cross-linked proteins, polyacrylamide gel electrophoresis was performed. The results are representative of at least three separate experiments. It is indicated that 30  μ M Cu 2+ induced the release of 4.3 nmol Ca 2+ per mg protein. However, in the presence of 100  μ M ortho -phenanthroline only 2  μ M Cu 2+ was required to attain the total release of the accumulated Ca 2+ , it should be noted that such a reaction is not inhibited by cyclosporin. The increased permeability corresponds to cross-linking of membrane proteins in which approximately 4 nmol thiol groups per mg protein appear to be involved. Such a linking process is inhibited by carboxyatractyloside. By using the fluorescent probe eosin-5-maleimide the label was found in a cross-linking 60 kDa dimer of two 30 kDa monomers. From the data presented it is concluded that copper– o -phenanthroline induces the intermolecular cross-linking of the adenine nucleotide translocase which in turn is converted to non-specific pore.