MITOCHONDRIAL PERMEABILITY TRANSITION AS INDUCED BY CROSS-LINKING OF THE ADENINE-NUCLEOTIDE TRANSLOCASE

Mitochondrial permeability transition is caused by the opening of a tr ansmembrane pore whose chemical nature has not been well established y et. 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 i nability of rat kidney mitochondria to take up and accumulate Ca2+, as well as their failure to build up a transmembrane potential, after th e cross-linking of membrane proteins by copper plus ortho-phenanthroli ne. To identify; the cross-linked proteins. polyacrylamide gel electro phoresis was performed. The results are representative of at least thr ee separate experiments. It is indicated that 30 mu M Cu2+ induced the release of 4.3 nmol Ca2+ per mg protein. However, in the presence of 100 mu M ortho-phenanthroline only 2 mu M C2+ was required to attain t he total release of the accumulated Ca2+, it should be noted that such a reaction is not inhibited by cyclosporin. The increased permeabilit y corresponds to cross-linking of membrane proteins in which approxima tely 4 nmol thiol groups per mg protein appear to be involved. Such a linking process is inhibited by carboxyatractyloside. By using the flu orescent probe eosin-5-maleimide the label was found in a cross-linkin g 60 kDa dimer of two 30 kDa monomers. From the data presented it is c oncluded that copper-o-phenanthroline induces the intermolecular cross -linking of the adenine nucleotide translocase which in turn is conver ted to nonspecific pore. (C)1998 Elsevier Science Ltd. All rights rese rved.