Induction of nonselective permeability of the inner membrane in deenergized mitochondria

Induction of the nonselective cyclosporin-sensitive pore in the inner mitoc hondrial membrane under conditions of complete dissipation of ion gradients and transmembrane potential was studied. This approach allows the kinetics of Ca2+-dependent pore opening and the preceding processes of induction to be studied separately. The effects of mitochondrial heterogeneity were als o minimized. We found that the kinetics of pore opening can be described by a minimal two-step scheme where only the rate constant at the first step d epends on Ca2+ concentration. Oxidation of pyridine nucleotides in the matr ix caused a slow transition in the pore complex and decreased the apparent dissociation constant of the Ca2+-binding site from >1 mM to similar to 30 mu M. N-Ethylmaleimide (but not disulfide-reducing agents) prevented and sl owly reverted the pore induction process. Data suggesting allosteric modula tion of the pore by pyridine nucleotides are presented.