(8) THE MOLECULAR MECHANISM OF THE MITOCHONDRIAL PERMEABILITY TRANSITION PORE

A.P. Halestrap, S.J. Clarke, G.P. McStay and J.P. Gillespie
Department of Biochemistry, School of Medical Sciences, University of Bristol, Bristol BS8 1TD

We have proposed that the mitochondrial permeability transition pore (MPTP) is formed through a calcium mediated conformational change of the adenine nucleotide translocase (ANT), facilitated by the binding of mitochondrial cyclophilin (CyP-D) which has peptidyl-prolyl cis-trans isomerase activity.  Data will be presented to demonstrate that the MPTP can be reconstituted into proteoliposomes using purified ANT and CyP-D, and assayed using a novel continuous spectrophotometric technique.  Cyclosporin A can inhibit pore opening provided it is added before CyP-D binds to the ANT.  The formation of the MPTP requires neither porin nor any other outer membrane protein, although this does not preclude such proteins having a regulatory role.  A major problem with the reconstitution procedure is to maintain the ANT in its native state during purification.  New data will be presented to confirm that binding of CyP-D to the ANT can be increased by oxidation or chemical modification of specific thiol groups on the ANT.  However, activation of pore opening by oxidative stress is not associated with cross-linking of two ANT monomers through disulfide formation between their cysteine 56 residues.  Indeed such cross-linking induced in sub-mitochondrial particles by copper/phenanthrolene can be inhibited by binding of exogenous CyP-D, suggesting that binding may occur at or close to Cys₅₆, perhaps at Pro₆₁ as originally proposed.

[1]  Halestrap, A.P. (1999) In Biochemical Society Symposium no 66 “Mitochondria and Cell Death” (Brown, G.C., Nicholls, D.G. & Cooper, C.E., eds.), Portland Press, London, pp. 181-203 
[2]  Halestrap, A.P. et al. (2000) Biochem. Soc. Trans. 28:170-177

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