Studies on incorporation of membrane protein into liposomes--effect of divalent cations on reconstitution of pig heart mitochondrial H+-ATPase into liposomes.

Abstract

During the process of reconstitution of pig heart mitochondrial H+-ATPase on liposomes by cholate dialysis method, 1 mM Mg2+ in the dialysis medium can obviously increase activities of the reconstituted H+-ATPase (32Pi-ATP exchange, enzyme activity and its sensitivity to oligomycin or DCCD and ATP-dependent ANS 1) fluorescence). Besides Mg2+, effects of other divalent cations and spermidine on the H+-ATPase reconstitution have been compared. The effectiveness of the divalent cation activation of 32Pi-ATP exchange of the reconstituted H+-ATPase, decreased in the order: Mg2+ greater than Ca2+ greater than Mn2+ greater than Sr2+, while that of increasing the sensitivity to the inhibitors: Ca2+ greater than Mg2+ greater than Mn2+ greater than Sr2+. No significant effect on the H+-ATPase reconstitution has been found with Cd2+, Zn2+ and spermidine3+. Mechanism of action of the divalent cations on reconstitution has been discussed.