Studies on the purified, lipid-reconstituted (Na+ + Mg2+)-ATPase from Acholeplasma laidlawii B membranes. Dependence of enzyme activity on lipid headgroup and hydrocarbon chain structure.

The purified (Na+ + Mg2+)-ATPase from Acholeplasma laidlawii B membranes was successfully reconstituted with a number of different phospho- and glycolipids, and the ability of these lipids to support the function of this enzyme was evaluated by their ability to increase the specific activity of the purified enzyme and by their ability to restore its lipid-phase state-dependent properties which were lost during purification. The incorporation of this ATPase into liposomes composed of the endogenous membrane lipids of the organism, or of zwitterionic phospholipids such as phosphatidylcholine or phosphatidylethanolamine, results in a full reconstitution of its activity and its lipid-phase state-dependent properties. In contrast, anionic phospholipids alone, or in combination with zwitterionic phospholipids at concentrations higher than 10 mol % of the anionic phospholipid, cause an irreversible inhibition of this ATPase. However, when combined with neutral glycolipids, larger amounts of anionic phospholipid can be tolerated without enzyme inhibition. Phosphatidylcholines with acyl chains of 14-24 linear carbon atoms and varying degrees of branching and unsaturation successfully reconstitute the enzyme, in marked contrast to the shorter chain homologues, which were ineffective. Our results indicate that the full expression of the activity of the A. laidlawii B ATPase requires a host lipid bilayer membrane of low to moderate negative surface charge which is predominantly liquid-crystalline and of a minimal bilayer thickness. Once such requirements are met, the enzyme exhibits considerable flexibility regarding the nature of the lipids which can effectively support its function. In particular, the activity of the A. laidlawii B ATPase is not very sensitive to lipid "fluidity" in the liquid-crystalline state.