Lipid requirement of membrane-bound ATPase. Studies on human erythrocyte ghosts.

Abstract

The (Na++ K+)‐stimulated ATPase of human erythrocyte ghosts is completely inactivated by treatment with pure phospholipase A2 or phospholipase C. These phospholipases also cause considerable or even complete loss of Mg2+‐dependent ATPase activity. Enzymatic hydrolysis of sphingomyelin does not significantly influence either of the ATPase activities. Conversion of the glycero‐phospholipids into phosphatidic acid by phospholipase D causes a drastic decrease in the Mg2+‐dependent ATPase activity, whereas the (Na++ K+)‐stimulated activity is increased. Enzymatic decarboxylation of the phosphatidylserine fraction of the ghosts does not influence the (Na++ K+)‐stimulated ATPase, unless the last 13% of this fraction is converted into phosphatidyethanolamine, when there is a complete loss of this particular ATPase activity. On the other hand, the Mg2+‐dependent activity tends to be increased by this treatment. The (Na++ K+)‐stimulated ATPase activity of erythrocyte ghosts treated with phospholipase C and subsequently extracted with dry ether, can be completely reconstituted by the addition of phosphatidylserine. Phosphatidylcholine, phosphatidylethanolamine and phospholipid mixtures are found to be ineffective. Phosphatidic acid produced a partial restoration of activity. It is concluded that the (Na++ K+)‐stimulated ATPase activity strongly depends upon the presence of phosphatidylserine, of which only a minor fraction in the erythrocyte membrane is directly involved. It cannot be concluded from the experiments described in this paper that the Mg2+‐dependent ATPase also requires a specific phospholipid. However, a direct involvement of either cholesterol or sphingomyelin in this activity can be precluded.