Role of phospholipid in the activation of Na +, Ka +-activated adenosine triphosphatase of beef brain

Abstract

Na +, K +-activated ATPase was prepared in soluble form from a membrane fraction of cerebral cortex of beef brain. It is probable that the ATPase has an absolute requirement for phospholipid since it was observed that only in the presence of phospholipid was the ATPase activated by Na + plus K +, and it was inhibited by ouabain. Lecithin protected the enzyme against the effect of the sulfhydryl inhibitor, N-ethylmaleimide, as well as against heat inactivation, but did not modify the effect of ouabain or Na + plus K +. The phospholipid did not affect the rate of either the ATP-ADP 32 exchange reaction or the labeling of the phosphoprotein fraction of the enzyme preparation by ATP 32. The ATPase was inhibited by various sulfhydryl inhibitors including Hg ++, p-mercuribenzoate, Cd ++ and arsenite (with 2,3 dimercaptopropanol). Inhibition by the latter two could be consistent with the view that a dithiol exists in the site concerned with activity. The preparation was found to be contaminated with another type of ATPase, which differed from the Na +, K +-activated ATPase in that the former showed either little or no response to Na + plus K +, phospholipid, sulfhydryl reagents, and heat inactivation.