RADICAL-INDUCED INACTIVATION OF KIDNEY NA+,K+-ATPASE - SENSITIVITY TO MEMBRANE LIPID-PEROXIDATION AND THE PROTECTIVE EFFECT OF VITAMIN-E

The Na+,K+-ATPase is a membrane-bound, sulfhydryl-containing protein whose activity is critical to maintenance of cell viability. The susceptibility of the enzyme to radical-induced membrane lipid peroxidation was determined following incorporation of a purified Na+,K+-ATPase into soybean phosphatidylcholine liposomes. Treatment of liposomes with Fenton's reagent ( Fe2+ H2O2) resulted in malondialdehyde formation and total loss of Na+,K+-ATPase activity. At 150 μM Fe2+ 75 μM H2O2, vitamin E (5 mol%) totally prevented lipid peroxidation but not the loss of enzyme activity. Lipid peroxidation initiated by 25 μM Fe2+ 12.5 μM H2O2 led to a loss of Na+,K+-ATPase activity, however, vitamin E (1.2 mol%) prevented both malondialdehyde formation and loss of enzyme activity. In the absence of liposomes, there was complete loss of Na+,K+-ATPase activity in the presence of 150 μM Fe2+ 75 μM H2O2, but little effect by 25 μM Fe2+ 12.5 μM H2O2. The activity of the enzyme was also highly sensitive to radicals generated by the reaction of Fe2+ with cumene hydroperoxide, t-butylhydroperoxide, and linoleic acid hydroperoxide. Lipid peroxidation initiated by 150 μM Fe2+ 150 μM Fe3+, an oxidant which may be generated by the Fenton's reaction, inactivated the enzyme. In this system, inhibition of malondialdehyde formation by vitamin E prevented loss of Na+,K+-ATPase activity. These data demonstrate the susceptibility of the Na+,K+-ATPase to radicals produced during lipid peroxidation and indicate that the ability of vitamin E to prevent loss of enzyme activity is highly dependent upon both the nature and the concentration of the initiating and propagating radical species. © 1990.