2,4-DINITROPHENOL AND CARBONYLCYANIDE P-TRIFLUOROMETHOXYPHENYLHYDRAZONE ACTIVATE THE GLUTATHIONE S-CONJUGATE TRANSPORT ATPASE OF HUMAN ERYTHROCYTE-MEMBRANES

2,4-Dinitrophenol (DNPOH) and carbonylcyanide p-triffuoromethoxyphenyl hydrazone (FCCP), two classical uncouplers of mitochondrial oxidative phosphorylation, were found to stimulate human erythrocyte membrane ve sicle ATPase activity. Both compounds competed with S-(2,4-dinitrophen yl) glutathione (DNPSG) for activation of the glutathione S-conjugate transport ATPase. Stimulation of the ATPase by DNPOH or FCCP occurred with V-max values 4-6 times greater than that with DNPSG. The K-0.5 fo r DNPOH (195 mu M) was similar to that of DNPSG (196 mu M), while that for FCCP (4.3 mu M) was 40 times lower. Vanadate inhibits both the DN POH- and FCCP-stimulated ATPase activities, as previously reported for the glutathione S-conjugate ATPase. The stimulation of erythrocyte ve sicle ATPase activities by these classical uncoupling agents does not result from increased proton conductance across the vesicle membrane: monensin, gramicidin and nystatin, all of which increase proton conduc tance, but by different mechanisms, do not stimulate erythrocyte vesic le ATPase activity. Verapamil, a known P-glycoprotein ATPase activator also does not stimulate human erythrocyte membrane ATPase activity. T hese results show that relatively small, monoanionic lipophilic compou nds such as DNPOH and FCCP can activate the glutathione S-conjugate tr ansport ATPase. The higher V-max values for activation by these agents than by DNPSG make possible a more sensitive assay of this transport ATPase activity. The results raise the question of whether these subst ances and other small anionic, lipophilic compounds are also transport ed by this system. (C) 1994 Academic Press, Inc.