Evidence for zinc function in the Mg ++/Ca ++ stimulated ATPase of escherichiacoli membranes

Abstract

Summary At low Mg ++ concentration the soluble Mg ++ /Ca ++ activated ATPase extracted from E. coli membranes is inhibited by several chelators including bathophenanthroline sulfonate, diphenylthiocarbazone and zincon. Incubation of the enzyme with zincon shows a gradual extraction of zinc. Activity is lost in proportion to zinc removal and is specifically restored by readdition of zinc to the extracted enzyme. The soluble enzyme in presence of high concentration of Mg ++ is not inhibited by chelators. The presence of high Mg ++ concentration also prevents reconstitution of the zinc depleted enzyme activity with zinc. In the membrane bound form the activity is not inhibited by zincon or other water soluble chelators regradless of Mg ++ concentration whereas the activity is inhibited by various lipophilic chelators such as thenoyltrifluoroacetone and tert-octyl catechol. These latter chelators do not inhibit the soluble ATPase. We conclude that the ATPase contains zinc which is protected from chelators by high Mg ++ concentration or by binding to the membrane and that there is another lipophilic chelator site in the membrane which also controls activity of the membrane-bound ATPase.