Effects of calcium and soluble cytoplasmic activator protein (calmodulin) on various states of (Ca2+ + Mg2+)-ATPase activity in isolated membranes of human red cells.
(Ca2+ + Mg2+)-ATPase activity of red cells and their isolated membranes was investigated in the presence of various Ca2+ concentrations and cytoplasmic activator protein. Red cell ATPase activity was high at low Ca2+ concentrations, and low at moderate and high concentrations of Ca2+. In the case of isolated membranes, both low and moderate ca2+ concentrations produced higher (Ca2+ + Mg2+)-ATPase activity than high Ca2+ concentration. Membrane-free hemolysate containing soluble activator of (Ca2+ + Mg2+)-ATPase produced a significant increase in (Ca2+ + Mg2+)-ATPase activity only at low ca2+ concentration. Regardless of Ca2+ and activator concentrations, the enzyme activity in the membrane was lower than lysed red cells. The low level of (Ca2+ + Mg2+)-ATPase activity seen at high Ca2+ concentration can be augmented by lowering the Ca2+ concentration of EGTA in the assay medium. However, once the membrane was exposed to a high Ca2+ concentration, the activator could no longer exert it maximum stimulation at the low Ca2+ concentration brought about by addition of EGTA. This loss of activation was not attributable to the Ca2+-induced denaturation of activator protein but rather related to the alteration of (Ca2+ + Mg2+)-ATPase states in the membrane. On the basis of these data, it is suggested that only a small portion of (Ca2+ + Mg2+)-ATPase activity of isolated membranes can be stimulated by the soluble activator and that (ca2+ + Mg2+)ATPase most likely exists in various states depending upon ca2+ concentration and the presence of activator. The enzyme state exhibiting the high degree of stimulation by activator may undergo irreversible damage in the presence of high Ca2+ concentrations.
D017301:Ca(2+) Mg(2+)-ATPase; D002118:Calcium / Q000494:pharmacology*; D002135:Calcium-Binding Proteins / Q000097:blood*; D000252:Calcium-Transporting ATPases / Q000097:blood*; D002147:Calmodulin / Q000097:blood*; D004533:Egtazic Acid / Q000494:pharmacology; D004789:Enzyme Activation; D004910:Erythrocyte Membrane / Q000201:enzymology*; D004912:Erythrocytes / Q000201:enzymology*; D006461:Hemolysis; D006801:Humans
D002135:Calcium-Binding Proteins; D002147:Calmodulin; D004533:Egtazic Acid; D017301:Ca(2+) Mg(2+)-ATPase; D000252:Calcium-Transporting ATPases; D002118:Calcium