Properties of (Mg2 + Ca2+)-ATPase of erythrocyte membranes prepared by different procedures: influence of Mg2+, Ca2+, ATP, and protein activator.
Erythrocyte membranes prepared by three different procedures showed (Mg2+ + Ca2+)-ATPase activities differing in specific activity and in affinity for Ca2+. The (Mg2+ + Ca2+)-ATPase activity of the three preparations was stimulated to different extents by a Ca2+-dependent protein activator isolated from hemolysates. The Ca2+ affinity of the two most active preparations was decreased as the ATP concentration in the assay medium was increased. Lowering the ATP concentration from 2 mM to 2-200 microM or lowering the Mg:ATP ratio to less than one shifted the (Mg2+ + Ca2+)-ATPase activity in stepwise hemolysis membranes from mixed "high" and "low" affinity to a single high Ca2+ affinity. Membranes from which soluble proteins were extracted by EDTA (0.1 mM) in low ionic strength, or membranes prepared by the EDTA (1-10 mM) procedure, did not undergo the shift in the Ca2+ affinity with changes in ATP and MgCl2 concentrations. The EDTA-wash membranes were only weakly activated by the protein activator. It is suggested that the differences in properties of the (Mg2+ + Ca2+)-ATPase prepared by these three procedures reflect differences determined in part by the degree of association of the membrane with a soluble protein activator and changes in the state of the enzyme to a less activatable form.
D000251:Adenosine Triphosphatases / Q000097:blood* / Q000302:isolation & purification; D000255:Adenosine Triphosphate / Q000494:pharmacology; D001798:Blood Proteins / Q000502:physiology*; D002118:Calcium / Q000494:pharmacology*; D004789:Enzyme Activation; D004910:Erythrocyte Membrane / Q000201:enzymology*; D004912:Erythrocytes / Q000201:enzymology*; D006801:Humans; D007700:Kinetics; D008274:Magnesium / Q000494:pharmacology*
D001798:Blood Proteins; D000255:Adenosine Triphosphate; D000251:Adenosine Triphosphatases; D008274:Magnesium; D002118:Calcium