title: Irreversible effects of calcium ions on the plasma membrane calcium pump.
abstract: The calcium pump of human red cells can be irreversibly activated by preincubation of the membranes in the presence of calcium ions, with a pattern reminiscent of that produced by controlled trypsin attack. With 1 mM Ca2+, the activity of the basal enzyme increases three to fourfold over 30 to 60 min, to levels about half those obtained in the presence of calmodulin. On the whole, the effect occurs slowly, with a very low Ca2+ affinity at 37 degrees C and is unaffected by serine-protease inhibitors. The activation caused by 1 mM Ca2+ is little affected by leupeptin (a thiol-protease inhibitor) and that obtained at 10 microM Ca2+ is not inhibited. Preincubations at 0 degrees C also lead to activation, to a level up to half that seen at 37 degrees C, and the effect is not affected by leupeptin or antipain. No activation is observed by preincubating soluble purified Ca,Mg-ATPase in Ca(2+)-containing solutions at 37 degrees C. Instead, calcium ions protect the detergent-solubilized enzyme from thermal inactivation, the effect being half-maximal between 10 and 20 microM Ca2+. We conclude that the activation of the membrane-bound Ca,Mg-ATPase by Ca2+ should result from an irreversible conformational change in the enzyme and not from attack by a membrane-bound protease, and that this change presumably arises from the release of inhibitory particles existing in the original membrane preparations.
authors.affiliation: Department of Cell Physiology and Pharmacology, Leicester University, England.
authors.forename: D G
authors.initials: DG
authors.lastname: Ward
authors.forename: T J
authors.initials: TJ
authors.lastname: Walton
authors.forename: J D
authors.initials: JD
authors.lastname: Cavieres
mesh_terms: D001665:Binding Sites; D017301:Ca(2+) Mg(2+)-ATPase / Q000037:antagonists & inhibitors / Q000378:metabolism; D002118:Calcium / Q000378:metabolism / Q000494:pharmacology*; D000252:Calcium-Transporting ATPases / Q000187:drug effects* / Q000378:metabolism; D004789:Enzyme Activation / Q000187:drug effects; D004910:Erythrocyte Membrane / Q000187:drug effects* / Q000378:metabolism*; D006801:Humans; D066298:In Vitro Techniques; D007700:Kinetics; D015842:Serine Proteinase Inhibitors / Q000494:pharmacology
chemical_list: D015842:Serine Proteinase Inhibitors; D017301:Ca(2+) Mg(2+)-ATPase; D000252:Calcium-Transporting ATPases; D002118:Calcium