Studies on the properties of the calcium ion pumping ATPase of plasma membranes will be continued. These calcium ion pumps exist in the plasma membranes of heart, smooth muscle, kidney, brain, intestinal epithelium, erythrocyte and many other cell types. They are involved in control of intracellular free calcium levels in cells of all kinds, and in movement of calcium ions across cell layers such as the intestinal and kidney tubule epithelia. Information about the regulation of these pumps may thus give information relevant to hypertension and kidney disease, among other conditions. The pump consists of a single polypeptide chain of molecular weight 127,000-137,000 (the exact molecular weight depends on the isoform). Four genes encode different isoforms, and additional forms are created by alternate splices. We have succeeded in expressing this pump in COS and insect cells and have developed assay systems for determining the activity of the expressed enzyme, its binding of calmodulin and of calcium ions. We propose studies designed to extend our knowledge of the function of the different isoforms of this enzyme, and of the calmodulin-binding and calcium ion-binding residues. We will make measurements of the nitrophenyl phosphatase activity, of the rates of the activation of the pump and of the binding of calmodulin. Measurements will also be made to observe the memory of the pump for its previous history of calcium ion exposure. These measurements will be made on appropriately altered forms of the pump and on the different isoforms.
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