Ca2+-ATPase from chicken (Gallus domesticus) erthrocyte plasma membrane: effects of calmodulin and taurine on the Ca2+-dependent ATPase activity and Ca2+ uptake

In the present report we describe a method for purifying plasma membranes f rom chicken erythrocytes using sonication under conditions that facilitate preferential lysis of plasma membrane, followed by centrifugation through a sucrose gradient. The Ca2+-dependent ATP hydrolysis by plasma membranes is activated by nanomolar levels of calmodulin, similarly to that from anucle ated erythrocytes. Inside-out vesicles display a calmodulin-activated Ca2uptake. Purified Ca2+-ATPase is obtained from the plasma membrane by Sephar ose-calmodulin affinity chromatography, and exhibits an apparent molecular mass of 150 kDa on SDS-polyacrylamide gel electrophoresis, clearly showing that the enzyme is distinct from that described in anucleated erythrocytes (140 kDa). The enzyme is insensitive to physiological concentrations of tau rine, a beta-amino acid that has been proposed to be involved in Ca2+ homeo stasis of nucleated erythrocytes, suggesting that the effect of taurine is not mediated by the Ca2+-ATPase. Taken together, these data suggest that th e enzyme may be an isoform that resembles the previously described plasma m embrane Ca2+-ATPase from anucleated erythrocytes in its regulation by calmo dulin, but differs in its apparent molecular weight. (C) 1999 Elsevier Scie nce Inc. All rights reserved.