The plasma membrane calcium pump is the preferred calpain substrate within the erythrocyte

Abstract

The activation of calpain in normal human erythrocytes incubated in the presence of Ca 2+ and the Ca 2+ ionophore A23187 led to the decline of the Ca 2+-dependent ATPase activity of the cells. Preloading of the erythrocyte with an anticalpain antibody prevented the decline. The pump was also inactivated by applied to isolated erythrocyte plasma membranes. The decline of the pump activity corresponded to the degradation of the pump protein and was inversely correlated to the amount of the natural inhibitor of calpain, calpastatin, present in the cells. In erythrocytes containing only 50% of the normal level the degradation started at a concentration of Ca 2+ significantly lower than in normal cells. A comparison of the concentrations of Ca 2+ required for the degradation of a number of erythrocyte membrane proteins showed that the Ca 2+ pump and band 3 were the most sensitive. All other membrane proteins tested were attacked at higher levels of intracellular Ca 2+. Thus, the degradation of the Ca 2+ pump protein may be a simple and sensitive means to monitor calpain activation in vivo. Furthermore, the results have shown that the calpastatin level correlated directly with the amount of activable calpain and with the concentration of Ca 2+ required to trigger the activation process.