Modulation of human erythrocyte Ca²⁺-ATPase activity by glycerol: The role of calmodulin

The effect of an intracellular cryoprotectant glycerol on human erythrocyte Ca²⁺-ATPase activity and possible involvement of calmodulin in the regulation of Ca²⁺-pump under these conditions were investigated. The experiments were carried out using saponin-permeabilized cells and isolated erythrocyte membrane fractions (white ghosts). Addition of rather low concentrations of glycerol to the medium increased Ca²⁺-ATPase activity in the saponin-permeabilized cells; the maximal effect was observed at 10% glycerol. Subsequent increase in glycerol concentrations above 20% was accompanied by inhibition of Ca²⁺-ATPase activity. Lack of stimulating effect of glycerol on white ghost Ca²⁺-ATPase may be attributed to removal of endogenous compounds regulating activity of this ion transport system. Inhibitory analysis using R24571 revealed that activation of Ca²⁺-ATPase by 10% glycerol was observed only in the case of inhibitor administration after modification of cells with glycerol; in the case of inhibitor addition before erythrocyte contact with glycerol, this phenomenon disappeared. These data suggest the possibility of regulation of human erythrocyte Ca²⁺-ATPase by glycerol; this regulatory effect may be attributed to both glycerol-induced structural changes in the membrane and also involvement of calmodulin in modulation of catalytic activity of the Ca ²⁺-pump.