Abstract
In calmodulin-stripped inside-out human red cell membrane vesicles /IOV/ ATP + Mg 2+-dependent active calcium uptake is stimulated by the addition of calmodulin. Calmodulin increases the maximum calcium transport rate /V max/, decreases K Ca, and does not affect K ATP of calcium uptake. The action of both membrane bound and external calmodulin is competitively inhibited by phenothiazines. Drugs reacting with SH groups of proteins reversibly inhibit calcium pumping by decreasing V max and not affecting K Ca and K ATP. The relative magnitude of calmodulin stimulation of calcium transport is unaltered by SH reagents. Mild proteolytic digestion of IOVs stimulates active calcium uptake and mimics the effects of calmodulin on the kinetic parameters — that is converts the system to a “high calcium-affinity” state. Proteolysis eliminates calcium-dependent calmodulin binding to IOV membranes and any further stimulation of calcium uptake by calmodulin. Based on these results the presence of a calmodulin-binding regulatory subunit of the red cell calcium pump at the internal membrane surface is postulated.
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