The Ca(2+)-pumping activity of constructs containing various portions of the putative 28-residue calmodulin-binding domain (C domain) of hPMCA4b were compared. As the length of the C domain in the pump increased, the pump's activity decreased and the ability of calmodulin to stimulate the activity increased. Study of the calmodulin dependence of activity showed that the construct containing all 28 residues of the C domain had a K1/2 for calmodulin equal to that of the complete molecule; the constructs containing less of the C domain interacted less strongly with calmodulin. On the other hand, incorporation of all 28 residues of the C domain did not decrease the activity of the pump (in the absence of calmodulin) as low as the activity of the complete molecule. This indicates that other segments of the molecule, further toward the COOH terminus, are also required for the degree of inhibition seen in the complete molecule.