Sequence differences in the IQ motifs of Cav1.1 and Cav1.2 strongly impact calmodulin binding and calcium-dependent inactivation

The proximal C terminus of the cardiac L-type calcium channel (CaV1.2) contains structural elements important for the binding of calmodulin (CaM) and calcium-dependent inactivation, and exhibits extensive sequence conservation with the corresponding region of the skeletal L-type channel (CaV1.1). However, there are several CaV1.1 residues that are both identical in six species and are non-conservatively changed from the corresponding CaV1.2 residues, including three of the “IQ motif.” To investigate the functional significance of these residue differences, we used native gel electrophoresis and expression in intact myotubes to compare the binding of CaM to extended regions (up to 300 residues) of the C termini of CaV1.1 and CaV1.2. We found that in the presence of Ca 2 (either millimolar or that in resting myotubes), CaM bound strongly to C termini of CaV1.2 but not of CaV1.1. Furthermore, replacement of two residues (Tyr 1657 and Lys 1662 ) within the IQ motif of a C-terminal CaV1.2 construct with the divergent residues of CaV1.1 (His 1532 and Met 1537 ) led to a weakening of CaM binding (native gels), whereas the reciprocal substitution in CaV1.1 caused a gain of CaM binding. In full-length CaV1.2, substitution of these same two divergent residues with those of CaV1.1 (Y1657H, K1662M) eliminated calcium-dependent inactivation of the heterologously expressed channel. Thus, our results reveal that a conserved difference between the IQ motifs of CaV1.2 and CaV1.1 has a profound effect on both CaM binding and calciumdependent inactivation.