Structural Insights into L-Type Voltage-Gated Ca²⁺ Channel (Ca_V1.2) Activation by CaBP1

The L-type voltage-gated Ca²⁺ channel (Ca_V1.2) controls gene expression, cardiac function, and neuronal excitability. Mutations in Ca_V1.2 that disrupt channel function are implicated in cardiac arrhythmias, vascular dysfunction, Timothy Syndrome, and epilepsy. Calcium-binding protein 1 (CaBP1) binds to the IQ-motif in Ca_V1.2 (residues 1640-1665), blocks Ca²⁺-dependent inactivation (CDI), and promotes Ca²⁺-dependent facilitation (CDF). CaBP1 is 56% identical in sequence to calmodulin (CaM), and both proteins bind competitively to the IQ-motif. Our binding studies reveal that Ca²⁺ binding to CaBP1 is enhanced more than 40-fold when CaBP1 is bound to the IQ peptide. Also, the IQ peptide binds to Ca²⁺-bound CaBP1 (dissociation constant of 45 ± 10 nM) with 100-fold higher affinity than IQ binding to Ca²⁺-free CaBP1. We present NMR structures of Ca²⁺-CaBP1 bound to the IQ peptide, which reveal CaBP1 residues (A107, F111, M128, L131, I144, and M165) that contact IQ residues (I1654, Y1657, and F1658). Also, IQ residue K1662 forms a salt bridge with CaBP1 residue D140, which may explain why a K1662 charge reversal mutation causes 4-fold weaker IQ binding to CaBP1. Electrophysiology studies suggest that CaBP1 acts to increase the Ca_V1.2 channel open probability (Po). We propose that Ca²⁺ binding to the third and fourth EF-hands of CaBP1 and the binding of Ca²⁺-bound CaBP1 to the IQ-motif are important for Ca_V1.2 channel activation.