Half-calcified calmodulin promotes basal activity and inactivation of the L-type calcium channel Ca_V1.2

The L-type Ca²⁺ channel Ca_V1.2 controls gene expression, cardiac contraction, and neuronal activity. Calmodulin (CaM) governs Ca_V1.2 open probability (Po) and Ca²⁺-dependent inactivation (CDI) but the mechanisms remain unclear. Here, we present electrophysiological data that identify a half Ca²⁺-saturated CaM species (Ca₂/CaM) with Ca²⁺ bound solely at the third and fourth EF-hands (EF3 and EF4) under resting Ca²⁺ concentrations (50-100 nM) that constitutively preassociates with Ca_V1.2 to promote Po and CDI. We also present an NMR structure of a complex between the Ca_V1.2 IQ motif (residues 1644-1665) and Ca₂/CaM_12', a calmodulin mutant in which Ca²⁺ binding to EF1 and EF2 is completely disabled. We found that the CaM_12' N-lobe does not interact with the IQ motif. The CaM_12' C-lobe bound two Ca²⁺ ions and formed close contacts with IQ residues I1654 and Y1657. I1654A and Y1657D mutations impaired CaM binding, CDI, and Po, as did disabling Ca²⁺ binding to EF3 and EF4 in the CaM₃₄ mutant when compared to WT CaM. Accordingly, a previously unappreciated Ca₂/CaM species promotes Ca_V1.2 Po and CDI, identifying Ca₂/CaM as an important mediator of Ca signaling.