Disrupted binding dynamics of the CaMD130G mutant to Cav1.2 channels: Implications for long QT syndrome pathogenesis

Abstract

Background: Recent studies have identified a significant association between mutations in Calmodulin (CaM), a pivotal calcium-sensing protein, and the pathogenesis of LQTS. CaM interacts with various cellular proteins, notably the myocardial L-type calcium channel (LTCC), where the Cav1.2 channel serves as the principal subtype in cardiac myocytes. The CaM_D130G mutant is specifically linked to the development of Long QT Syndrome (LQTS). This study aimed to investigate the altered binding of CaM_D130G mutants to Cav1.2 channels, clarifying their role in LQTS pathogenesis.

Methods: Molecular docking techniques were employed to analyze the structural changes at the binding interfaces of CaM_D130G mutants with specific motifs of the Cav1.2 channel, as well as molecular dynamics simulations to verify the reliability of the docking complex. Then, we utilized pull-down experiments to validate the bioinformatic predictions.

Results: Compared to wild-type CaM, the results indicate that the spatial structure of CaM_D130G mutant proteins is significantly altered, and Ca²⁺ ligands have a diminished ability to bind with the C lobe of mutant CaM_D130G proteins. GST pull-down binding assays in vitro demonstrated that CaM binds to the CT1, preIQ, and IQ motifs of the Cav1.2 channel in a Ca²⁺ concentration-dependent and CaM concentration-dependent manner; and this binding was significantly diminished when CaM_D130G was used.

Conclusion: The CaM_D130G mutation disrupts the precise regulation of myocardial Cav1.2 channels, with the CT1 motif being a particularly critical site of interaction. CaM_D130G mutants exhibit a decreased binding affinity for Cav1.2 channel fragments, which may impair Ca²⁺-dependent inactivation (CDI), contributing to the LQTS phenotype.

Keywords: Calmodulin; Calmodulin mutant CaM(D130G); Cardiac L-type calcium channel; Long QT syndrome.