Key result
MYBPC3 N-terminal missense mutations enhance actin interactions and destabilize protein structure, contributing to HCM pathogenesis.
Why the study?
Although most cMyBP-C mutations cause HCM through haploinsufficiency from truncated proteins, the mechanisms by which missense mutations cause disease remain poorly understood.
MYBPC3 missense mutations linked to hypertrophic cardiomyopathy enhance actin binding and reduce protein thermal stability, providing mechanistic insights into HCM pathogenesis.
May inform HCM mechanisms via MYBPC3-actin effects; leaves open human validation and therapeutic targeting.
Mutations in cardiac myosin-binding protein C (cMyBP-C) are a leading cause of hypertrophic cardiomyopathy (HCM). Although most cMyBP-C mutations produce truncated proteins and cause HCM via haploinsufficiency, the mechanisms by which missense mutations result in disease remain poorly understood. Here, we have evaluated three mutations in immunoglobulin-like domains C1 (P161S, Y237S) and C2 (P371R), predicted to be pathogenic for HCM, assessing their effects on cMyBP-C actin-binding function, protein thermal stability, and residue mobility. Using a fluorescence lifetime-based actin-binding assay, we found that N-terminal mutants P161S, Y237S, and P371R enhanced C0-C2 interactions with actin in both unphosphorylated and phosphorylated states, suggesting that the mutations strengthen actin binding and make the binding resistant to phosphorylation-mediated regulation. Differential scanning calorimetry revealed that mutants exhibit destabilized thermal melting profiles with reduced unfolding temperature, energy, and cooperativity. Molecular dynamics simulations indicated that these mutations induce allosteric effects, increasing fluctuations of unstructured loops in C1 or C2 that contain key actin-binding residues. These alterations in protein stability and residue mobility may promote domains to visit binding-competent conformations more frequently, reduce the energetic cost of complex formation, and/or expose actin-interacting interfaces, thereby enhancing C0-C2 binding and contributing to HCM pathogenesis.
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Kanassatega et al. (2025) studied Hypertrophic cardiomyopathy. MYBPC3 N-terminal missense mutations (P161S, Y237S, P371R) vs. Wild-type cMyBP-C was evaluated on cMyBP-C actin-binding function, protein thermal stability, and residue mobility. MYBPC3 N-terminal missense mutations (P161S, Y237S, P371R) enhanced C0-C2 interactions with actin and destabilized thermal melting profiles, contributing to hypertrophic cardiomyopathy pathogenesis.
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