Inherited mutations in β-cardiac myosin disrupt the molecular mechanics of contraction and contribute to cardiomyopathies. One such variant, A223T, resides near the nucleotide binding pocket and alters communication pathways linking nucleotide, switch I, and actin-binding elements. These perturbations reduce actin affinity and impair chemo-mechanical transitions during cross-bridge cycling. Small molecules and nucleotides, such as the therapeutic analog 2′-deoxy-ATP (dATP), have emerged as strategies to modulate myosin function; however, their mechanisms of action in the context of pathogenic variants remain unresolved. Here, we combined atomistic molecular dynamics simulations with kinetic assays to examine the mechanistic interplay between A223T and dATP. Strikingly, we found that both A223T and dATP act through overlapping structural communication pathways, yet they produce opposite outcomes. In simulations of pre-powerstroke myosin, A223T disrupted coupling between nucleotide and actin-binding regions to decrease the accessible surface area of the actin binding residues and reducing actin-myosin affinity, while dATP reinforces these connections and enhances acto-myosin affinity. In post-powerstroke simulations, A223T reduced the buried surface area at the actin-myosin interface by 31% and caused a twisting of the myosin motor relative to actin, which is predicted to favor crossbridge dissociation. These findings establish that dATP corrects structural deficiencies introduced by A223T, providing both a mechanistic explanation and a therapeutic rationale for nucleotide-based interventions. Our combination of simulations and experimental validation highlights a shared molecular mechanism that underlies divergent phenotypic outcomes, advancing precision strategies to treat inherited cardiomyopathies.
Wasley et al. (2026) studied this question.