Changes in cardiac myosin acetylation disrupt the super-relaxed state in genotype-negative hypertrophic cardiomyopathy with type 2 diabetes

Patients with hypertrophic cardiomyopathy (HCM) and type 2 diabetes (T2D) have a more severe cardiac phenotype and worse clinical course than non‑diabetic patients. To identify how T2D aggravates the disease and whether the most abundant cardiac protein, myosin, is involved, we combined functional, structural and mass spectrometry analyses of human samples. Left ventricular septal myectomy samples from genotype‑negative (G−) HCM patients without T2D (G− , N = 19) and with T2D (G−T2D, N = 15) were analyzed mainly using fluorescent ATP chase experiments, small‑angle X‑ray diffraction and targeted myosin heavy chain proteomics. Mant‑ATP chase measurements showed a lower fraction of myosin heads in the energy‑conserving super‑relaxed (SRX) state in G−T2D compared to non-diabetic myocardium. In parallel, X‑ray diffraction showed trends toward structural alterations in myosin organization in G−T2D tissue, consistent with altered OFF/ON state equilibrium. Targeted mass spectrometry identified hyperacetylation of several myosin lysine residues in G−T2D, including K847 within the S2 region. All‑atom molecular dynamics simulations indicated that K847 acetylation disrupts stabilizing electrostatic interactions in the interacting‑heads motif, which is associated with the OFF state. Disruption of myosin super‑relaxation emerges as a central cellular defect in G−T2D HCM myocardium and can be mechanistically linked to site‑specific myosin hyperacetylation at K847, providing a potential therapeutic target for genotype‑negative HCM with T2D.