The L348P gain-of-function mutation in the M-domain of cardiac myosin-binding protein-C significantly increased its binding affinity for actin and enhanced force activation independent of calcium.
The tri-helix bundle of the cMyBP-C M-domain is crucial for regulating cardiac muscle contraction via actin binding, and its mutation can lead to functional changes associated with hypertrophic cardiomyopathy.
Tasa de eventos absoluta: 3.2% vs 7.4%
valor p: p=<0.05
The M-domain is the major regulatory subunit of cardiac myosin-binding protein-C (cMyBP-C) that modulates actin and myosin interactions to influence muscle contraction. However, the precise mechanism(s) and the specific residues involved in mediating the functional effects of the M-domain are not fully understood. Positively charged residues adjacent to phosphorylation sites in the M-domain are thought to be critical for effects of cMyBP-C on cross-bridge interactions by mediating electrostatic binding with myosin S2 and/or actin. However, recent structural studies revealed that highly conserved sequences downstream of the phosphorylation sites form a compact tri-helix bundle. Here we used site-directed mutagenesis to probe the functional significance of charged residues adjacent to the phosphorylation sites and conserved residues within the tri-helix bundle. Results confirm that charged residues adjacent to phosphorylation sites and residues within the tri-helix bundle are important for mediating effects of the M-domain on contraction. In addition, four missense variants within the tri-helix bundle that are associated with human hypertrophic cardiomyopathy caused either loss-of-function or gain-of-function effects on force. Importantly, the effects of the gain-of-function variant, L348P, increased the affinity of the M-domain for actin. Together, results demonstrate that functional effects of the M-domain are not due solely to interactions with charged residues near phosphorylatable serines and provide the first demonstration that the tri-helix bundle contributes to the functional effects of the M-domain, most likely by binding to actin. Background: Cardiac myosin-binding protein C (cMyBP-C) regulates heart muscle contraction by influencing actomyosin interactions. Results: Amino acids within the tri-helix bundle of the M-domain contribute to the functional effects of cMyBP-C. Conclusion: Amino acids outside of phosphorylation sites influence function of the M-domain. Significance: The tri-helix bundle is important to the regulatory role of cMyBP-C, likely through actin-binding interactions.
Bezold et al. (Thu,) conducted a other in Hypertrophic cardiomyopathy (context). L348P mutation in the M-domain of cMyBP-C vs. Wild-type C1C2 protein was evaluated on Actin binding affinity (Kd in μM) (p=<0.05). The L348P gain-of-function mutation in the M-domain of cardiac myosin-binding protein-C significantly increased its binding affinity for actin and enhanced force activation independent of calcium.