Electron Microscopy Of The Complex Formed By Heavy Meromyosin And C-Protein

Most studies of regulation of vertebrate striated muscle contraction have focussed on activation of the thin filament via calcium binding to the troponin-tropomyosin complex. There is accumulating evidence, however, that thick filament activation is an additional regulatory mechanism. Under relaxing conditions, the two heads of a myosin can interact to form an inactive asymmetric off-state, called the interacting-heads motif (IHM). The off-state inhibits cross-bridge formation and cycling by blocking actin binding in one head and ATPase activity in the other. Docking of the IHM onto the thick filament backbone results in the super-relaxed state (SRX) observed in muscle fibres, where the myosin ATPase is strongly inhibited. What regulates formation and disruption of the IHM is largely uncharacterised. Myosin binding protein C, also known as C-protein, is a key sarcomere component, bound at sites spaced by 43 nm in the cross-bridge regions of thick filaments. The arrangement, mode of action, and interactions of C-protein with myosin are poorly understood. In the heart, phosphorylation of C-protein increases the rate of contraction and its ablation disrupts the SRX. To investigate the interaction of human cardiac C-protein with β-cardiac heavy meromyosin (HMM), we have imaged the complex by electron microscopy and single particle averaging. Cardiac HMM in the IHM motif has the characteristic blocked and inhibited head conformations. The fraction of HMM in the IHM motif is increased when dephosphorylated C-protein is bound, suggesting it stabilises the IHM. Characterization of the complex should provide insight into the regulatory and structural roles of the cardiac C-protein/myosin interaction in heart.