Cardiac Myosin binding Protein-C: Molecular Mechanisms of Actomyosin Modulation

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DESCRIPTION (provided by applicant): Cardiac Myosin Binding Protein-C (cMyBP-C) is critical to normal cardiac performance as evidenced by genetic mutations in cMyBP-C being one of the leading causes of familial hypertrophic cardiomyopathy. Despite its functional importance, the molecular mechanism by which cMyBP-C exerts its effect on the myosin molecular motor as it interacts with actin to generate force and motion remains largely undefined. A yet unanswered question is with its low ratio relative to myosin and it being located in distinct regions of the thick filament, we will determine how cMyBP-C's modulates actomyosin's power generation by interacting with only a limited population of crossbridges within the thick filament. To address this in Aim #1, we will use state-of- the-art single molecule biophysical techniques (e.g. laser trap assay) to probe the effect that cMyBP-C exerts on actomyosin function along the length a single native thick filament. In Aim #2, we will use expressed N-terminal fragments of cMyBP-C to probe the binding affinity of these fragments for actin, the regulated thin filament, and/or myosin. In combination with motility and laser trap assays, we will determine if the N- terminus of CMyBP-C limits myosin's attachment rate to the thin filament or if it directly affects myosin's inherent molecular mechanics and kinetics. Finally, in Aim #3 we will characterize how phosphorylation regulates cMyBP-C action. Using existing transgenic mouse models expressing cMyBP C mutants having alanine or aspartic acid substitutions for all three phosphorylatable serines, we will determine the functional importance of phosphorylation using native thick filaments containing mutant cMyBP-C as well as N-terminal fragments having the same mutations. Once the molecular mechanism of cMyBP-C is defined, the potential for novel therapeutics or clinical intervention may be possible in cases of heart failure associated with genetic mutations in cMyBP-C. PUBLIC HEALTH RELEVANCE Cardiac Myosin Binding Protein-C (cMyBP-C) is critical to normal cardiac performance as evidenced by genetic mutations in cMyBP-C being one of the leading causes of familial hypertrophic cardiomyopathy. Despite its functional importance, the molecular mechanism by which cMyBP-C exerts its effect on the myosin molecular motor as it interacts with actin to generate force and motion remains largely undefined.