Modulation of striated-muscle contractility by a high-affinity myosin-targeting peptide

Myosin-based regulation has emerged as a fundamental new concept governing both cardiac and skeletal muscle contractile function during both health and disease states. Myosin-targeted therapeutics have the potential to treat heart failure with either systolic or diastolic dysfunction based on either activating or inhibiting the function of myosin. In this study, we developed a striated-muscle myosin-specific high-affinity peptide that targeted the proximal subfragment 2 (S2) region of the MYH7 myosin, which has been shown to undergo conformational changes associated with force generation by the myosin head domains. We characterized the peptide called Stabilizer using a wide range of biochemical, biophysical, and physiological methods, creating a multi-scale structure-activity relationship ranging from single-molecule assays to contractile measurements in intact cardiac muscle cells. The Stabilizer binds myosin S2 with low nanomolar affinity and strongly increases its mechanical stability as measured by single-molecule gravitational force spectroscopy and Förster resonance energy transfer measurements. The Stabilizer significantly inhibits myofibrilar contractility and ATPase activity, and it reduces myosin crossbridge kinetics in demembranated cardiac muscle cells. Biochemical modification of the Stabilizer further allowed measurements in intact porcine cardiomyocytes showing decreased contraction and relaxation kinetics in the presence of the peptide. Our results show that myosin S2-targeting peptides are biologicals with potential therapeutic applications for muscle diseases.