Myotropes unveil two myosin cycles with distinct kinetics and stroke size

Abstract

Cardiac contraction relies on cyclic interactions between myosin II motors and actin filaments, powered by the free energy of ATP turnover. Targeting of this fundamental process has emerged as a promising therapeutic strategy; Mavacamten, a cardiac myosin inhibitor was recently approved for treating obstructive hypertrophic cardiomyopathy whereas omecamtiv mecarbil was developed as a myosin activator for heart failure. Both myotropes bind to the same myosin pocket but their mechanisms of action remain incompletely understood. Here, we investigate these using optical tweezers mechanoenzymology and single-molecule fluorescence-based actomyosin kinetics applied to {beta}-myosin from donor hearts and recombinant human {beta}-myosin subfragment 1. Our findings reveal unexpected layers of complexity. Both mavacamten and omecamtiv mecarbil partially slow the ATP turnover, giving one fast (similar to physiological) and one drug-induced slow ATP turnover cycle at saturating drug concentrations with different rate-limiting steps for the two compounds. In one of the parallel cycles, mavacamten increases stroke size and appreciably delays the power-stroke, allowing direct visualization of a strongly bound pre-power-stroke state. No power-stroke delay is seen in the other cycle, but a reduced stroke size is observed. Our findings uncover a new paradigm in cardiac regulation at the single-molecule level, demonstrating partial inhibition of myosin as a powerful therapeutic strategy.