Effects of Hypertrophic Cardiomyopathy Causing R403Q Mutation on Human Beta-Cardiac Myosin Biomechanical Function: Single Molecule to Ensemble Studies

Abstract

Over 300 known mutations in human beta-cardiac myosin, the motor that powers ventricular contraction, causes familial hypertrophic cardiomyopathy (HCM) by altering the functional capacity of the cardiac sarcomere. The precise mechanism(s) by which these point mutations alter cardiac function and lead to the clinically diverse HCM phenotypes is not well understood. The human heart is optimally designed to cyclically contract under varying loads and it is likely that the mutations in beta-cardiac myosin affect the power output of the heart by either altering force, velocity of contraction, or both, which ultimately results in the observed clinical phenotypes.Using ensemble in vitro motility experiments and single molecule force spectroscopy, we show that a truncated version (1-808 aa, containing human ELC) of human beta-cardiac HCM-causing R403Q myosin mutant has ∼15% lower unitary force generated by a single head but has ∼15% higher unloaded velocity of contraction with pure actin. However, when studied with the six-component regulatory thin filament (RTF) system (containing tropomyosin and the troponin complex) the unloaded velocity of R403Q is unchanged. Loaded in vitro motility measurements with both pure actin and RTFs indicate lower ensemble force and therefore possibly lower power generated by this molecule. Additional data suggests that R403Q myosin may have lower affinity for RTFs than WT myosin, signifying a change in the duty ratio. Current efforts exploring the effects of the R403Q mutation with a two-headed HMM construct are underway.