Significance Heart disease is the leading cause of death worldwide, and hypertrophic cardiomyopathy (HCM) is the most common inherited form of heart disease, affecting over 1 in 200 people. Mutations in myosin, the motor protein responsible for contraction of the heart, are a common cause of HCM but have diverse effects on the biomechanics of the myosin protein. We demonstrate that complex biomechanical effects of mutations associated with heart disease can be effectively studied and understood using a multiscale experimental and computational modeling approach. This work confirms an important role for disruption of the super relaxed state for one particular HCM mutation and provides an approach that can be extended to aid in the understanding of disease mechanisms for different mutations.