Impact of Regulatory Light Chain Mutation (K104E) on the Atpase and Motor Properties of Human Cardiac Myosin

Abstract

Mutations in the cardiac myosin heavy chain (MYH7) as well as the essential (MYL3) and regulatory (MYL2) light chains are known to lead to cardiomyopathies with variable phenotypes, including hypertrophic (HCM) and dilated (DCM) cardiomyopathy. We investigated the impact of the HCM-associated regulatory light chain (RLC) mutation (K104E) on myosin ATPase kinetics and in vitro motility. We expressed and purified human cardiac myosin subfragment 1 (M2B-S1) and exchanged on the human ventricular RLC, with the mutant or WT RLC. A mouse model of K104E was found to cause cardiac hypertrophy, fibrosis, and diastolic dysfunction in older animals suggesting a slow development of HCM, while variable penetrance of the mutation in human populations suggest the impact of K104E may be subtle. We found that maximum actin-activated ATPase activity and the actin-dependence of ATPase were similar in K104E and WT. Actin gliding velocities in the in vitro motility assay were slightly increased in K104E. We found that the ADP release rate constant in the presence of actin also was slightly increased in K104E compared to WT. Our results suggest that HCM-associated RLC mutations may only have a minor impact on unloaded ATPase kinetics and in vitro actin gliding, which may suggest other factors could be altered by the mutation such as mechanical load and myosin regulation. Future experiments will address the impact of the mutation in mouse cardiac vs. human cardiac myosin as well as the impact of the mutation on load-dependent motility and the off state (super-relaxed state) of cardiac myosin.