Using 2'-deoxy-ADP to probe stability of the myosin interacting heads motif at atomic resolution.

Abstract

In addition to active cycling states, myosins within the thick filament can access an ‘inactive’ conformation called the interacting heads motif (IHM) that has been associated with the energy-conserving super relaxed (SRX) state of muscle. Thus, accessing the IHM conformation is a means of thick filament-based regulation of muscle. Only a handful of cryo-EM structures of IHM myosin are available. The large size of the IHM has similarly challenged all-atom molecular simulations of the structure. Using ANTON2, we have performed explicit solvent, all-atom molecular dynamics simulations of human cardiac β-myosin in the IHM confirmation on the microsecond timescale. In recent experimental studies, the small molecule ATP analogue, 2’-deoxy-ATP (dATP), has been shown to destabilize myosin heads from the IHM into disordered, more active states. To complement these experimental studies, we simulated β-myosin in the IHM conformation in which ADP.Pi was replaced by dADP.Pi. These simulations showed that dADP reduced the stability of the IHM by reducing the number of interactions between S1 heads. They also show that the tails of dADP.Pi-bound heads adopt conformations distinct from existing atomic models obtained with cryoEM. Thus, simulations suggest that dynamics in the RLC-binding region of the tail and at the head-head interface both contribute to IHM stability. Further, they suggest that departure from the IHM state involves coordinated motions in regions of myosin separated by over 100 Å. Ongoing analysis of the degree to which altered dynamics within the nucleotide binding pocket influence the conformation of both the head interface and RLC-binding region of the tail. These novel simulations should also prompt further research into the contribution of tail dynamics into IHM stability as well as interest in mutations that influence tail dynamics.