Journal Article

Dilated cardiomyopathy mutation E525K in human beta-cardiac myosin stabilizes the interacting-heads motif and super-relaxed state of myosin

Desetty, Rohini, Sadler, Fredrik R, Sivaramakrishnan, Sivaraj, Yengo, Christopher M, Rasicci, David V, Craig, Roger, Bodt, Skylar ML, Tiwari, Prince

2022

Overview

The auto-inhibited, super-relaxed (SRX) state of cardiac myosin is thought to be crucial for regulating contraction, relaxation, and energy conservation in the heart. We used single ATP turnover experiments to demonstrate that a dilated cardiomyopathy (DCM) mutation (E525K) in human beta-cardiac myosin increases the fraction of myosin heads in the SRX state (with slow ATP turnover), especially in physiological ionic strength conditions. We also utilized FRET between a C-terminal GFP tag on the myosin tail and Cy3ATP bound to the active site of the motor domain to estimate the fraction of heads in the closed, interacting-heads motif (IHM); we found a strong correlation between the IHM and SRX state. Negative stain electron microscopy and 2D class averaging of the construct demonstrated that the E525K mutation increased the fraction of molecules adopting the IHM. Overall, our results demonstrate that the E525K DCM mutation may reduce muscle force and power by stabilizing the auto-inhibited SRX state. Our studies also provide direct evidence for a correlation between the SRX biochemical state and the IHM structural state in cardiac muscle myosin. Furthermore, the E525 residue may be implicated in crucial electrostatic interactions that modulate this conserved, auto-inhibited conformation of myosin.

• Publisher: eLife Sciences Publications Ltd,eLife Sciences Publications, Ltd
• Subject: Adenosine Triphosphate / Biosensors / Cardiac muscle / cardiac muscle myosin / Cardiac Myosins / Cardiomyopathy / Cardiomyopathy, Dilated - genetics / Conformation / Dilated cardiomyopathy / Ejection fraction / Electron microscopy / Electrostatic properties / Energy conservation / Fluorescence resonance energy transfer / FRET / Genotype & phenotype / Humans / Hypotheses / interacting-heads motif / Ionic strength / Mutation / Myocardium / Myosin / Myosins - genetics / Regulation / Smooth muscle / Structural Biology and Molecular Biophysics / super-relaxed state / Ventricular Myosins - genetics