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bioRxiv · 10.1101/2023.12.07.570656

Tail Length and E525K Dilated Cardiomyopathy Mutant Alter Human β-Cardiac Myosin Super-Relaxed State

Duno-Miranda, S.·Nelson, S. R.·Rasicci, D. V.·Bodt, S. L. M.·Cirilo, J. A.·Vang, D.·Sivaramakrishnan, S.·Yengo, C. M.·Warshaw, D. M.
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Abstract

Dilated cardiomyopathy (DCM) is characterized by impaired cardiac function due to myocardial hypo-contractility and is associated with point mutations in {beta}-cardiac myosin, the molecular motor that powers cardiac contraction. Myocardial function can be modulated through sequestration of myosin motors into an auto-inhibited "super relaxed" state (SRX), which is further stabilized by a structural state known as the "Interacting Heads Motif" (IHM). Therefore, hypo-contractility of DCM myocardium may result from: 1) reduced function of individual myosin, and/or; 2) decreased myosin availability due to increased IHM/SRX stabilization. To define the molecular impact of an established DCM myosin mutation, E525K, we characterized the biochemical and mechanical activity of wild-type (WT) and E525K human {beta}-cardiac myosin constructs that differed in the length of their coiled-coil tail, which dictates their ability to form the IHM/SRX state. Single-headed (S1) and a short-tailed, double-headed (2HEP) myosin constructs exhibited low ([~]10%) IHM/SRX content, actin-activated ATPase activity of [~]5s-1 and fast velocities in unloaded motility assays ([~]2000nm/s). Double-headed, longer-tailed (15HEP, 25HEP) constructs exhibited higher IHM/SRX content ([~]90%), and reduced actomyosin ATPase (<1s-1) and velocity ([~]800nm/s). A simple analytical model suggests that reduced velocities may be attributed to IHM/SRX-dependent sequestration of myosin heads. Interestingly, the E525K 15HEP and 25HEP mutants showed no apparent impact on velocity or actomyosin ATPase at low ionic strength. However, at higher ionic strength, the E525K mutation stabilized the IHM/SRX state. Therefore, the E525K-associated DCM human cardiac hypo-contractility may be attributable to reduced myosin head availability caused by enhanced IHM/SRX stability. SummaryThis research investigates the E525K mutation in human {beta}-cardiac myosin, crucial for heart contraction, and its role in causing Dilated Cardiomyopathy (DCM). It demonstrates that the length of the myosin tail influences its self-inhibition, and the E525K mutation strengthens this effect, potentially reducing heart contractility in DCM.

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Duno-Miranda, S., Nelson, S. R., Rasicci, D. V., Bodt, S. L. M., Cirilo, J. A., Vang, D., Sivaramakrishnan, S., Yengo, C. M., Warshaw, D. M.. 2023-12-08. Tail Length and E525K Dilated Cardiomyopathy Mutant Alter Human β-Cardiac Myosin Super-Relaxed State. https://doi.org/10.1101/2023.12.07.570656

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