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Exploring the super-relaxed state of myosin in myofibrils from fast-twitch, slow-twitch, and cardiac muscle

Walklate, Jonathan, Kao, Kerry, Regnier, Michael, Geeves, Michael A. (2022) Exploring the super-relaxed state of myosin in myofibrils from fast-twitch, slow-twitch, and cardiac muscle. Journal of Biological Chemistry, 298 (3). Article Number 101640. ISSN 0021-9258. (doi:10.1016/j.jbc.2022.101640) (KAR id:93569)

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Muscle myosin heads, in the absence of actin, have been shown to exist in two states, the relaxed (turnover ~0.05 s-1) and super-relaxed states (SRX, 0.005 s-1) using a simple fluorescent ATP chase assay (Hooijman, P. et al (2011) Biophys. J. 100, 1969–1976). Studies have normally used purified proteins, myosin filaments or muscle fibers. Here we use muscle myofibrils, which retain most of the ancillary proteins and 3-D architecture of muscle and can be used with rapid mixing methods. Recording time scales from 0.1 – 1000 sec provides a precise measure of the two populations of myosin heads present in relaxed myofibrils. We demonstrate that the population of SRX states is formed from rigor cross bridges within 0.2 s of relaxing with fluorescently labelled ATP, and the population of SRX states is relatively constant over the temperature range of 5 °C – 30 °C. The SRX population is enhanced in the presence of mavacamten and reduced in the presence of deoxy-ATP. Compared to myofibrils from fast twitch muscle, slow-twitch and cardiac muscles myofibrils require a 10-fold lower concentration of mavacamten to be effective, and mavacamten induced a larger increase in the population of the SRX state. Mavacamten is less effective, however, at stabilizing the SRX state at physiological temperatures than at 5 °C. These assays require small quantities of myofibrils, making them suitable for studies of model organism muscles, human biopsies, or human derived iPSCs.

Item Type: Article
DOI/Identification number: 10.1016/j.jbc.2022.101640
Uncontrolled keywords: Myosin, Actin, Cardiac muscle, Skeletal muscle, induced pluripotent stem cells (iPSC), Mavacamten, myofibrils, myosin filament, regulation, deoxy-ATP
Subjects: Q Science > QP Physiology (Living systems) > QP517 Biochemistry
Divisions: Divisions > Division of Natural Sciences > Biosciences
Funders: [29] EU
Depositing User: Michael Geeves
Date Deposited: 12 Mar 2022 16:25 UTC
Last Modified: 14 Mar 2022 15:00 UTC
Resource URI: (The current URI for this page, for reference purposes)
Walklate, Jonathan:
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