Biology, Illinois Institute of Technology, Chicago, IL 60616.
Bioengineering, University of Washington, Seattle, WA 98109.
Proc Natl Acad Sci U S A. 2023 Jan 31;120(5):e2207615120. doi: 10.1073/pnas.2207615120. Epub 2023 Jan 25.
Contraction in striated muscle is initiated by calcium binding to troponin complexes, but it is now understood that dynamic transition of myosin between resting, ordered OFF states on thick filaments and active, disordered ON states that can bind to thin filaments is critical in regulating muscle contractility. These structural OFF to ON transitions of myosin are widely assumed to correspond to transitions from the biochemically defined, energy-sparing, super-relaxed (SRX) state to the higher ATPase disordered-relaxed (DRX) state. Here we examined the effect of 2'-deoxy-ATP (dATP), a naturally occurring energy substrate for myosin, on the structural OFF to ON transitions of myosin motors in porcine cardiac muscle thick filaments. Small-angle X-ray diffraction revealed that titrating dATP in relaxation solutions progressively moves the myosin heads from ordered OFF states on the thick filament backbone to disordered ON states closer to thin filaments. Importantly, we found that the structural OFF to ON transitions are not equivalent to the biochemically defined SRX to DRX transitions and that the dATP-induced structural OFF to ON transitions of myosin motors in relaxed muscle are strongly correlated with submaximal force augmentation by dATP. These results indicate that structural OFF to ON transitions of myosin in relaxed muscle can predict the level of force attained in calcium-activated cardiac muscle. Computational modeling and stiffness measurements suggest a final step in the OFF to ON transition may involve a subset of DRX myosins that form weakly bound cross-bridges prior to becoming active force-producing cross-bridges.
在横纹肌中，收缩是由钙与肌钙蛋白复合物结合引发的，但现在人们已经明白，肌球蛋白在粗丝上的静止、有序的 OFF 状态和能够与细丝结合的活跃、无序的 ON 状态之间的动态转换，对于调节肌肉收缩力至关重要。这些肌球蛋白的结构 OFF 到 ON 的转变被广泛认为与从生化定义的、节能的、超松弛（SRX）状态到更高的 ATPase 无序松弛（DRX）状态的转变相对应。在这里，我们研究了 2'-脱氧-ATP（dATP）对猪心肌粗丝中肌球蛋白马达结构 OFF 到 ON 转变的影响。小角 X 射线衍射表明，在松弛溶液中滴定 dATP 会使肌球蛋白头部从粗丝骨架上有序的 OFF 状态逐渐转变为更接近细丝的无序 ON 状态。重要的是，我们发现结构 OFF 到 ON 的转变不等同于生化定义的 SRX 到 DRX 转变，并且 dATP 诱导的松弛肌肉中肌球蛋白马达的结构 OFF 到 ON 的转变与 dATP 引起的亚最大力增强密切相关。这些结果表明，松弛肌肉中肌球蛋白的结构 OFF 到 ON 转变可以预测钙激活心肌中达到的力水平。计算建模和刚性测量表明，OFF 到 ON 转变的最后一步可能涉及一组 DRX 肌球蛋白，它们在成为有活性的力产生横桥之前形成弱结合的横桥。
