Cellular Biophysics, Institute for Biophysical Chemistry, Hannover Medical School, 30625 Hannover, Germany.
Structural Bioinformatics and Chemical Biology, Institute for Biophysical Chemistry, Hannover Medical School, 30625 Hannover, Germany.
Int J Mol Sci. 2020 Dec 24;22(1):104. doi: 10.3390/ijms22010104.
The actomyosin system generates mechanical work with the execution of the power stroke, an ATP-driven, two-step rotational swing of the myosin-neck that occurs post ATP hydrolysis during the transition from weakly to strongly actin-bound myosin states concomitant with P release and prior to ADP dissociation. The activating role of actin on product release and force generation is well documented; however, the communication paths associated with weak-to-strong transitions are poorly characterized. With the aid of mutant analyses based on kinetic investigations and simulations, we identified the W-helix as an important hub coupling the structural changes of switch elements during ATP hydrolysis to temporally controlled interactions with actin that are passed to the central transducer and converter. Disturbing the W-helix/transducer pathway increased actin-activated ATP turnover and reduced motor performance as a consequence of prolonged duration of the strongly actin-attached states. Actin-triggered P release was accelerated, while ADP release considerably decelerated, both limiting maximum ATPase, thus transforming myosin-2 into a high-duty-ratio motor. This kinetic signature of the mutant allowed us to define the fractional occupancies of intermediate states during the ATPase cycle providing evidence that myosin populates a cleft-closure state of strong actin interaction during the weak-to-strong transition with bound hydrolysis products before accomplishing the power stroke.
肌球蛋白系统通过执行动力冲程产生机械功，这是一个 ATP 驱动的、两步旋转摆动肌球蛋白颈的过程，发生在 ATP 水解后，从弱结合肌动蛋白状态到强结合肌动蛋白状态的转变过程中，伴随着 P 释放和 ADP 解离之前。肌动蛋白在产物释放和力产生中的激活作用已有充分的文献记载；然而，与弱结合到强结合转变相关的通讯途径还没有很好地描述。通过基于动力学研究和模拟的突变分析，我们确定了 W 螺旋作为一个重要的连接点，它将 ATP 水解过程中开关元件的结构变化与肌动蛋白的时间控制相互作用联系起来，并传递到中央转换器。干扰 W 螺旋/转换器途径会增加肌球蛋白的 ATP 周转率，降低运动性能，因为强肌动蛋白结合状态的持续时间延长。肌动蛋白触发的 P 释放加速，而 ADP 释放显著减速，这两种情况都限制了最大 ATP 酶的活性，从而将肌球蛋白-2转化为高比例的运动蛋白。突变体的这种动力学特征使我们能够在 ATP 酶循环中定义中间状态的分数占有率，这证明了肌球蛋白在弱结合到强结合转变过程中存在一个与结合水解产物的强肌动蛋白相互作用的裂缝关闭状态，然后再完成动力冲程。
