Department of Molecular & Cellular Biochemistry, Indiana University, Bloomington, Indiana, USA.
Medical Sciences, Indiana University School of Medicine-Bloomington, Bloomington, Indiana, USA.
Cytoskeleton (Hoboken). 2021 Jan;78(1):3-13. doi: 10.1002/cm.21650. Epub 2021 Jan 11.
Myosin active site elements (i.e., switch-1) bind both ATP and a divalent metal to coordinate ATP hydrolysis. ATP hydrolysis at the active site is linked via allosteric communication to the actin polymer binding site and lever arm movement, thus coupling the free energy of ATP hydrolysis to force generation. How active site motifs are functionally linked to actin binding and the power stroke is still poorly understood. We hypothesize that destabilizing switch-1 movement at the active site will negatively affect the tight coupling of the ATPase catalytic cycle to force production. Using a metal-switch system, we tested the effect of interfering with switch-1 coordination of the divalent metal cofactor on force generation. We found that while ATPase activity increased, motility was inhibited. Our results demonstrate that a single atom change that affects the switch-1 interaction with the divalent metal directly affects actin binding and productive force generation. Even slight modification of the switch-1 divalent metal coordination can decouple ATP hydrolysis from motility. Switch-1 movement is therefore critical for both structural communication with the actin binding site, as well as coupling the energy of ATP hydrolysis to force generation.
肌球蛋白活性位点元件（即开关 1）结合 ATP 和二价金属以协调 ATP 水解。通过变构通讯，活性位点的 ATP 水解与肌动蛋白聚合物结合位点和杠杆臂运动相连接，从而将 ATP 水解的自由能与力的产生相偶联。活性位点基序如何与肌动蛋白结合和力产生的功紧密连接仍知之甚少。我们假设破坏活性位点的开关 1运动将对 ATP 酶催化循环与力产生的紧密偶联产生负面影响。我们使用金属开关系统测试了干扰二价金属辅因子与开关 1协调对力产生的影响。我们发现，尽管 ATP 酶活性增加，但运动性受到抑制。我们的结果表明，影响开关 1与二价金属相互作用的单个原子变化直接影响肌动蛋白结合和产生力。即使对开关 1 二价金属配位的微小修饰也可以使 ATP 水解与运动解偶联。因此，开关 1 的运动对于与肌动蛋白结合位点的结构通讯以及将 ATP 水解的能量与力的产生相偶联都是至关重要的。
