Guilford W H, Dupuis D E, Kennedy G, Wu J, Patlak J B, Warshaw D M
Department of Molecular Physiology and Biophysics, University of Vermont, Burlington 05405, USA.
Biophys J. 1997 Mar;72(3):1006-21. doi: 10.1016/S0006-3495(97)78753-8.
Purified smooth muscle myosin in the in vitro motility assay propels actin filaments at 1/10 the velocity, yet produces 3-4 times more force than skeletal muscle myosin. At the level of a single myosin molecule, these differences in force and actin filament velocity may be reflected in the size and duration of single motion and force-generating events, or in the kinetics of the cross-bridge cycle. Specifically, an increase in either unitary force or duty cycle may explain the enhanced force-generating capacity of smooth muscle myosin. Similarly, an increase in attached time or decrease in unitary displacement may explain the reduced actin filament velocity of smooth muscle myosin. To discriminate between these possibilities, we used a laser trap to measure unitary forces and displacements from single smooth and skeletal muscle myosin molecules. We analyzed our data using mean-variance analysis, which does not rely on scoring individual events by eye, and emphasizes periods in the data with constant properties. Both myosins demonstrated multiple but similar event populations with discrete peaks at approximately +11 and -11 nm in displacement, and 1.5 and 3.5 pN in force. Mean attached times for smooth muscle myosin were longer than for skeletal-muscle myosin. These results explain much of the difference in actin filament velocity between these myosins, and suggest that an increased duty cycle is responsible for the enhanced force-generating capacity of smooth over skeletal-muscle myosin.
在体外运动分析中，纯化的平滑肌肌球蛋白推动肌动蛋白丝的速度仅为骨骼肌肌球蛋白的1/10，但产生的力却是骨骼肌肌球蛋白的3 - 4倍。在单个肌球蛋白分子水平上，这些力和肌动蛋白丝速度的差异可能反映在单个运动和力产生事件的大小和持续时间上，或者反映在横桥循环的动力学上。具体而言，单位力或工作循环的增加可能解释了平滑肌肌球蛋白增强的力产生能力。同样，附着时间的增加或单位位移的减少可能解释了平滑肌肌球蛋白降低的肌动蛋白丝速度。为了区分这些可能性，我们使用激光阱来测量单个平滑肌和骨骼肌肌球蛋白分子的单位力和位移。我们使用均值 - 方差分析来分析数据，该分析不依赖于肉眼对单个事件进行评分，并强调数据中具有恒定特性的时间段。两种肌球蛋白都表现出多个但相似的事件群体，位移在约 +11和 -11 nm处有离散峰值，力在1.5和3.5 pN处有离散峰值。平滑肌肌球蛋白的平均附着时间比骨骼肌肌球蛋白长。这些结果解释了这些肌球蛋白之间肌动蛋白丝速度差异的很大一部分，并表明工作循环的增加是平滑肌肌球蛋白比骨骼肌肌球蛋白力产生能力增强的原因。
