静息肌肉中的肌球蛋白马达通过在有序螺旋阵列中对肌球蛋白丝的骨架折叠而失活,并且必须从该构象中释放以参与力的产生。两栖动物肌肉单纤维的时间分辨X射线衍射表明,可以通过在刺激开始时施加空载缩短来抑制肌球蛋白丝的激活,表明细丝被机械应力激活。在这里,我们使用小鼠在28°C时收缩的整个趾长伸肌改善了该方法的信噪比。与肌球蛋白丝激活相关的X射线信号变化,包括与螺旋马达阵列相关的一阶肌球蛋白层线的减少,增加基于肌球蛋白的反射的间距,与肌球蛋白尾巴在细丝主链中的堆积有关,并增加与电机远离主干运动相关的1,1和1,0赤道反射的比率,通过在刺激开始时施加10毫秒的空载缩短来延迟。这些结果表明,肌球蛋白丝主要被丝应力激活,就像两栖动物肌肉一样。然而,检测到零负载时灯丝激活的一小部分,暗示部分细丝激活的独立机制。X射线干扰测量表明,在力量发展开始时,肌球蛋白运动构象发生了开关状变化,伴随着肌球蛋白细丝中点附近区域电机的瞬时紊乱,这表明细丝的带状动力学也在其激活中起作用。关键点:通过从刺激开始施加快速缩短,可以延迟小鼠趾长伸肌中肌球蛋白丝的激活。应激是这些肌肉肌球蛋白丝激活的主要机制,但是在零应力下的电刺激过程中,细丝激活的成分很小。肌球蛋白马达在力发育早期迅速从折叠的抑制构象切换到肌动蛋白附着的力产生构象。
Myosin motors in resting muscle are inactivated by folding against the backbone of the myosin filament in an ordered helical array and must be released from that conformation to engage in force generation. Time-resolved X-ray diffraction from single fibres of amphibian muscle showed that myosin filament activation could be inhibited by imposing unloaded shortening at the start of stimulation, suggesting that filaments were activated by mechanical stress. Here we improved the signal-to-noise ratio of that approach using whole extensor digitorum longus muscles of the mouse contracting tetanically at 28°C. Changes in X-ray signals associated with myosin filament activation, including the decrease in the first-order myosin layer line associated with the helical motor array, increase in the spacing of a myosin-based reflection associated with packing of myosin tails in the filament backbone, and increase in the ratio of the 1,1 and 1,0 equatorial reflections associated with movement of motors away from the backbone, were delayed by imposing 10-ms unloaded shortening at the start of stimulation. These results show that myosin filaments are predominantly activated by filament stress, as in amphibian muscle. However, a small component of filament activation at zero load was detected, implying an independent mechanism of partial filament activation. X-ray interference measurements indicated a switch-like change in myosin motor conformation at the start of force development, accompanied by transient disordering of motors in the regions of the myosin filament near its midpoint, suggesting that filament zonal dynamics also play a role in its activation. KEY POINTS: Activation of myosin filaments in extensor digitorum longus muscles of the mouse is delayed by imposing rapid shortening from the start of stimulation. Stress is the major mechanism of myosin filament activation in these muscles, but there is a small component of filament activation during electrical stimulation at zero stress. Myosin motors switch rapidly from the folded inhibited conformation to the actin-attached force-generating conformation early in force development.