Caremani Marco, Melli Luca, Dolfi Mario, Lombardi Vincenzo, Linari Marco
Laboratory of Physiology, Department of Biology, University of Florence, Sesto Fiorentino, 50019, Italy.
J Physiol. 2015 Aug 1;593(15):3313-32. doi: 10.1113/JP270265. Epub 2015 Jul 7.
Muscle contraction is due to cyclical ATP-driven working strokes in the myosin motors while attached to the actin filament. Each working stroke is accompanied by the release of the hydrolysis products, orthophosphate and ADP. The rate of myosin-actin interactions increases with the increase in shortening velocity. We used fast half-sarcomere mechanics on skinned muscle fibres to determine the relation between shortening velocity and the number and strain of myosin motors and the effect of orthophosphate concentration. A model simulation of the myosin-actin reaction explains the results assuming that orthophosphate and then ADP are released with rates that increase as the motor progresses through the working stroke. The ADP release rate further increases by one order of magnitude with the rise of negative strain in the final motor conformation. These results provide the molecular explanation of the relation between the rate of energy liberation and shortening velocity during muscle contraction. The chemo-mechanical cycle of the myosin II--actin reaction in situ has been investigated in Ca(2+)-activated skinned fibres from rabbit psoas, by determining the number and strain (s) of myosin motors interacting during steady shortening at different velocities (V) and the effect of raising inorganic phosphate (Pi) concentration. It was found that in control conditions (no added Pi ), shortening at V ≤ 350 nm s(-1) per half-sarcomere, corresponding to force (T) greater than half the isometric force (T0 ), decreases the number of myosin motors in proportion to the reduction of T, so that s remains practically constant and similar to the T0 value independent of V. At higher V the number of motors decreases less than in proportion to T, so that s progressively decreases. Raising Pi concentration by 10 mM, which reduces T0 and the number of motors by 40-50%, does not influence the dependence on V of number and strain. A model simulation of the myosin-actin reaction in which the structural transitions responsible for the myosin working stroke and the release of the hydrolysis products are orthogonal explains the results assuming that Pi and then ADP are released with rates that increase as the motor progresses through the working stroke. The rate of ADP release from the conformation at the end of the working stroke is also strain-sensitive, further increasing by one order of magnitude within a few nanometres of negative strain. These results provide the molecular explanation of the relation between the rate of energy liberation and the load during muscle contraction.
肌肉收缩是由于肌球蛋白马达在附着于肌动蛋白丝时，由ATP驱动的周期性工作冲程所致。每个工作冲程都伴随着水解产物正磷酸盐和ADP的释放。肌球蛋白与肌动蛋白相互作用的速率随着缩短速度的增加而增加。我们利用快速半肌节力学对去表皮肌纤维进行研究，以确定缩短速度与肌球蛋白马达数量和应变之间的关系，以及正磷酸盐浓度的影响。肌球蛋白 - 肌动蛋白反应的模型模拟解释了这些结果，假设正磷酸盐然后是ADP以随着马达在工作冲程中推进而增加的速率释放。在最终马达构象中，随着负应变的增加，ADP释放速率进一步增加一个数量级。这些结果为肌肉收缩过程中能量释放速率与缩短速度之间的关系提供了分子解释。通过确定在不同速度（V）下稳定缩短过程中相互作用的肌球蛋白马达数量和应变（s）以及提高无机磷酸盐（Pi）浓度的影响，研究了兔腰大肌Ca(2+)激活的去表皮纤维中肌球蛋白II - 肌动蛋白反应的化学 - 机械循环。发现在对照条件下（不添加Pi），每半肌节以V≤350 nm s(-1)缩短，对应于大于等长力（T0）一半的力（T），肌球蛋白马达数量与T的降低成比例减少，因此s实际上保持恒定且类似于与V无关的T0值。在较高的V下，马达数量的减少与T不成比例，因此s逐渐降低。将Pi浓度提高10 mM，这会使T0和马达数量减少40 - 50%，但不会影响数量和应变对V的依赖性。肌球蛋白 - 肌动蛋白反应的模型模拟中，负责肌球蛋白工作冲程和水解产物释放的结构转变是正交的，该模型解释了这些结果，假设Pi然后是ADP以随着马达在工作冲程中推进而增加的速率释放。工作冲程结束时构象的ADP释放速率也对应变敏感，在负应变的几纳米范围内进一步增加一个数量级。这些结果为肌肉收缩过程中能量释放速率与负载之间的关系提供了分子解释。
