Swartz D R, Moss R L
Department of Physiology, University of Wisconsin, Madison 53706.
J Biol Chem. 1992 Oct 5;267(28):20497-506.
The ability of strong-binding myosin heads to activate the thin filament was investigated by incubating skinned single muscle fibers with N-ethylmaleimide-(NEM) modified myosin subfragment-1 (S1). Isometric force was influenced in a complex manner: during maximal calcium activation, NEM-S1 inhibited force with half-maximal inhibition at 20 microM while at submaximal calcium, NEM-S1 potentiated force with greatest effect at 6 microM. When fibers were treated with NEM-S1 (4-8 microM), the tension-pCa (-log [Ca2+]) relationship became less steep (i.e. the Hill coefficient decreased from 5.4 to 3.0 upon treatment with NEM-S1), but the midpoint was unchanged. These results support the idea that strong binding of intrinsic heads contributes to the cooperativity observed in Ca2+ activation of force. The NEM-S1-induced increase in force at low Ca2+ was associated with an acceleration of a kinetic transition, and this transition was activated to near maximum while force was not. The rate of force redevelopment following restretch (ktr) at submaximal calcium was increased by NEM-S1 in a concentration-dependent manner, yielding a maximum rate at low [Ca2+] which was similar to that observed during full activation. The effects of NEM-S1 on force and ktr indicate that strong-binding myosin cross-bridges are involved in activation of the thin filament.
通过用N - 乙基马来酰亚胺（NEM）修饰的肌球蛋白亚片段1（S1）孵育去表皮单根肌纤维，研究了强结合肌球蛋白头部激活细肌丝的能力。等长力受到复杂的影响：在最大钙激活期间，NEM - S1抑制力，在20 microM时抑制作用达到半数最大抑制，而在亚最大钙浓度下，NEM - S1增强力，在6 microM时效果最佳。当纤维用NEM - S1（4 - 8 microM）处理时，张力 - pCa（-log[Ca2 +]）关系变得不那么陡峭（即，用NEM - S1处理后希尔系数从5.4降至3.0），但中点不变。这些结果支持这样的观点，即内在头部的强结合有助于在Ca2 +激活力时观察到的协同性。在低Ca2 +时NEM - S1诱导的力增加与动力学转变的加速有关，并且这种转变在力未达到最大值时就被激活到接近最大值。在亚最大钙浓度下，NEM - S1以浓度依赖的方式增加了再拉伸后力的恢复速率（ktr），在低[Ca2 +]时产生的最大速率与完全激活期间观察到的相似。NEM - S1对力和ktr的影响表明，强结合肌球蛋白横桥参与细肌丝的激活。
