Yamaguchi M, Takemori S
Department of Physiology, The Jikei University School of Medicine, Minato-ku, Tokyo 105-8461, Japan.
Biophys J. 2001 Jan;80(1):371-8. doi: 10.1016/S0006-3495(01)76021-3.
To assess the activating efficiency of Ca2+ and cross-bridges, the release rates of phosphate analogs from skinned fibers were estimated from the recovery of contractility and that of stiffness. Estimations were performed based on the assumptions that contractility was indicative of the population of analog-free myosin heads and that stiffness reflected the population of formed cross-bridges. Aluminofluoride (AlFx) and orthovanadate (Vi) were used as phosphate analogs with mechanically skinned fibers from rabbit psoas muscle. The use of the analogs enabled the functional assessment of activation level in the total absence of ATP. Fibers loaded with the analogs gradually recovered contractility and stiffness in normal plain rigor solution. The addition of Ca2+ to the plain rigor solution significantly accelerated their recovery, whereas ADP had no appreciable effect. ATP plus Ca2+(contracting condition) accelerated the recovery by several tens of times. These results indicate that the cross-bridges formed during contraction have prominent activating efficiency, which is indispensable to attain full activation. A comparison between the activating efficiency evaluated from stiffness and that from contractility suggested that Ca2+ is more potent in accelerating the binding of actin to analog-bound myosin heads whereas cross-bridges mainly accelerate the subsequent analog-releasing step.
为了评估Ca2+和横桥的激活效率，通过收缩力和刚度的恢复来估算脱膜纤维中磷酸类似物的释放速率。估算基于以下假设：收缩力代表不含类似物的肌球蛋白头部的数量，而刚度反映已形成横桥的数量。使用铝氟化物（AlFx）和原钒酸盐（Vi）作为磷酸类似物，对来自兔腰大肌的机械脱膜纤维进行研究。使用这些类似物能够在完全没有ATP的情况下对激活水平进行功能评估。加载了类似物的纤维在正常的普通强直溶液中逐渐恢复收缩力和刚度。向普通强直溶液中添加Ca2+显著加速了它们的恢复，而ADP没有明显影响。ATP加Ca2+（收缩条件）使恢复加速了几十倍。这些结果表明，收缩过程中形成的横桥具有显著的激活效率，这是实现完全激活所必需的。从刚度评估的激活效率与从收缩力评估的激活效率之间的比较表明，Ca2+在加速肌动蛋白与结合了类似物的肌球蛋白头部的结合方面更有效，而横桥主要加速随后的类似物释放步骤。
