Konishi M, Wakabayashi K, Kurihara S, Higuchi H, Onodera N, Umazume Y, Tanaka H, Hamanaka T, Amemiya Y
Department of Physiology, Jikei University School of Medicine, Tokyo, Japan.
Biophys Chem. 1991 Mar;39(3):287-97. doi: 10.1016/0301-4622(91)80007-e.
Time-resolved X-ray equatorial diffraction studies on a single frog skeletal muscle fiber were performed with a 10 ms time resolution using synchrotron radiation in order to compare the time courses of the molecular changes of contractile proteins and the intracellular Ca2+ transient during an isometric twitch contraction at 2.7 degrees C. Measurements of the Ca2+ transient using aequorin as an intracellular Ca2+ indicator were conducted separately just before and after the X-ray experiments under very similar experimental conditions. The results, which showed a similar time course of tension to that observed in the X-ray experiment, were compared with the aequorin light signal, tension and the intensity changes of the 1,0 and 1,1 equatorial reflections. No appreciable change in both reflection spacings indicated that the effect of internal shortening of the muscle was minimized during contraction. The intensity change of the equatorial reflections generally occurred after the aequorin light signal. In the rising phase, the time course of increase in the 1,1 intensity paralleled that of the rise of the light signal and the intensity peak occurred 20-30 ms after the peak of the light signal. The decrease in the 1,0 intensity showed a time course similar to that of tension and the intensity minimum roughly coincided with the tension peak, coming at 80-90 ms and about 60 ms after the peaks of the light signal and the 1,1 intensity change, respectively. In the relaxation phase, the 1,1 intensity seemed to fall rapidly just before the tension peak and then returned to the original level in parallel with the decay of tension. The 1,0 intensity returned more slowly than the tension relaxation. Thus, the change of the 1,1 intensity was faster than that of the 1,0 intensity in both the rising and relaxation phases. When the measured aequorin light signal was corrected for the kinetic delay of the aequorin reaction with a first-order rate constant of either 50 or 17 s-1, the peak of the corrected light signal preceded that of the measured one by approx. 30 ms. Thus, the peak of the Ca2+ transient appeared earlier than the peaks of the 1,1 and 1,0 intensity changes by 50-60 and 110-120 ms, respectively. The time lag between the extent of structural change and the Ca2+ transient is discussed in relation to the double-headed attachment of a cross-bridge to actin.
为了比较2.7摄氏度下等长单收缩过程中收缩蛋白分子变化和细胞内钙离子瞬变的时间进程，利用同步辐射对单个青蛙骨骼肌纤维进行了时间分辨X射线赤道衍射研究，时间分辨率为10毫秒。在与X射线实验非常相似的实验条件下，分别在X射线实验之前和之后，使用水母发光蛋白作为细胞内钙离子指示剂进行钙离子瞬变的测量。将与X射线实验中观察到的张力时间进程相似的结果，与水母发光蛋白光信号、张力以及1,0和1,1赤道反射的强度变化进行比较。两个反射间距没有明显变化，表明收缩过程中肌肉内部缩短的影响最小化。赤道反射的强度变化通常在水母发光蛋白光信号之后发生。在上升阶段，1,1强度增加的时间进程与光信号上升的时间进程平行，强度峰值在光信号峰值后20 - 30毫秒出现。1,0强度的下降显示出与张力相似的时间进程，强度最小值大致与张力峰值同时出现，分别在光信号峰值和1,1强度变化峰值后80 - 90毫秒和60毫秒左右出现。在松弛阶段，1,1强度似乎在张力峰值之前迅速下降，然后随着张力的衰减平行恢复到原始水平。1,0强度的恢复比张力松弛更慢。因此，在上升和松弛阶段，1,1强度的变化都比1,0强度的变化更快。当用50或17 s-1的一级速率常数校正测量的水母发光蛋白光信号的动力学延迟时，校正后的光信号峰值比测量的光信号峰值提前约30毫秒。因此，钙离子瞬变的峰值分别比1,1和1,0强度变化的峰值早50 - 60毫秒和110 - 120毫秒出现。结合横桥与肌动蛋白的双头附着，讨论了结构变化程度与钙离子瞬变之间的时间滞后。
