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ATP类似物AMPPNP对脊椎动物骨骼肌中横桥结构的影响:X射线衍射和力学研究。

The effect of the ATP analogue AMPPNP on the structure of crossbridges in vertebrate skeletal muscles: X-ray diffraction and mechanical studies.

作者信息

Padrón R, Huxley H E

出版信息

J Muscle Res Cell Motil. 1984 Dec;5(6):613-55. doi: 10.1007/BF00713923.

Abstract

Adenylylimidodiphosphate (AMPPNP), a nonhydrolysable analogue of ATP, has been used to arrest the crossbridge cycle of muscular contraction in one of its hypothetical intermediate states. Whole frog sartorius muscles were chemically demembranated, and it was found possible to cycle such skinned muscles reversibly between the relaxed and rigor states. The effect of binding of AMPPNP on the structure and spatial arrangement of the crossbridges of such muscles was studied using low-angle X-ray diffraction, with simultaneous recording of the mechanical effects, starting from the rigor state. Saturating concentrations of MgAMPPNP produce a characteristic decrease of about 50% in the original rigor isometric tension with a concomitant increase in muscle length by 0.13%. The equatorial X-ray diffraction pattern is modified in the following way: the lattice dimensions and the intensity of the (10) equatorial reflection do not change, while the intensity of the (11) equatorial reflection increases slightly. These observations of very small equatorial changes could be explained by assuming that in these muscles (as distinct from others such as rabbit psoas) the analogue does not produce a significant degree of detachment of crossbridges; that is, there are only AMPPNP-modified attached ones. The changes in the meridional X-ray diffraction pattern are more pronounced: the meridional reflection at 14.5 nm decreases in intensity, and the meridional reflection at 7.2 nm increases considerably: the intensity of all the actin-based off-meridional layer-lines decreases. There are no signs of the characteristic relaxed layer-lines, and the changes in the layer-line intensities are probably due to there being a single population of AMPPNP-modified attached crossbridges, rather than a mixture of attached and detached crossbridges. Thus the AMPPNP X-ray pattern, both equatorially and meridionally, is somewhat similar to the rigor one, indicating that most of the crossbridges remain attached. On the other hand, the fact that there are some changes in the layer-line intensities of the AMPPNP frog pattern, without the appearance of any signs of a relaxed equatorial pattern, indicates that the attached crossbridges are in a structural state that is different from rigor, one is not seeing, apparently, simply a mixture of rigor and relaxed states. Our tentative interpretation of this result is that there may be a structural change in the crossbridge near to the junction with S2, with less significant changes occurring in the parts of the crossbridge close to actin.

摘要

腺苷酰亚胺二磷酸(AMPPNP)是一种不可水解的ATP类似物,已被用于在肌肉收缩的一个假设中间状态下阻止横桥循环。对完整的青蛙缝匠肌进行化学去膜处理,发现可以使这种去皮肌肉在松弛状态和强直状态之间可逆地循环。使用低角度X射线衍射研究了AMPPNP结合对这种肌肉横桥结构和空间排列的影响,并同时记录从强直状态开始的力学效应。饱和浓度的MgAMPPNP会使原始强直等长张力特征性地降低约50%,同时肌肉长度增加0.13%。赤道X射线衍射图谱的变化如下:晶格尺寸和(10)赤道反射强度不变,而(11)赤道反射强度略有增加。这些非常小的赤道变化的观察结果可以通过假设在这些肌肉中(与其他肌肉如兔腰大肌不同)该类似物不会使横桥产生显著程度的脱离来解释;也就是说,只有被AMPPNP修饰的附着横桥。子午X射线衍射图谱的变化更为明显:14.5 nm处的子午反射强度降低,7.2 nm处的子午反射强度大幅增加:所有基于肌动蛋白的离子午线层线的强度都降低。没有特征性松弛层线的迹象,层线强度的变化可能是由于存在单一群体的被AMPPNP修饰的附着横桥,而不是附着和脱离横桥的混合物。因此,AMPPNP的X射线图谱在赤道和子午方向上都与强直图谱有些相似,表明大多数横桥仍然附着。另一方面,AMPPNP青蛙图谱的层线强度有一些变化,而没有出现任何松弛赤道图谱的迹象,这一事实表明附着的横桥处于与强直不同的结构状态,显然不是简单地看到强直和松弛状态的混合物。我们对这一结果的初步解释是,在靠近与S2连接处的横桥可能存在结构变化,而在靠近肌动蛋白的横桥部分发生的变化不太显著。

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