MOLECULAR DYNAMICS OF MUSCLE CONTRACTION

这项研究的长期目标是了解 肌肉受力过程中的蛋白质旋转运动 收缩。位点选择性分子探针(自旋标记或 发光染料)将附着在肌球蛋白或肌动蛋白上，以及 光谱实验(电子顺磁共振(EPR)和 时间分辨光学各向异性)将提供直接信息 关于He标记物的取向和转动 蛋白质。这些实验将在纯化的 蛋白质，其中的生化条件可以精确地 控制和监测，以及皮肤肌肉纤维，在 哪些机械性能是可以控制和监测的。 将选择实验条件，以提供直接测试 肌球蛋白头部(交叉桥)和肌动蛋白的拟议运动 在力量产生过程中的子单位。高定向度 EPR的分辨率和光学各向异性的时间分辨率将 用于解析复杂结构下存在的多种构象 生理条件。尽管主要重点将是 在这些应用程序中，该项目的一个组成部分是 光谱方法的发展。五个密切相关的 该项目的主要目标是(1)开发EPR技术 (包括常规转移和饱和转移)(2)应用 这些EPR技术用来检测物体的方向和旋转 肌球蛋白和肌动蛋白，(3)时间分辨光学的发展 各向异性技术(瞬时吸收、磷光和 荧光)，(4)这些光学技术在 解决肌球蛋白和肌动蛋白的旋转运动，以及(5) 分子动力学与结构、力学、 并对相同的制剂进行生化测量。这些 研究应该为分子的作用提供直接的见解 在这个基本的生理过程中的动力学，以及 我们正在开发的技术应该适用范围很广 构象动力学的其他生物物理系统 再加上酶的作用。

The long-term goal of this research is to understand the role of protein rotational motions in the generation of force during muscle contraction. Site-selective molecular probes (spin labels or luminescent dyes) will be attached to myosin or actin, and spectroscopy experiments (electron paramagnetic resonance (EPR) and time-resolved optical anisotropy) will provide direct information about the orientation and rotational motions of he labeled proteins. These experiments will be performed on purified proteins, in which biochemical conditions can be precisely controlled and monitored, and also on skinned muscle fibers, in which mechanical performance can be controlled and monitored. Experimental conditions will be chosen to provide direct tests for the proposed motions of myosin heads (cross-bridges) and actin subunits during the generation of force. The high orientational resolution of EPR and time resolution of optical anisotropy will be used to resolve the multiple conformations present under complex physiological conditions. Although the primary emphasis will be on these applications, an integral apart of this project is the development of the spectroscopic methods. The five closely related principal aims of the project are (1) development of EPR techniques (both conventional and saturation-transfer) (2) application of these EPR techniques to detect the orientations and rotations of myosin and actin, (3) development of time-resolved optical anisotropy techniques (transient absorption, phosphorescence, and fluorescence), (4) application of these optical techniques to resolve the rotational motions of myosin and actin, and (5) correlation of the molecular dynamics with structural, mechanical, and biochemical measurements on the same preparations. These studies should provide direct insight into the role of molecular dynamics in this fundamental physiological process, and the technology we are developing should be applicable to a wide range of other biophysical system in which conformational dynamics are coupled to enzyme action.