THICK FILAMENT MYOSIN AND C-PROTEIN ARRANGEMENT

粗丝肌球蛋白和 C 蛋白排列

基本信息

批准号：
3155810
负责人：
Robert W. Kensler
金额：
$ 13.54万
依托单位：
UNIVERSITY OF MISSOURI KANSAS CITY
依托单位国家：
美国
项目类别：
财政年份：
1988
资助国家：
美国
起止时间：
1988-10-01 至 1993-12-31
项目状态：
已结题

项目摘要

The arrangement of myosin and accessory protein such as C- protein in the thick filaments of fish, avian, and mammalian skeletal muscles is still not clearly elucidated. Recent work in our laboratory on the structure of frog thick filaments however, has demonstrated that the structure of vertebrate muscles may be amenable to analysis by a combination of electron microscopy, optical diffraction analysis, and computer image analysis. Additionally, it has been shown that thick filaments from fich, avian, and mammalian muscle can be isolated with a similarly well ordered array of crossbridges; thus making these filaments suitable for analysis by the same type of techniques which have proven useful for the frog thick filaments. The specific aim of the proposed research is to extend the techniques used for the frog thick filaments to the determination of the structure of the thick filaments of fish, avian, and mammalian skeletal muscles. The major objective of theses studies will be the determination of the arrangement of the myosin and accessory proteins in the filaments of these vertebrate classes. The studies to accomplish these aims will include: 1) the use of a combination of negative staining, platinum shadowing, rapid-freeze deep-etch cryo studies, and optical diffraction analysis to elucidate the major ultrastructural features of the filaments and to assess the preservation of the filaments as compared to the structural parameters expected from X-ray diffraction patterns of living muscle; 2) the use of computer image analysis of the filament images in the best electron micrographs to determine the rotational symmetry and arrangement of the myosin heads, and to produce a three-dimensional reconstruction of the structure of the filament; 3) the use of immunolabelling studies to determine the probable location of accessory proteins such as C-protein and X-protein on the filaments, and to correlate this information with the structure of the filament as seen in the three dimensional reconstructions. These studies will provide basic scientific information about the structure of muscle, and will thus provide a better foundation for understanding how pathological conditions may affect the functioning of this tissue.

肌球蛋白和辅助蛋白的排列，例如C- 鱼类，禽和哺乳动物的厚细丝中的蛋白质骨骼肌仍未清楚地阐明。最近的工作但是，我们关于青蛙厚细丝结构的实验室已经证明脊椎动物肌肉的结构可能是可以通过电子显微镜组合进行分析，光学衍射分析和计算机图像分析。此外，已经显示出Fich的厚细丝，禽和哺乳动物肌肉可以与类似井井有条的越野阵列；从而使这些细丝适用于具有相同类型的技术的分析被证明对青蛙厚细丝有用。特定目的的拟议的研究是扩展用于青蛙厚细丝以确定结构鱼，禽和哺乳动物骨骼肌的厚细丝。这些研究的主要目的是确定肌球蛋白和附件蛋白的排列这些脊椎动物类的细丝。完成的研究这些目标包括：1）使用负面的组合染色，白金阴影，快速冻结深蚀刻冷冻研究，和光学衍射分析以阐明主要细丝的超微结构特征，并评估与结构相比，细丝的保存 X射线衍射模式的参数肌肉; 2）使用灯丝的计算机图像分析最佳电子显微照片中的图像以确定肌球蛋白头的旋转对称性和排列产生三维结构的重建细丝； 3）使用免疫标签研究来确定附件蛋白（例如C蛋白和）的可能位置细丝上的X蛋白，并将这些信息与灯丝的结构如三维中所见重建。这些研究将提供基本的科学有关肌肉结构的信息，因此将提供理解病理状况的更好基础可能会影响该组织的功能。