GENETIC AND IMMUNOLOGIC ANALYSIS OF MICROTUBULE PROTEINS

真核细胞使用基于微管的运输来完成 细胞内的细胞器运动，细胞分裂，以及可能的细胞 形态发生 这些活动都需要微管马达的活动 蛋白质如驱动蛋白。 在这个建议中，我们描述了实验 旨在了解驱动蛋白如何将化学能转化为 机械力，驱动蛋白如何附着在它移动的元素上， 驱动蛋白的哪些结构特征对于体内功能是必需的， 在驱动蛋白适合所有基于MT的运动的情况下， 这可能是由类似驱动蛋白的马达产生的。 完成这些 目标，我们将：1）继续我们的研究驱动蛋白的元素 力产生所需重链马达结构域和开发方法 用于大规模生产驱动蛋白运动域， 高分辨率结构研究; 2）开始蛋白质分析， 驱动蛋白重链尾部与其相互作用以附着 细胞货物3)以确定的方式改变驱动蛋白重链， 将这些改变的分子引入生物体中进行研究， 体内; 4）分析我们最近发现的五个新基因， 似乎编码驱动蛋白样蛋白。 总的来说，我们的调查 揭示了一个特定的微管运动功能，并将开始 揭示真核生物的功能和策略， 完成基于微管的运动

Eukaryotic cells use microtubule-based transport to accomplish intracellular organelle movement, cell division, and possibly cellular morphogenesis. These events require the activities of microtubule-motor proteins such as kinesin. In this proposal we describe experiments designed to understand how kinesin converts chemical energy into mechanical force, how kinesin attaches to the elements that it moves, what structural features of kinesin are essential for in vivo function, and where kinesin fits in the scheme of all MT-based movements, some of which might be generated by kinesin-like motors. To accomplish these goals, we will: 1) continue our studies of the elements of the kinesin heavy chain motor domain needed for force generation and develop methods for large-scale production of the kinesin motor domain for use in high-resolution structural studies; 2) begin analyses of proteins with which the kinesin heavy chain tail interacts in order to attach to cellular cargoes. 3) alter kinesin heavy chain in defined ways and introduce these altered molecules back into the organism for study in vivo; and 4) analyze five new genes that we have recently discovered that appear to encode kinesin-like proteins. In toto, our investigations will reveal how one particular microtubule motor functions and will begin to unravel the range of functions and strategies used by eukaryotes to accomplish microtubule-based movements.