BIOPHYSICS AND ENZYMOLOGY OF KINESIN MOVEMENT

驱动蛋白运动的生物物理学和酶学

• 批准号: 6180396
• 负责人: JEFF GELLES
• 金额: $ 28.94万
• 依托单位: BRANDEIS UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1991
• 资助国家: 美国
• 起止时间: 1991-04-01 至 2002-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6180396
• 关键词: biomechanics; cell motility; chemical conjugate; conformation; enzyme activity; enzyme structure; intracellular transport; kinesin; microtubules; nanotechnology; protein structure function; site directed mutagenesis

Kinesin is a mechanoenzyme that drives microtubule-based intracellular organelle transport processes. Kinesin couples a free-energy-liberating chemical reaction (the hydrolysis of ATP) to a cycle of mechanical processes that move the enzyme molecules and attached organelles along microtubules. We want to characterize the cycle of mechanical processes by which kinesin moves and to determine how these processes are coupled to the reactions of ATP hydrolysis. We have developed novel experimental systems that allow us to directly monitor nanometer-scale mechanical processes, domain movements, and chemical steps in single isolated kinesin molecules using light microscope-based instruments. Intracellular organelle transport by kinesin and kinesin homologs plays an essential role in the physiology of eukaryotic cells. Its functions include transport of materials, chromosome and nuclear movements in mitosis/meiosis, and morphogenesis of membranous organelles. To explore these functions at the molecular level we will: Characterize the conformational changes in the kinsin alpha-helical coiled-coil neck domain that are required for processive motility along microtubules. Test the hand-over-hand hypothesis for the kinesin movement mechanism Detect and characterize the power stroke of an isolated kinesin head domain. The experiments will combine single-molecule biophysics techniques with conventional enzymology methods including protein chemical modification, site-directed mutagenesis, and chemical kinetics techniques.

驱动蛋白是一种机械酶，可驱动基于微管的细胞内 细胞器传输过程。 动力学夫妻伴侣 化学反应（ATP的水解）至机械循环 移动酶分子和附着的细胞器的过程 微管。 我们想表征机械过程的周期 运动蛋白移动并确定如何耦合这些过程 对ATP水解的反应。 我们已经开发了小说 实验系统使我们能够直接监视纳米尺度 机械过程，域运动和化学步骤 使用基于光显微镜的仪器进行分离的驱动蛋白分子。 驱动蛋白和动力蛋白同源物的细胞内细胞器转运 在真核细胞生理学中的重要作用。 它的功能 包括材料的运输，染色体和核运动 有丝分裂/减数症以及膜细胞器的形态发生。 探索 这些功能在分子水平上：我们将： 表征Kinsin alpha螺旋中的构象变化 沿途运动式运动所需 微管。 测试动力蛋白运动机制的手动假设 检测并表征孤立的驱动蛋白头的功率中风 领域。 实验将将单分子生物物理技术与 常规酶学方法，包括蛋白质化学修饰， 定点诱变和化学动力学技术。