Direct Observation of Dynein Motility Using Biophysics

利用生物物理学直接观察动力蛋白运动

• 批准号: 6994090
• 负责人: JENNIFER L ROSS
• 金额: $ 4.4万
• 依托单位: UNIVERSITY OF PENNSYLVANIA
• 依托单位国家: 美国
• 财政年份: 2005
• 资助国家: 美国
• 起止时间: 2005-09-01 至 2008-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6994090
• 关键词: biophysics; dynein ATPase; fluorescence microscopy; green fluorescent proteins; intracellular transport; kinesin; microtubule associated protein; molecular biology; nanotechnology; optical polarization; postdoctoral investigator; protein binding; protein purification; protein transport; single cell analysis

DESCRIPTION (provided by applicant): Cytoplasmic dynein is the main minus-end directed motor protein to walk along microtubules inside cells, but the complex structure has made dynein difficult to purify and study in vitro. This study aims to purify dynein and label the motors with fluorophores or beads to probe their stalling force, step size, and processivity. Dynein coated beads will be grabbed by optical tweezers to detect steps and forces. Single fluorescent dyneins can be accurately located as they walk along microtubules using total internal reflection fluorescence microscopy and fluorescence imaging with one nanometer accuracy to determine the step size and processivity. This work will not only reveal the inner workings of the dynein motor protein, but will also further inform us of dynein malfunction related to motor neuron diseases.

描述（由申请人提供）：细胞质动力蛋白是沿细胞内部微管行走的主要负端导向蛋白，但是复杂的结构使动力蛋白难以在体外纯化和研究。 这项研究旨在净化动力蛋白并用荧光团或珠子标记电动机，以探测其停滞力，步长和加工性。 Dynein涂层的珠子将被光学镊子抓住，以检测步骤和力。单个荧光动力蛋白可以在沿微管行走时使用总内反射荧光显微镜和荧光成像，具有一个纳米的精度，以确定步骤尺寸和加工性。 这项工作不仅会揭示动力蛋白运动蛋白的内部起作用，而且还将进一步告知我们与运动神经元疾病有关的动力蛋白故障。