MECHANICS OF KINESIN--MICROTUBULE-BASED MOTOR PROTEIN

驱动蛋白的机制——基于微管的运动蛋白

• 批准号: 3161026
• 负责人: Jonathon Howard
• 金额: $ 14.65万
• 依托单位: UNIVERSITY OF WASHINGTON
• 依托单位国家: 美国
• 财政年份: 1990
• 资助国家: 美国
• 起止时间: 1990-06-30 至 1995-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3161026
• 关键词: adenosine triphosphate; biophysics; chemical bond; chemical models; dynein ATPase; ear hair cell; eukaryote; high energy compound; kinesin; membrane activity; membrane channels; microtubules; myosins; phosphates; protein structure function

The long-term objective of the proposed studies is to understand the physical chemistry of chemomechanical transduction, the conversion of chemical energy contained in high-energy phosphate bonds to mechanical energy used to power intracellular movement. This process is accomplished by several enzymes, termed motor proteins, that include myosin from muscle cells, dynein from cilia and flagella, and cytoplasmic dynein and kinesin from eukaryotic cells in general. Current models, such as the crossbridge model, postulate that the underlying transduction process is a cyclic reaction of a motor molecule with a cytoskeletal polymer, an actin filament in the case of myosin and a microtubule in the instances of dynein and kinesin. After binding to the filament, the motor protein is thought to undergo a conformational change, the power stroke, that produces an increment of movement. The protein then releases the filament before rebinding at another site along the filament and initiating another cycle. The specific aim of the proposed experiments is to test directly such models by making mechanical measurements of the transduction reaction at the single-molecular level. The movement of microtubules across surfaces coated at low density with purified kinesin will be visualized by dark- field microscopy. Special apparatus, previously used by the investigator to study force-sensitive ion channels in hair cells of the ear, will be used to exert forces and to measure displacements with subnanometer precision on a millisecond timescale. The distance that a single kinesin molecule moves a microtubule upon the hydrolysis of a molecule of ATP will be determined. After characterizing the movement of microtubules by single kinesin molecules, the nature of the interactions between several kinesin molecules moving one microtubules by single kinesin molecules, the nature of the interactions between several kinesin molecules moving one microtubule will be studied in order to predict the behavior of large assemblies of motor proteins such as those found in muscles and cilia. Because of the structural and biochemical similarities between kinesin and other motor proteins, the elucidation of the molecular events underlying transduction by kinesin should significantly increase the understanding of cellular motility in general. It is hoped that this understanding may lead to more rational treatments of muscle disorders such as heart disease, or to better methods of selectively interfering with pathological cellular movements such as the invasion and proliferation of tumor cells, and the transport of viruses between the cell membrane and the nucleus.

拟议研究的长期目标是了解 化学机械转导的物理化学， 高能磷酸盐中所含的化学能与机械能 用于驱动细胞内运动的能量。 这个过程就完成了 由几种酶（称为运动蛋白）产生，其中包括来自肌肉的肌球蛋白 细胞，来自纤毛和鞭毛的动力蛋白，以及细胞质动力蛋白和驱动蛋白 一般来自真核细胞。 当前型号，例如横桥 模型，假设底层转导过程是一个循环 运动分子与细胞骨架聚合物（肌动蛋白丝）的反应 在肌球蛋白和微管的情况下，在动力蛋白和微管的情况下 驱动蛋白。 与细丝结合后，运动蛋白被认为 经历构象变化，动力冲程，产生 运动增量。 然后蛋白质在释放丝之前 沿着细丝在另一个位点重新结合并启动另一个循环。 所提出的实验的具体目的是直接测试这样的 通过对转导反应进行机械测量来建立模型 单分子水平。 微管在表面上的运动 以低密度涂覆有纯化的驱动蛋白将通过暗-可视化 场显微镜。 调查员之前使用过的特殊设备 研究耳毛细胞中的力敏感离子通道 用于施加力并用亚纳米测量位移 精确到毫秒时间尺度。 单个驱动蛋白的距离 ATP 分子水解后，分子会移动微管 被确定。 在通过单个微管运动表征后 驱动蛋白分子，几种驱动蛋白之间相互作用的性质 分子通过单个驱动蛋白分子移动一个微管，性质 几个驱动蛋白分子之间的相互作用使一个 将研究微管以预测大分子的行为 运动蛋白的组装，例如在肌肉和纤毛中发现的运动蛋白的组装。 由于驱动蛋白和驱动蛋白之间的结构和生化相似性 其他运动蛋白，阐明潜在的分子事件 驱动蛋白的转导应该会显着增加对 一般细胞运动。 希望这种理解能够引导 对心脏病等肌肉疾病进行更合理的治疗，或者 寻找更好的选择性干扰病理细胞的方法 肿瘤细胞的侵袭、增殖等运动 病毒在细胞膜和细胞核之间的运输。