Structural basis of motility by dimeric kinesin and myosin motor proteins

二聚驱动蛋白和肌球蛋白运动蛋白运动的结构基础

• 批准号: 10296683
• 负责人: CHARLES VAUGHN SINDELAR
• 金额: $ 34.39万
• 依托单位: YALE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-05-01 至 2023-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10296683
• 关键词: 3-Dimensional; Actins; Active Sites; Address; Automobile Driving; Binding; Biochemical; Cardiomyopathies; Cell division; Cells; Chemical Structure; Chemicals; Classification; Communication; Complex; Consumption; Couples; Cryoelectron Microscopy; Cytokinesis; Cytoskeletal Filaments; Cytoskeleton; DNA Structure; Data; Defect; Development; Disease; Electron Microscope; Filament; Freezing; Funding; Generations; Goals; Head; Hydrolysis; Image; Individual; Investigation; Kinesin; Life; Malignant Neoplasms; Maps; Mechanics; Methods; Microfilaments; Microtubules; Mitosis; Molecular; Molecular Conformation; Molecular Machines; Molecular Motors; Motor; Movement; Myosin ATPase; Myosin Type V; Nature; Nerve; Neurotransmitters; Nonmuscle Myosin Type IIA; Nucleotides; Pharmacologic Substance; Power stroke; Preparation; Process; Production; Proteins; Research; Resolution; Role; Sampling; Shapes; Spastic Paraplegia; Structure; Synapses; System; Time; Walking; Work; analog; arm; cancer therapy; cell motility; design; dimer; experimental study; foot; image processing; improved; inhibitor; inorganic phosphate; insight; molecular rearrangement; nanometer resolution; three dimensional structure; trait

Project Summary Kinesin and myosin are so-called `motor proteins' that can use two `feet' to walk along microtubule and actin filaments (respectively) that make up the cytoskeleton. These motors support many vital functions with the cell, including pulling DNA structures apart during cell division and resupplying nerve junctions (the synapse) with neurotransmitters (such as seratonin). The precise mechanisms by which these elaborate molecular machines, which are composed of tens of thousands of exquisitely arranged atoms, are able to `walk' are complex and incompletely understood. To see how they work, it is necessary to visualize these complex structures in three dimensions at sufficient levels of detail to resolve individual atoms– and to follow molecular rearrangements that happen while the motors step forward. This goal, however, has long remained out of reach due to the extreme technical challenges involved. We have addressed this problem by developing new methods to analyze images of frozen motor-filament assemblies collected by latest-generation electron microscopes. This approach, known as cryo-electron microscopy, allows us to directly visualize the three dimensional shape of individual molecular motor proteins attached to their partner filaments. During the previous funding period, we solved 3D structures of truncated single `feet' (one motor domain) of kinesin and myosin motors attached to their partner filaments, showing in atomic detail how these structures changed when molecules of ATP fuel were bound and consumed. We also captured a 3D structure of an intact pair of kinesin molecules (dimer) caught in mid- step on a microtubule. This allowed us to visualize, for the first time, a way in which the two `feet' of kinesin can pull on each other in a way to stay coordinated while walking. In our ongoing research we are improving our methods to capture more intermediates in the stepping process of kinesin, in order to gain a complete more understanding of how it walks. We are improving our analysis methods to better resolve precise chemical details within these structures. Finally, we are extending our approach to understand how a pair of myosin molecules can walk along the actin filament. Results of our studies are expected to aid the development of a new generation of pharmaceutical agents for treating cancer and a wide variety of other diseases.

项目摘要 驱动蛋白和肌球蛋白是所谓的“运动蛋白”，它们可以用两只“脚”来沿着行走 微管和肌动蛋白丝（分别），构成细胞骨架。这些电机 它支持细胞的许多重要功能，包括在细胞分裂过程中将DNA结构分开。 分裂和再供应神经接头（突触）与神经递质（如 血清素）。这些精细的分子机器， 由成千上万个精巧排列的原子组成，能够“行走”的原子是复杂的， 不完全理解。为了了解它们是如何工作的，有必要将这些复杂的 在三维结构在足够的细节水平，以解决个别原子-和 跟随马达前进时发生的分子重排。这个目标， 然而，由于所涉及的极端技术挑战，长期以来一直无法实现。 我们已经解决了这个问题，通过开发新的方法来分析图像冻结 由最新一代电子显微镜收集的电机灯丝组件。这种方法， 低温电子显微镜，使我们能够直接观察到 分子马达蛋白附着在它们的伴侣细丝上。 在上一个资助期间，我们解决了截断单脚（一个电机）的3D结构 结构域）的驱动蛋白和肌球蛋白马达连接到他们的伴侣丝，显示在原子 详细说明了当ATP燃料分子被结合和消耗时，这些结构是如何变化的。 我们还捕获了一对完整的驱动蛋白分子（二聚体）的3D结构， 踩在微管上。这让我们第一次看到， 驱动蛋白的“脚”可以在行走时以某种方式相互拉动以保持协调。在我们正在进行的 研究我们正在改进我们的方法，以捕获更多的中间体在步进过程中， 为了更全面地了解它是如何行走的我们正在改善 我们的分析方法，以更好地解决这些结构中的精确化学细节。最后， 我们正在扩展我们的方法，以了解一对肌球蛋白分子是如何沿着 肌动蛋白丝我们的研究结果有望帮助开发一种新的 产生用于治疗癌症和多种其它疾病的药剂。