Structure and Conformational Changes of Assembled Dynamin

组装动力的结构和构象变化

• 批准号: 7329726
• 负责人: Joshua S Chappie
• 金额: $ 2.8万
• 依托单位: SCRIPPS RESEARCH INSTITUTE, THE
• 依托单位国家: 美国
• 财政年份: 2007
• 资助国家: 美国
• 起止时间: 2007-07-18 至 2009-07-17
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7329726
• 关键词: Action Potentials; Adopted; Biochemistry; Biological; Cells; Chemicals; Clathrin; Clathrin-Coated Vesicles; Collaborations; Cryoelectron Microscopy; Cysteine; Data; Dynamin; Dynamin I; Electrons; Endocytosis; Event; GTP Binding; Gold; Guanosine Triphosphate; Guanosine Triphosphate Phosphohydrolases; Housing; Hydrolysis; Individual; Knowledge; Label; Letters; Link; Lipids; Localized; Location; Maps; Mediating; Membrane; Mental Health; Microscopy; Molecular; Mutagenesis; Nature; Neurologic; Neurons; Neurotransmitters; Nucleotides; Polymers; Positioning Attribute; Process; Proteins; Purpose; Recycling; Research Institute; Resolution; Resources; Role; Series; Structure; Surface; Synapses; Synaptic Vesicles; Techniques; Translating; Vesicle; Work; image reconstruction; insight; interest; nucleotide analog; reconstruction; response; self assembly; undecagold

DESCRIPTION (provided by applicant): Dynamin is a large, multidomain GTPase essential for the final steps of clathrin mediated endocytosis and is a key regulator of synaptic vesicle recycling. In order to understand the mechanisms by which it modulates these biological events, it is crucial to determine how dynamin translates nucleotide hydrolysis into structural changes to facilitate membrane fission and the liberation of clathrin coated vesicles into the cell. Despite large amounts of functional data, the exact nature of the conformational changes that drive dynamin's mechanochemical activities is still ill-defined due to a lack of structural information. To better understand how dynamin structure influences its functional activities, we seek to visualize the structure and conformational changes of assembled dynamin using a combination of cryo-electron microscopy, biochemistry, and image reconstruction techniques. These studies will yield three-dimensional reconstructions of dynamin at sufficient resolution to allow us to see the key structural changes that are linked to different nucleotide states. Attachment of electron-dense labels will elucidate the overall organization of the dynamin polymer and will unambiguously define the positions of dynamin's domains within these structural maps. This knowledge will greatly increase our understanding of how dynamin transmits the chemical energy of GTP hydrolysis into the physical work of membrane scission and ultimately pre-synaptic recycling. The ability of the body to transmit neurological impulses is highly dependent on the efficiency of a process known as pre-synaptic recycling, which maintains a large pool of vesicles pre-loaded with neurotransmitters and primes the neuron for rapid response to an action potential. The GTPase dynamin is essential for this process, though the underlying mechanisms of its function remain unknown due to a lack of structural information about the protein. We seek to determine how dynamin structure influences its function, thereby establishing the key changes in the protein that drive its biological activities. These findings will ultimately enhance our understanding of dynamin's role in mental health.

描述（由申请人提供）：Dynamin是一个大型多域GTPase，对于网格蛋白介导的内吞作用的最后一步必不可少，并且是突触囊泡回收的关键调节剂。为了理解调节这些生物事件的机制，确定动力蛋白如何将核苷酸水解转化为结构变化至关重要，以促进膜裂变并将细胞蛋白涂层的囊泡释放到细胞中。尽管有大量的功能数据，但由于缺乏结构信息，构象变化的确切性质仍未定义。为了更好地理解元素结构如何影响其功能活动，我们试图使用冷冻电子显微镜，生物化学和图像重建技术的组合来可视化组装动力蛋白的结构和构象变化。这些研究将在足够的分辨率下产生动力蛋白的三维重建，以使我们能够看到与不同核苷酸状态相关的关键结构变化。电子致密标签的附着将阐明Dynamin聚合物的整体组织，并明确定义Dynamin域在这些结构图中的位置。这些知识将大大提高我们对Dynamin如何将GTP水解的化学能传递到膜分裂的物理工作以及最终突触前再循环的理解。人体传播神经脉冲的能力高度取决于称为突触前回收的过程的效率，该过程维持了预载有神经递质的大量囊泡和神经元的神经元，以快速响应动作电位。 GTPase Dynamin对于此过程至关重要，尽管由于缺乏有关蛋白质的结构信息，其功能的基本机制仍然未知。我们试图确定动力蛋白结构如何影响其功能，从而确立驱动其生物活性的蛋白质的关键变化。这些发现最终将增强我们对Dynamin在心理健康中的作用的理解。