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Microtubule Polymerization and Depolymerization Mechanisms by Conserved Proteins

保守蛋白的微管聚合和解聚机制

基本信息

批准号：
8042154
负责人：
Jawdat MH Al-Bassam
金额：
$ 7.23万
依托单位：
HARVARD MEDICAL SCHOOL
依托单位国家：
美国
项目类别：
财政年份：
2010
资助国家：
美国
起止时间：
2010-04-01 至 2011-03-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8042154
关键词：
Affect Binding Biochemical Biochemistry CDC2 Protein Kinase Caenorhabditis elegans Cell division Complex Crystallography Cyclin-Dependent Kinases Defect Dimerization Electron Microscopy Environment Family Fingers Fluorescence Microscopy Germany Goals Human Genetics In Vitro Institutes Interphase Kinesin Laboratories Learning Link Malignant Neoplasms Mediating Mentors Mentorship Methodology Microscopy Microtubule Depolymerization Microtubule Polymerization Microtubules Mitosis Mitotic Molecular Molecular Conformation Molecular Models Motor Mutation N-terminal Orthologous Gene Pathway interactions Phase Phosphorylation Phosphorylation Site Phosphotransferases Plus End of the Microtubule Polymerase Postdoctoral Fellow Property Protein Family Proteins Regulation Research Research Personnel Role Saccharomyces cerevisiae Saccharomycetales Structure Surface Tubulin Tubulin Interaction Work Xenopus base cancer therapy depolymerization dimer experience medical schools molecular modeling monomer mutant nucleotide analog polymerization programs

项目摘要

DESCRIPTION (provided by applicant): The goal of this research is to understand the molecular mechanism of microtubule (MT) polymerization and depolymerization which are mediated by conserved families of proteins. These proteins act as tubulin polymerases and depolymerases at MT plus ends. Under the mentorship of Dr. Stephen C. Harrison at Harvard Medical School, I have developed a molecular model using electron microscopy and biochemistry for how budding yeast Stu2 dimers and Xenopus XMAP215 monomers bind tubulin dimer via their conseved N-terminal TOG domains. I have determined the structure of a TOG domain using x-ray crystallography and show a narrow conserved surface on the TOG paddle-like structure is the tubulin binding interface. My work suggests a hypothesis for how TOG domains enhance tubulin polymerization at MT plus ends. We are currently studying the structure of TOG domains bound to tubulin dimer. In the mentored phase, I will determine the structure of TOG domains-tubulin dimer complex to understand how TOG domains bind and enhance tubulin assembly. I also intend to explore the role of different TOG domains in enhancing MT assembly using microscopy methodologies in collaberation with the laboratories of Anthony A Hyman and Jonathan Howard at the Max Planck institute in Germany. In the independent phase, I aim to understand how two classes of conserved kinesin superfamily proteins (kinesins-13 and 8) mediate the disassembly of MTs, acting as MT depoymerases. A hypothesis has been developed about how the motor domains may force tubulin at MT ends to curve leading to MT disassembly. I intend to determine structures for their motor domain in action, while bound to tubulin dimer to understand the mechanism of MT depolymerization. I also intend to determine the role of conservd features in their depolymerase activity. I will then explore the antagonism and interaction between MT polymerase and depolymerases in interphase and its regulation upon the onset of mitotic cell division. This antagonism leads to slow MT dynamics in interphase and fast MT dynamics in mitosis. A direct role of phosphorylation by mitotic kinases will be explored during the transition between interphase and mitosis. Defects in the regulation of cell division are directly linked to human genetic defects and cancers. The results from this study will broaden our understanding of this regulation and provide new targets for cancer therapy.

描述（由申请人提供）：本研究的目的是了解由保守蛋白质家族介导的微管（MT）聚合和解聚的分子机制。这些蛋白质在MT+末端充当微管蛋白聚合酶和解聚酶。在Stephen C博士的指导下，Harrison在哈佛医学院的研究中，我利用电子显微镜和生物化学开发了一个分子模型，用于研究芽殖酵母Stu 2二聚体和非洲爪蟾XMAP 215单体如何通过其保守的N-末端TOG结构域结合微管蛋白二聚体。我已经确定了结构的TOG域使用X-射线晶体学和显示一个狭窄的保守表面上的TOG桨状结构的微管蛋白结合界面。我的工作提出了一个假设，如何TOG域增强微管蛋白聚合在MT加结束。我们目前正在研究与微管蛋白二聚体结合的TOG结构域的结构。在指导阶段，我将确定TOG结构域-微管蛋白二聚体复合物的结构，以了解TOG结构域如何结合和增强微管蛋白组装。我还打算探索不同的TOG域的作用，在提高MT组装使用显微镜方法与安东尼海曼和乔纳森霍华德在马克斯普朗克研究所在德国的实验室合作。在独立阶段，我的目标是了解两类保守的驱动蛋白超家族蛋白（驱动蛋白-13和8）介导的拆卸MT，作为MT depolymerases。关于运动域如何迫使微管蛋白在MT末端弯曲导致MT解体的假说已经被提出。我打算确定其运动域的结构，同时结合微管蛋白二聚体，以了解MT解聚的机制。我还打算确定保守特征在解聚酶活性中的作用。然后，我将探讨MT聚合酶和解聚酶之间的拮抗作用和相互作用的间期和有丝分裂细胞分裂的开始后，其调节。这种拮抗作用导致在间期的慢MT动力学和在有丝分裂中的快MT动力学。有丝分裂激酶磷酸化的直接作用将探讨在间期和有丝分裂之间的过渡。细胞分裂调节的缺陷与人类遗传缺陷和癌症直接相关。这项研究的结果将拓宽我们对这种调控的理解，并为癌症治疗提供新的靶点。