Structure and Function of the Parkinson's disease associated protein LRRK2

帕金森病相关蛋白 LRRK2 的结构和功能

基本信息

批准号：
8887486
负责人：
Quyen Quoc Hoang
金额：
$ 29.96万
依托单位：
INDIANA UNIV-PURDUE UNIV AT INDIANAPOLIS
依托单位国家：
美国
项目类别：
财政年份：
2015
资助国家：
美国
起止时间：
2015-04-01 至 2020-03-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8887486
关键词：
Active Sites Address Adverse effects Affect Affinity Alzheimer&apos s Disease Amino Acid Sequence Architecture Area Bacteria Binding Binding Sites Biochemical Chemicals Clinic Complex Disease Drug Targeting Enzymes GTP-Binding Proteins Genes Guanosine Triphosphate Guanosine Triphosphate Phosphohydrolases Hydrolysis Impairment Investigation Lead Length Light Location Magnesium Maps Mediating Metals Molecular Molecular Conformation Mutation Nerve Degeneration Neurodegenerative Disorders Output Parkinson Disease Parkinson&apos s Disease Pathway Pathogenesis Phosphotransferases Procedures Process Proteins Rattus Recombinants Regulation Resolution Sampling Signal Pathway Signal Transduction Site Structure System Therapeutic Translations Work alpha synuclein base biophysical analysis drug development effective therapy inhibitor/antagonist insight leucine-rich repeat kinase 2 magnesium ion mutant novel polypeptide public health relevance therapy development treatment strategy

项目摘要

DESCRIPTION (provided by applicant): Here we seek to understand how structure and interactions between the domains of LRRK2 regulate its activities, and also how Parkinson's disease-associated mutations affect these processes. Those areas of study are significant because LRRK2 holds the strongest promise for developing effective treatments for Parkinson's disease (PD), for which there is currently none. Mutations in LRRK2 leading to disease clearly perturb LRRK2 kinase activity; however, the mechanism in doing so is complex as mutation sites remote from the kinase domain also exert similar effects on kinase activity. The major roadblock in unraveling these mechanisms had been the lack of protein samples amenable for detail biochemical and biophysical studies. We have overcome that by developing procedures that yield highly purified samples amenable for detail studies. Here we use these samples to study the structure and function of LRRK2, to tease out how its activities are regulated, and to define its mechanism in disease pathogenesis. The three areas to be investigated in this project are: 1. To determine the effects of PD-associated mutations on the structure and function of the Roc domain of LRRK2. We have shown that the PD-mutation R1441H impairs GTPase activity of the Roc domain of LRRK2. This impairment occurs through distorting the active-site, altering the conformation of the switch regions, or impeding the binding of magnesium ions. 2. To determine the GTPase activation domain of Roc and to define the structural basis for GTPase regulation. We have shown that the Roc domain of LRRK2 has low intrinsic GTPase activity, and we have seen that it could be activated 30-fold by an unknown domain within LRRK2. We will use a combination of biochemical and structural studies to identify and define this activation mechanism. 3. To define global structural interactions between the domains within LRRK2. Mutations are found in various parts of LRRK2, but they all affect kinase activity which can be several domains away from the mutation sites, thus indicating that these domains interact with one another working in concert to exert a common function. We will map the spatial arrangement of all domains within LRRK2 to gain insight into how they work together.

描述（由申请人提供）：在这里，我们试图了解 LRRK2 结构域之间的结构和相互作用如何调节其活性，以及帕金森病相关突变如何影响这些过程，这些研究领域非常重要，因为 LRRK2 具有最强的前景。开发帕金森病 (PD) 的有效治疗方法，目前尚无 LRRK2 突变导致疾病明显扰乱 LRRK2 激酶活性；然而，这样做的机制很复杂，因为突变位点远离。激酶结构域也对激酶活性产生类似的影响。阐明这些机制的主要障碍是缺乏适合详细生化和生物物理研究的蛋白质样品，我们通过开发可产生适合详细研究的高度纯化样品的程序克服了这一问题。在这里，我们使用这些样本来研究 LRRK2 的结构和功能，梳理其活性如何受到调节，并确定其在疾病发病机制中的机制。本项目要研究的三个领域是： 1. 确定 LRRK2 的影响。 LRRK2 Roc 结构域的 PD 相关突变我们已经证明，PD 突变 R1441H 会损害 LRRK2 Roc 结构域的 GTPase 活性，这种损害是通过扭曲活性位点、改变其构象而发生的。开关区域，或阻碍镁离子的结合 2. 确定 Roc 的 GTPase 激活结构域并定义 GTPase 调节的结构基础。 LRRK2 的结构域具有较低的内在 GTP 酶活性，我们发现它可以被 LRRK2 内的未知结构域激活 30 倍。我们将结合生化和结构研究来识别和定义这种激活。 3. 定义 LRRK2 内结构域之间的整体结构相互作用突变存在于 LRRK2 的各个部分，但它们都会影响距离突变位点数个结构域的激酶活性，从而表明这些结构域彼此相互作用。我们将绘制 LRRK2 内所有域的空间排列图，以深入了解它们如何协同工作。