Structural Basis for RCC1 Directed Recruitment of Ran GTPase to Chromatin

RCC1 定向招募 Ran GTP 酶至染色质的结构基础

• 批准号: 8311671
• 负责人: SONG TAN
• 金额: $ 29.88万
• 依托单位: PENNSYLVANIA STATE UNIVERSITY, THE
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-08-01 至 2013-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8311671
• 关键词: Address; Affinity; Binding; Biochemical; Cell Nucleus; Cell division; Cell physiology; Cells; Chromatin; Chromosomes; Complement; Complex; Cytoplasm; DNA; Data; Docking; Ensure; Enzymes; Eukaryotic Cell; Fluorescence Resonance Energy Transfer; GTP Binding; Genetic; Genome; Goals; Guanine; Guanine Nucleotide Exchange Factors; Guanosine; Herpesviridae; Histone H4; Histones; Interferometry; Investigation; Kaposi Sarcoma; Malignant Neoplasms; Maps; Mediating; Methods; Mitosis; Mitotic; Mitotic spindle; Modeling; Molecular; Molecular Models; Mutation; Neutrons; Nuclear Pore Complex; Nucleosome Core Particle; Nucleosomes; Nucleotides; Peptides; Physical condensation; Positioning Attribute; Procedures; Process; Protein Binding; Proteins; Recruitment Activity; Reporting; Resolution; Roentgen Rays; Role; Running; Solutions; Structural Models; Structure; System; Testing; Viral; Viral Genome; abstracting; base; daughter cell; electron density; improved; insight; macromolecule; molecular modeling; nucleocytoplasmic transport; ran GTP-Binding Protein; research study

Project Summary/Abstract A eukaryotic cell must be able to transport macromolecules directionally between its nucleus and cytoplasm, and to divide the cell through mitosis. These fundamental processes are controlled by localizing the small Ran guanosine triphosphatase (GTPase) protein in its GDP or GTP bound state within the cytoplasm or the nucleus respectively, and by generating a gradient of RanGTP around the chromosomes. The spatial localization of RanGTP in the nucleus is achieved through chromatin bound RCC1 (regulator of chromosomal condensation) protein. RCC1 recruits Ran to the chromosomes and promotes the exchange of RanGDP for RanGTP, thereby creating a high concentration of RanGTP around chromosomes. We currently lack a molecular understanding of how RCC1 binds to the nucleosome and how RCC1 recruits Ran to the nucleosome, despite the critical importance of these interactions for basic cellular processes. Our overall goal is therefore to develop atomic models which describe how RCC1 and Ran bind to the nucleosome core particle. Our specific aims are: 1. Define how RCC1 binds to nucleosomes through biochemical methods. We will challenge structural models for how RCC1 interacts with the nucleosome through pulldown, biolayer interferometry and fluorescence resonance energy transfer experiments. 2. Determine the structure of the RCC1/nucleosome complex. We will use single crystals of the RCC1/nucleosome complex we have grown to determine the structure of the complex. These crystallographic studies will be complemented with small angle X-ray and neutron scattering experiments to provide a solution structure of the complex. 3. Determine how chromatin-bound RCC1 binds to and activates Ran. We will test models for the Ran/RCC1/nucleosome complex by analyzing the effects of directed mutations on binding of Ran to the RCC1/nucleosome complex and on Ran's nucleotide exchange activity in the presence of RCC1 and the nucleosome.

项目摘要/摘要 真核细胞必须能够在其核之间方向转运大分子 细胞质，并通过有丝分裂分裂细胞。这些基本过程是通过本地化来控制的 在细胞质或GTP结合状态下，小型RAN RAN鸟嘌呤三磷酸酶（GTPase）蛋白在细胞质或GTP结合状态或 细胞核分别通过在染色体周围产生RANGTP的梯度。空间 RANGTP在细胞核中的定位是通过染色质结合的RCC1（染色体调节剂）实现的 凝结）蛋白质。 RCC1招募染色体并促进Rangdp的交换 Rangtp，从而在染色体周围产生高浓度的Rangtp。 目前，我们对RCC1如何与核小体结合以及RCC1的结合缺乏分子理解 尽管这些相互作用对于基本细胞的重要性至关重要，但招募人员仍奔向核小体 过程。因此，我们的总体目标是开发原子模型，以描述RCC1和运行如何结合 核小体核心粒子。我们的具体目的是： 1。定义RCC1如何通过生化方法与核小体结合。我们将挑战结构 RCC1如何通过下拉，Biolayer干涉法和核小体相互作用的模型 荧光共振能量传递实验。 2。确定RCC1/核小体复合物的结构。我们将使用 RCC1/核小体络合物我们已经成长以确定复合物的结构。这些晶体学 研究将与小角度X射线和中子散射实验相辅相成，以提供溶液 复合物的结构。 3。确定结合染色质的RCC1如何与RAN结合并激活RAN。我们将测试模型 通过分析定向突变对RAN与RAN的影响，RAN/RCC1/核小体复合物 RCC1/核小体复合物以及在RCC1和RCC1和RAN的核苷酸交换活动上 核小体。