Structural Biology Studies of Ribosome Biogenesis Network

核糖体生物发生网络的结构生物学研究

• 批准号: 10249225
• 负责人: Hong Li
• 金额: $ 40.91万
• 依托单位: FLORIDA STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-09-10 至 2023-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10249225
• 关键词: ATP phosphohydrolase; Affinity; Antineoplastic Agents; Architecture; Basic Science; Binding; Binding Proteins; Biochemical; Biogenesis; Biological Assay; Biology; Biophysics; C-terminal; Calorimetry; Cell Death; Cell Nucleolus; Cell Proliferation; Cells; Chemicals; Client; Clinical Trials; Complex; Cryoelectron Microscopy; Data; Defect; Deuterium; Disease; Environment; Environmental Impact; Enzymes; Eukaryotic Cell; Event; Evolution; Excision; Functional disorder; Genetic Diseases; Goals; Heat-Shock Proteins 90; Hydrogen; Individual; Laboratories; Learning; Liquid substance; Malignant Neoplasms; Mass Spectrum Analysis; Mediating; Methods; Molecular; Molecular Chaperones; Molecular Conformation; Molecular Machines; Nucleotides; Pathway interactions; Pharmacology; Physiological; Process; Production; Property; Protein Analysis; Protein Biochemistry; Protein Biosynthesis; Proteins; RNA; Regulation; Research; Resistance development; Resolution; Ribosomal RNA; Ribosomes; Risk; Role; Sampling; Scientist; Site; Small Nucleolar RNA; Small Nucleolar Ribonucleoproteins; Structure; Surface; System; Tail; Testing; Therapeutic; Therapeutic Studies; Thermodynamics; Titrations; Work; X-Ray Crystallography; Yeasts; analytical ultracentrifugation; anti-cancer therapeutic; base; biophysical techniques; cancer cell; crosslink; design; drug action; experience; improved; in vivo; interfacial; overexpression; particle; protein folding; reconstitution; structural biology; success; therapeutic development

Description: Ribosome is the evolutionarily conserved molecular machine responsible for synthesis of proteins. In eukaryotic cells, ribosome is produced in a biophysically distinct subnuclear compartment, the nucleolus, through a cascade of energy-driven events. This process underlies a number of genetic diseases and is a major target for anticancer therapeutics. However, the physical interactions of the network required for this process remain largely uncharacterized. This application will characterize a key molecular complex called R2TP that acts early in ribosome production by facilitating assembly of several ribosome production enzymes. R2TP has a wide client base, collaborates with a general protein folding chaperone, heat shock protein 90 (Hsp90), and delivers clients to the nucleolus. The Li laboratory has identified effective methods to purify, reconstitute, and structurally characterize R2TP and its interaction with a client protein by use of a powerful set of structural biology and biochemical methods. The long- term goal is to understand the molecular basis for R2TP/Hsp90-mediated assembly process and uncover the molecular basis for its function in ribosome production. Two specific aims are designed to 1) dissect the client binding and release cycle of R2TP and its regulation by nucleotides and the liquid immiscibility of the nucleolus; 2) characterize at a molecular level how R2TP impacts pre-ribosome structure purified from yeast cells. Results of this study promise to reveal new interfacial surfaces for anticancer drugs and to explain the action of those currently in use. The Li laboratory has assembled a team of scientists with complementary expertise in x-ray crystallography, high-throughput electron cryomicroscopy, mass spectrometry, biophysics, and protein biochemistry in order to maximize the chance of successes while mitigating risks. Relevance: Ribosome production is accelerated in multiple cancer cells. Correspondingly, R2TP/Hsp90 and their clients are overexpressed in these cells. More than fifteen types of anticancer drugs have been investigated or in clinical trials that target molecules involved in this pathway. Though effective in cell-death assays, the pharmacological basis of many compounds remains to be explained and some develop resistance in cells. The proposed research provides a platform to test the actions of these drugs outside the cell and to target previously unknown sites or processes.

核糖体是进化上保守的分子机器， 用于蛋白质的合成。在真核细胞中，核糖体是在一个生物蛋白酶促反应中产生的。 不同的亚核区室，核仁，通过级联的能量驱动 事件这一过程是许多遗传疾病的基础，也是治疗的主要目标。 抗癌疗法然而，所需的网络的物理交互， 这一过程在很大程度上仍然没有特征。此应用程序将描述一个关键 一种称为R2 TP的分子复合物，在核糖体产生的早期起作用， 几种核糖体生产酶的组装。R2 TP拥有广泛的客户群， 与一般蛋白质折叠分子伴侣，热休克蛋白90（Hsp 90）， 并将客户端传送到细胞核。Li实验室已经确定了有效的方法， 纯化、重组和结构表征R2 TP及其与客户端的相互作用 蛋白质通过使用一套强大的结构生物学和生物化学方法。很长的- 本学期的目标是了解R2 TP/Hsp 90介导的组装的分子基础 处理并揭示其在核糖体产生中功能的分子基础。两 具体目标是：1）剖析R2 TP的客户端绑定和发布周期， 它的调节核苷酸和液体不溶性的核仁; 2）表征 在分子水平上，R2 TP如何影响从酵母细胞中纯化的前核糖体结构。 这项研究的结果有望揭示抗癌药物的新界面， 来解释目前使用的那些的作用。李实验室组建了一个团队 在X射线晶体学、高通量、 电子低温显微镜，质谱，生物物理学和蛋白质生物化学， 以最大限度地提高成功的机会，同时降低风险。 相关性：核糖体的产生在多种癌细胞中加速。 相应地，R2 TP/Hsp 90及其客户端在这些细胞中过表达。更 超过15种抗癌药物已经被研究或在临床试验中， 参与这一途径的靶分子。尽管在细胞死亡测定中有效， 许多化合物的药理学基础仍有待解释，有些化合物正在开发中 细胞的抵抗力。拟议的研究提供了一个平台，以测试的行动， 这些药物的细胞外，并针对以前未知的网站或过程。