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.

描述：核糖体是进化上保守的分子机器 用来合成蛋白质。在真核细胞中，核糖体是在生物物理学中产生的 独特的亚核隔间，即核仁，通过能量驱动的级联 事件。这一过程是许多遗传性疾病的基础，也是 抗癌疗法。但是，网络的物理交互需要 这一进程在很大程度上仍未确定特征。这个应用程序将描述一个密钥 称为R2TP的分子复合体，在核糖体生产的早期通过促进 几种核糖体生产酶的组装。R2TP拥有广泛的客户基础， 与一种通用的蛋白质折叠伴侣热休克蛋白90(Hsp90)合作， 并将客户送到核仁。LI实验室已经确定了有效的方法来 对R2TP及其与客户端的交互进行提纯、重构和结构化表征 通过使用一套强大的结构生物学和生化方法来研究蛋白质。长的- 学期目标是了解R2TP/Hsp90介导的组装的分子基础 并揭示其在核糖体生产中作用的分子基础。二 设计了具体的目标来1)剖析R2TP的客户端绑定和发布周期，并 它受核苷酸的调节和核仁的液体不混溶；2)表征 在分子水平上，R2TP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和 来解释那些当前正在使用的人的行为。李实验室已经组建了一个团队 在x射线结晶学方面拥有互补专业知识的科学家，高通量 电子冷冻显微镜、质谱学、生物物理学和蛋白质生物化学 以最大限度地增加成功的机会，同时降低风险。 相关性：在多种癌细胞中，核糖体的产生加速。 相应地，R2TP/Hsp90及其客户在这些细胞中过表达。更多 已经研究或正在进行临床试验的抗癌药物超过15种 参与这一途径的靶分子。虽然在细胞死亡分析中有效，但 许多化合物的药理基础仍有待解释，一些化合物已有进展 细胞的抵抗力。这项拟议的研究提供了一个平台来测试 这些药物在细胞外并针对以前未知的部位或过程。