Mechanism of GroEL-assisted Protein Folding

GroEL 辅助蛋白质折叠机制

• 批准号: 7222693
• 负责人: LINGLING CHEN
• 金额: $ 27.91万
• 依托单位: TRUSTEES OF INDIANA UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2003
• 资助国家: 美国
• 起止时间: 2003-05-01 至 2010-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7222693
• 关键词: Address; Affinity; Alzheimer' s Disease; Apical; Binding; Binding Proteins; Biochemical; Biology; Cataract; Cell physiology; Complex; Coupled; Crystallography; Cystic Fibrosis; Environment; Escherichia coli; Event; Fluorescence Spectroscopy; Goals; Hydrolysis; Knowledge; Lead; Mediating; Methods; Modeling; Molecular; Molecular Chaperones; Molecular Conformation; Nature; Nucleotides; Pathway interactions; Peptide Library; Peptides; Phage Display; Prion Diseases; Process; Property; Protein Biosynthesis; Proteins; Range; Research; Resolution; Roentgen Rays; Role; Series; Staging; Structure; Substrate Interaction; Techniques; X-Ray Crystallography; base; combinatorial; design; human disease; in vivo; molecular assembly/self assembly; novel therapeutics; polypeptide; prevent; protein folding; structural biology

DESCRIPTION (provided by applicant): Molecular chaperones are involved in a wide range of essential cellular processes: protein synthesis, molecular assembly, translocation, degradation, and folding. The E. coli molecular chaperone GroEL, along with its co-chaperone GroES, increases the efficiency of protein folding in vivo, using an ATP-driven mechanism. In the GroEL-facilitated folding process, first GroEL sequesters the aggregation-prone nonnative forms of proteins from the complex cellular environment within its central cavity. Then with the actions of ATP binding/hydrolysis and GroES binding, the protein is allowed to carry out its initial folding events within an isolated "folding chamber" formed by GroEL/GroES. The long-term goals of this proposal are to elucidate the structural features of the interaction of GroEL and substrate proteins, to understand the mechanism of GroEL-facilitated protein folding in a structural context, and to further structural knowledge of molecular chaperones function in general. Various biochemical and biophysical techniques, including phage display, fluorescence spectroscopy/polarization and X-ray crystallography, will be used to determine how GroEL recognizes the substrate proteins, and to understand the energetics of ATP binding/hydrolysis. Three specific aims are to: 1) Select for small peptides that interact with the substrate binding domain of GroEL using a phage display method. 2) Study the interplay of GroEL-substrate by determining the structures of the substrate-trapped GroEL assemblies using X-ray protein crystallography and NMR, and carry out structure-guided mutational studies. 3) Create model polypeptide substrates for GroEL based on the peptides selected in 1) and use them to study the functional role of nucleotide binding/hydrolysis and the mechanism of GroEL-assisted protein folding. Knowledge of protein folding and the role of molecular chaperones in facilitating the folding process will contribute to a better understanding of folding-related human diseases, such as Cystic Fibrosis, Alzheimer's, Prion diseases and cataracts, at the molecular level, and could lead to the design of novel therapeutic approaches.

描述（由申请人提供）：分子伴侣参与广泛的必需细胞过程：蛋白质合成，分子组装，易位，降解和折叠。大肠杆菌分子伴侣凹槽及其副酮凹槽，使用ATP驱动的机制提高了体内蛋白质折叠的效率。在凹槽的折叠过程中，首先是凹槽将其中央腔内复杂细胞环境中的蛋白质易发的非本质形式隔离。然后，随着ATP结合/水解和GROES结合的作用，蛋白质被允许在凹槽/凹槽形成的分离的“折叠室”中执行其初始折叠事件。该提案的长期目标是阐明凹槽和底物蛋白相互作用的结构特征，以了解结构背景下的凹槽 - 硅化蛋白折叠的机理，并进一步了解分子伴侣的结构知识。各种生化和生物物理技术，包括噬菌体显示，荧光光谱/极化和X射线晶体学，将用于确定Groel如何识别底物蛋白，并了解ATP结合/水解的能量学。三个特定的目的是：1）选择使用噬菌体显示方法与凹槽底物结合域相互作用的小肽。 2）通过使用X射线蛋白质晶体学和NMR确定粘胶底物的相互作用，并进行结构引导的突变研究。 3）创建基于1）中选择的凹槽的模型多肽底物，并使用它们来研究核苷酸结合/水解的功能作用以及Groel辅助蛋白折叠的机制。了解蛋白质折叠的知识以及分子伴侣在促进折叠过程中的作用，将有助于更好地理解与折叠相关的人类疾病，例如囊性纤维化，阿尔茨海默氏症，阿尔茨海默氏症，prion疾病和caparactes，并在分子水平上，并可能导致新型治疗方法的设计。