Single molecule analysis of nascent protein folding

新生蛋白质折叠的单分子分析

基本信息

批准号：
7570874
负责人：
Christian Kaiser
金额：
$ 8.94万
依托单位：
UNIVERSITY OF CALIFORNIA BERKELEY
依托单位国家：
美国
项目类别：
财政年份：
2008
资助国家：
美国
起止时间：
2008-12-01 至 2010-11-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7570874
关键词：
Alzheimer&apos s Disease Area Binding Biological Process Cells Chemicals Complex DNA Degenerative Disorder Development Disease Doctor of Philosophy Elements Environment Event Excision Fluorescence Free Ribosome Functional disorder Human Immobilization In Vitro Individual Label Laboratories Length Life Maps Masks Measurement Methodology Methods Modeling Modification Molecular Molecular Chaperones Nature Parkinson Disease Pathway interactions Peptides Principal Investigator Process Protein Biosynthesis Proteins Research Research Personnel Ribosomes Site Spectrum Analysis Stretching Students Tertiary Protein Structure Therapeutic Intervention Time Translating Translations base fascinate insight interest laser tweezer neglect novel polypeptide protein folding protein misfolding research study single molecule three dimensional structure tool

项目摘要

DESCRIPTION (provided by applicant): The experiments proposed here will analyze the folding of nascent proteins in an increasingly more complex and biologically relevant context. We will analyze how the folding of purified polypeptide fragments of increasing length, mimicking nascent polypeptides, differs from that of the corresponding full length proteins. Then, we will characterize the folding of individual nascent polypeptides in the context of translating ribosomes. This will allow us to assess how the folding energy landscape is influenced by the vectorial nature of protein synthesis and the unique environment of the ribosome. Finally, we will investigate how ribosome-associated chaperones modulate protein folding of both free and ribosome-bound nascent chains. These findings will allow us to map the folding energy landscapes of nascent proteins and provide insight into the fundamental principles that govern de novo protein folding. I have had a strong interest in the mechanism of de novo protein folding and its modulation by molecular chaperones since the beginning of my doctoral studies. Particularly, I have developed novel fluorescence-based methodologies to investigate de novo protein folding in the context of translation by the ribosome. In recent years, single molecule methodologies have become invaluable tools for the characterization of intrinsically heterogeneous protein folding pathways. Many of the applications that utilize optical tweezers to analyze biological processes with single molecule force spectroscopy have been developed in the laboratory of Prof. Bustamante. Thus, his laboratory provides an unequaled environment for me to become proficient in the utilization of optical tweezers and their application to protein folding. A postdoctorate in Prof. Bustamante's laboratory will enable me to build and operate optical tweezers and to study de novo protein folding with single molecule methods as an independent researcher. Ultimately, I would like to establish my own laboratory to engage PhD students and postdoctoral trainees in this fascinating area of research. RELEVANCE: Prevalent human disorders, such as Parkinson's and Alzheimer's diseases, are associated with protein misfolding. A more thorough understanding of protein folding and the function of molecular chaperones is imperative to gain insight into the pathophysiology underlying these disease states and may ultimately allow the development of effective therapeutic intervention.

描述（由申请人提供）：此处提出的实验将分析越来越复杂且与生物学相关的环境中新生蛋白的折叠。我们将分析长度增加，模仿新生多肽的纯化多肽片段的折叠与相应的全长蛋白的折叠方式不同。然后，我们将在翻译核糖体的背景下表征单个新生多肽的折叠。这将使我们能够评估如何受蛋白质合成的矢量性质和核糖体的独特环境的影响。最后，我们将研究核糖体相关的伴侣如何调节自由和核糖体结合的新生链的蛋白质折叠。这些发现将使我们能够绘制新生蛋白的折叠能景观，并洞悉控制从头蛋白质折叠的基本原理。自从我的博士研究开始以来，我对从头蛋白折叠的机制及其对分子伴侣的调节感有很大的兴趣。特别是，我开发了基于荧光的新方法，以研究核糖体翻译的背景下从头蛋白质折叠。近年来，单分子方法论已成为表征本质上异质蛋白折叠途径的宝贵工具。在Bustamante教授的实验室中，已经开发了许多利用光学镊子用单分子力光谱分析生物过程的应用。因此，他的实验室为我提供了一个无与伦比的环境，使我能够精通光学镊子及其在蛋白质折叠中的应用。 Bustamante教授的实验室中的博士后术会使我能够建立和操作光学镊子，并以独立研究人员的单分子方法研究从头蛋白质折叠。最终，我想建立自己的实验室，以在这一有趣的研究领域与博士生和博士后学员联系。相关性：普遍的人类疾病，例如帕金森氏症和阿尔茨海默氏症疾病，与蛋白质错误折叠有关。必须对蛋白质折叠和分子伴侣的功能进行更透彻的理解，这对于深入了解这些疾病状态的病理生理学是必不可少的，并最终可能允许发展有效的治疗干预措施。