UNFOLDED STATES AND FOLDING PATHWAYS BY NMR

通过 NMR 观察未折叠状态和折叠通路

• 批准号: 2910402
• 负责人: HELEN JANE DYSON
• 金额: $ 22.85万
• 依托单位: SCRIPPS RESEARCH INSTITUTE, THE
• 依托单位国家: 美国
• 财政年份: 1998
• 资助国家: 美国
• 起止时间: 1998-05-01 至 2002-04-03
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2910402
• 关键词: DNA binding protein; Escherichia coli; bacterial proteins; conformation; corticosteroid receptors; molecular chaperones; myoglobin; nuclear magnetic resonance spectroscopy; plastocyanin; protein denaturation; protein folding; protein structure function; stop flow technique

DESCRIPTION (Adapted from applicant's abstract): Studies of the structure and dynamics of unfolded states of proteins have assumed great importance for the insights they provide into the initiation of protein folding. In addition, a number of disease states from defects in the folding and stability of specific proteins and an have recently been shown to arise increasing number of key intracellular proteins appear to be unfolded in their biologically functional states. The major goals of this project are focused on the elucidation of folding pathways and the structural characterization of unfolded states. There are three major specific aims, representing an integrated approach that utilizes recently-developed NMR methods: (1) Heteronuclear NMR methods will be used to obtain a quantitative description of the conformations sampled by two unfolded proteins, acid-unfolded apomyoglobin and low-salt-unfolded apoplastocyanin, which contain almost purely a single type of secondary structure in the folded state. The unfolded states are tractable according to preliminary results, and apoplastocyanin in particular has the added advantage that the unfolded form is present under "folding" conditions, neutral pH and ambient temperature, making the results highly applicable-to the protein folding process in vivo and in vitro. A detailed study of backbone structural propensities and polypeptide dynamics in the unfolded state of these two proteins will be made, as well as of denaturants on the structure in the unfolded state. (2) The folding of apoplastocyamin is slow on the NMR timescale and an MR study of the folding process in real time is proposed. The early, fast folding steps will be followed by multiple-step stopped-flow fluorescence and hydrogen exchange pulse labeling. 3) A long-term goal is to extend the work on the unfolded state to include studies of interactions with chaperones. The interaction of unfolded proteins and protpetides will be studies with a small domain from the E. coil chaperone protein DnaJ, which has been shown to bind zinc and to resemble the DNA-binding domain of glucocorticoid receptors. This surprising result typifies the new information that is constantly being uncovered in the protein folding and assembly a continual source of new and important information.

描述（改编自申请人摘要）：结构研究