Learning About Protein Unfolded States From Heterodimeric Fragment Complementation

从异二聚体片段互补中了解蛋白质未折叠状态

• 批准号: 0118252
• 负责人: Maria Luisa Tasayco
• 金额: $ 33万
• 依托单位: CUNY City College
• 依托单位国家: 美国
• 项目类别: Continuing grant
• 财政年份: 2001
• 资助国家: 美国
• 起止时间: 2001-09-01 至 2005-01-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=0118252&HistoricalAwards=false
• 关键词: Learning; About; Protein; Unfolded; States

This is a study of a family of complementary fragments from oxidized E. coli thioredoxin (Trx) and their heterodimeric reassemblies around different interfaces. The values of estimated buried surface from DSC studies of intrinsically unstructured protein fragments will be correlated with NMR studies of conformational preferences and rigidity of the backbone at the residue level. This project combines three areas of expertise (protein fragment complementation, calorimetry of proteins, and NMR analysis of structure and dynamics of proteins) and centers on two main hypotheses: (1) Isolated Trx fragments which comprise the regions of the native 2 and 4 strands have substantial nonlocal interactions between them, while remaining intrinsically unstructured. (2) The differences in delta G of folding/binding among the different heterodimeric reassemblies are mainly due to differences in the degree of folding and rigidity in the isolated fragments. In order to test these hypotheses, the delta G of folding/binding and low-resolution structural characterization of the initial and final states for a family of complementary fragments will be determined. High sensitivity DSC studies of selected isolated fragments and heterodimeric reassemblies will be undertaken, and NMR studies of the structures of the initial and final states of selected complementary fragments will be done to study the dynamics of such a selected pair in both states. The long-term objective of these studies is to characterize the conformational space of the unfolded state of proteins and understand protein-protein interactions covering a range of affinities and degree of folding associated with binding through a synergistic interaction between experimenation and theory.

这是一项针对氧化大肠杆菌硫氧还蛋白 (Trx) 的互补片段家族及其在不同界面周围的异二聚体重组的研究。 本质上非结构化蛋白质片段的 DSC 研究估计的埋藏表面值将与残基水平主链的构象偏好和刚性的 NMR 研究相关。该项目结合了三个专业领域（蛋白质片段互补、蛋白质量热法以及蛋白质结构和动力学的 NMR 分析），并以两个主要假设为中心：（1）包含天然 2 和 4 链区域的分离 Trx 片段在它们之间具有大量非局部相互作用，同时保持本质上的非结构化。 (2)不同异二聚体重组体之间折叠/结合δG的差异主要是由于分离片段的折叠程度和刚性的差异。 为了测试这些假设，将确定互补片段家族的折叠/结合的δG以及初始和最终状态的低分辨率结构表征。 将对选定的分离片段和异二聚体重组进行高灵敏度 DSC 研究，并对选定的互补片段的初始和最终状态的结构进行 NMR 研究，以研究此类选定对在两种状态下的动力学。这些研究的长期目标是表征蛋白质未折叠状态的构象空间，并通过实验和理论之间的协同相互作用了解蛋白质-蛋白质相互作用，涵盖与结合相关的一系列亲和力和折叠程度。