BIOPHYSICAL PROPERTIES OF PRION PROTEIN OLIGOMERS

朊病毒蛋白低聚物的生物物理特性

• 批准号: 7277489
• 负责人: WITOLD K SUREWICZ
• 金额: $ 35.78万
• 依托单位: CASE WESTERN RESERVE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2007
• 资助国家: 美国
• 起止时间: 2007-06-01 至 2012-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7277489
• 关键词: Accounting; Address; Amino Acids; Amyloid; Amyloid Fibrils; Animals; Binding; Bovine Spongiform Encephalopathy; Brain; C-terminal; Complement; Coupled; Data; Deuterium Exchange Measurement; Development; Digestion; Disease; Electron Spin Resonance Spectroscopy; Endopeptidase K; Endopeptidases; Engineering; Epidemic; Goals; Helix (Snails); Human; In Vitro; Infectious Agent; Label; Mass Spectrum Analysis; Measurement; Mesocricetus auratus; Methods; Modeling; Molecular; Molecular Biology; Molecular Models; N-terminal; Neurodegenerative Disorders; Numbers; Peptide Hydrolases; PrP amyloid; PrP variant; PrPSc Proteins; Prion Diseases; Prions; Procedures; Process; Property; Protein Isoforms; Proteins; Public Health; Reaction; Recombinants; Reporting; Research; Resistance; Resolution; Site; Spin Labels; Structural Models; Structure; Techniques; Technology; Testing; Theoretical model; Time; Variant; amyloid structure; base; ear helix; experience; glycosylation; in vivo; pathogen; prion hypothesis; programs; protein misfolding cyclic amplification; recombinant PrP; research study; single molecule; three dimensional structure

The long-term goal of this component of the Program Project is to elucidate the molecular basis of the pathogenic process in transmissible spongiform encephalopathies (TSEs), a group of fatal neurodegenerative diseases that afflict humans and animals. The protein-only model postulates that the infectious pathogen responsible for these diseases is a misfolded form of the prion protein, PrPSc, which self-propagates by binding to normal prion protein and catalyzing its conversion to the pathogenic form. TSE diseases have emerged as a major public health issue following recent epidemics of bovine spongiform encephalopathy (BSE) and indications that BSE might have crossed the species barrier to cause variant Creutzfeldt-Jakobdisease in humans. The main focus of this project is on understanding structural properties of various abnormally folded forms of the recombinaht prion protein. The first specific aim is to determine the structure of amyloid fibrils formed by the recombinant prion protein in vitro at a resolution close to amino acid residue. This will be accomplished using a number of newly emerged biophysical techniques that provide site- specific information about local protein mobility and accessibility and measurements of intermolecular and intramolecular distances between specific sites within the prion protein amyloid. Another specific aim is to expand and optimize the recently developed technique of protein misfolding by cyclic amplification (PMCA) to accomplish efficient brain PrP^-templated conversion of bacterially-expressed recombinant prion protein to proteinase K-resistantform, and assess the infectivity of this product in experimental animals. Finally, building on the experience gained in structural studies with spontaneously formed prion protein amyloid (Specific Aim 1), we will determine structural organization of the recombinant PrP oligomers generated in brain PrPSc- templated PMCA reaction. Our hypothesis is that the basic folding motif of PrPSc is similar to that of PrP amyloid fibrils, though some differences which may account for especially high resistance of brain PrPSc to proteolytic digestion are expected.

该计划项目的这一组成部分的长期目标是阐明分子基础 一组致命的致命性脑病（TSE）中的致病过程 神经退行性疾病会困扰人类和动物。仅蛋白质模型假设 负责这些疾病的感染性病原体是prion蛋白的一种错误折叠的形式， PRPSC，通过与正常的pr蛋白结合并催化其转化为 致病形式。最近，TSE疾病已成为主要的公共卫生问题 牛海绵状脑病（BSE）的流行病以及BSE可能具有的迹象 越过物种屏障，引起人类的变体creutzfeldt-jakobdisease。主要重点 该项目的是了解各种异常折叠形式的结构特性 重组prion蛋白。第一个具体目的是确定淀粉样蛋白纤维的结构 在接近氨基酸残基的分辨率下，由重组prion蛋白在体外形成。这会 可以使用多种新出现的生物物理技术来实现，这些技术提供站点 - 有关局部蛋白质迁移率以及可访问性和测量的特定信息 蛋白质淀粉样蛋白内特定部位之间的分子间和分子内距离。 另一个具体目的是扩展和优化最近开发的蛋白质技术 通过循环放大（PMCA）折叠以完成有效的大脑PRP^模拟转换 细菌表达的重组prion蛋白与蛋白酶K-抗抗性的蛋白质，并评估 该产品在实验动物中的感染力。最后，基于获得的经验 自发形成的prion蛋白淀粉样蛋白（特定目标1）的结构研究，我们将 确定大脑PRPSC-产生的重组PRP低聚物的结构组织 模板PMCA反应。我们的假设是，PRPSC的基本折叠基序与 PRP淀粉样蛋白原纤维，尽管有些差异可能特别是高度高的抗性 脑PRPSC至蛋白水解消化。