Structure and assembly of the fungal prion HET-s

真菌朊病毒HET-s的结构和组装

• 批准号: 8201967
• 负责人: William N Wan
• 金额: $ 2.62万
• 依托单位: VANDERBILT UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-09-30 至 2014-09-29
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8201967
• 关键词: Agreement; Alzheimer' s Disease; Amino Acid Sequence; Amyloid; Amyloid Fibrils; Asparagine; Behavior; Biological Models; Bovine Spongiform Encephalopathy; Brain; Cattle; Characteristics; Chronic Wasting Disease; Complex; Creutzfeldt-Jakob Syndrome; Crystallography; Data; Deer; Development; Disease; Electron Microscopy; Elements; Fiber; Fluorescence Spectroscopy; Genetic Polymorphism; Goals; Higher Order Chromatin Structure; Human; Hybrids; In Vitro; Indium; Infectious Agent; Joints; Link; Measures; Methodology; Methods; Modeling; Molecular; Molecular Structure; Nature; Neurodegenerative Disorders; Parkinson Disease; Pathology; Peptides; Phenotype; Podospora anserina; Prion Diseases; Prions; Process; Property; Proteins; Public Health; Publishing; Recording of previous events; Resolution; Roentgen Rays; Running; Scrapie; Sheep; Site-Directed Mutagenesis; Sodium Chloride; Structural Models; Structure; Symptoms; System; Techniques; Therapeutic; Variant; Work; X-Ray Crystallography; amyloid formation; amyloid structure; analog; base; fungus; human disease; in vivo; insight; meetings; prion-like; protein aggregate; protein folding; solid state nuclear magnetic resonance; therapy design; tool; yeast prion

DESCRIPTION (provided by applicant): Prions are aberrantly folded proteins implicated in the invariably fatal transmissible spongiform encephalopathies (TSEs). TSEs include scrapie in sheep, bovine spongiform encephalopathy (BSE) in cattle, chronic wasting disease (CWD) in deer and elk, and several forms of Creutzfeldt-Jakob disease (CJD) in humans. In addition to the TSEs, it has become apparent that other neurodegenerative diseases, such as Alzheimer's and Parkinson's diseases, contain prion-like behavior in their pathologies. One common feature of these diseases is the aggregation of aberrantly folded proteins and peptides into amyloid fibrils. While the mechanism by which amyloid fibrils are involved in disease is unknown, differences in their molecular structures have been linked with differences in pathologies, a property called the strain phenomenon, which indicates that structure and pathology are interdependent. In order to understand the pathology of these diseases, it is imperative to determine the molecular structures of the implicated amyloids. However, this is a particularly difficult task owing to the heterogeneous and disordered nature of amyloid fibrils. In efforts to develop techniques, in addition to expanding our mechanistic understanding of amyloid formation, the proposed work aims to develop the prion forming domain of the functional fungal prion HET-s as a model system in studying the structure of pathological prions and prion-related amyloids. The HET-s prion forming domain, (218-289), is an ideal model system because infectious fibrils can be formed reproducibly and easily, but the domain still possesses a relatively complex fold comparable to those of pathologically relevant prions. The first aim will be to determine the structure of the infectious form of HET-s(218-289) using joint refinement methods between X-ray fiber diffraction and solid state NMR (ssNMR). The second aim will be to use site- directed mutagenesis to probe the necessity and redundancy of structural elements in correct folding of HET- s(218-289). The third aim will be to characterize infectious and several non-infectious species of HET-s(218- 289), which together can serve as an analog to the strain phenomenon. The methods of characterization will include X-ray fiber diffraction, fluorescence spectroscopy and electron microscopy. These aims will allow for a deeper understanding of the mechanism of prion folding as well as the critical elements necessary for correct folding. Characterization of HET-s(218-289) species, taken with the results of the first two aims, will provide insight into the interplay between environmental conditions and amino acid sequence that determine the final amyloid structure. While these studies cannot replace the need for studies of brain-derived prions, they provide a framework for understanding the mechanisms of prion folding and polymorphism. Preliminary results demonstrate the amenability of HET-s(218-289) to structure determination and biophysical characterization as well as its similarities to and subsequent extensibility in the study of pathological prions. PUBLIC HEALTH RELEVANCE: Prion diseases such as bovine spongiform encephalopathy (BSE) and Creutzfeldt-Jakob disease (CJD) as well as prion-related diseases such as Alzheimer's and Parkinson's diseases are a continually growing public health concern. The pathologies of these diseases are intricately associated with the molecular structures of aberrantly folded protein aggregates, termed amyloid, that are implicated in these diseases. The proposed studies aim to establish the fungal prion HET-s(218-289) as a model system in efforts to obtain useful structural and mechanistic information that will further advance efforts in the study of disease-associated amyloids.

描述（由申请人提供）：朊病毒是一种异常折叠的蛋白质，与致命的传染性海绵状脑病（TSE）有关。 TSE 包括羊痒病、牛海绵状脑病 (BSE)、鹿和麋鹿慢性消耗性疾病 (CWD) 以及人类多种形式的克雅氏病 (CJD)。除了 TSE 之外，其他神经退行性疾病，例如阿尔茨海默病和帕金森病，在其病理学中也包含类似朊病毒的行为，这一点已经变得很明显。这些疾病的一个共同特征是异常折叠的蛋白质和肽聚集成淀粉样原纤维。虽然淀粉样原纤维参与疾病的机制尚不清楚，但其分子结构的差异与病理学的差异有关，这种特性称为应变现象，表明结构和病理学是相互依赖的。为了了解这些疾病的病理学，必须确定相关淀粉样蛋白的分子结构。然而，由于淀粉样原纤维的异质性和无序性，这是一项特别困难的任务。在开发技术的过程中，除了扩大我们对淀粉样蛋白形成机制的理解外，拟议的工作还旨在开发功能性真菌朊病毒HET-s的朊病毒形成结构域，作为研究病理性朊病毒和朊病毒相关结构的模型系统。淀粉样蛋白。 HET-s 朊病毒形成结构域 (218-289) 是一个理想的模型系统，因为感染性原纤维可以重复且容易地形成，但该结构域仍然具有与病理相关朊病毒相当的相对复杂的折叠。第一个目标是利用 X 射线纤维衍射和固态 NMR (ssNMR) 之间的联合精化方法确定 HET-s(218-289) 感染性形式的结构。第二个目标是利用定点诱变来探讨结构元件在HET-s(218-289)正确折叠中的必要性和冗余性。第三个目标是表征 HET-s(218-289) 的传染性和几种非传染性物种，它们一起可以作为菌株现象的类似物。表征方法包括 X 射线纤维衍射、荧光光谱和电子显微镜。这些目标将使人们更深入地了解朊病毒折叠机制以及正确折叠所需的关键要素。 HET-s(218-289) 物种的表征，结合前两个目标的结果，将深入了解环境条件和决定最终淀粉样蛋白结构的氨基酸序列之间的相互作用。虽然这些研究不能取代对脑源性朊病毒研究的需要，但它们为理解朊病毒折叠和多态性机制提供了一个框架。初步结果证明了 HET-s(218-289) 适合结构测定和生物物理表征，以及其与病理性朊病毒研究的相似性和随后的可扩展性。 公共卫生相关性：牛海绵状脑病 (BSE) 和克雅氏病 (CJD) 等朊病毒疾病以及阿尔茨海默病和帕金森病等朊病毒相关疾病是日益严重的公共卫生问题。这些疾病的病理学与异常折叠的蛋白质聚集体（称为淀粉样蛋白）的分子结构密切相关，而淀粉样蛋白与这些疾病有关。拟议的研究旨在建立真菌朊病毒 HET-s(218-289) 作为模型系统，以努力获得有用的结构和机制信息，从而进一步推进疾病相关淀粉样蛋白的研究工作。