Structures of Protein Aggregates by Solid-State NMR

通过固态 NMR 分析蛋白质聚集体的结构

• 批准号: 7931186
• 负责人: Chad M Rienstra
• 金额: $ 18.56万
• 依托单位: UNIVERSITY OF ILLINOIS AT URBANA-CHAMPAIGN
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-09-30 至 2010-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7931186
• 关键词: Abbreviations; Amyloid; Anisotropy; Argon; Behavior; Binding; Brain; Chad; Chemicals; Chemistry; Collaborations; Communities; Complex; Data Analyses; Data Set; Development; Diagnostic; Dimerization; Disease; Electron Microscopy; Event; Fluorescence; Human Resources; Kinetics; Knowledge; Label; Laboratories; Lecithin; Literature; Magic; Maintenance; Mass Spectrum Analysis; Measurement; Membrane; Methods; Micelles; Modeling; Modification; Molecular Conformation; Monitor; Mutation; N-terminal; NMR Spectroscopy; Oxygen; Parkinson Disease; Pathology; Phage Display; Phosphatidic Acid; Physiologic pulse; Physiological; Preparation; Proteins; Protocols documentation; Public Health; Regulation; Reporting; Research; Research Personnel; Research Project Grants; Resolution; Sampling; Sedimentation process; Side; Signal Transduction; Site; Sodium Dodecyl Sulfate; Solid; Solutions; Specificity; Spin Labels; Staging; Streptococcus IgG Fc-binding protein; Stress; Stretching; Structural Models; Structure; Surface; Synaptic Vesicles; Techniques; Technology; Testing; Thioflavin T; Variant; Vesicle; Wood material; alpha synuclein; base; design; dimer; early onset; endosulfine; globular protein; human disease; improved; in vitro Model; in vivo; instrumentation; interest; light scattering; mutant; neuron development; oxidation; planetary Atmosphere; programs; protein aggregate; protein aggregation; research study; scaffold; small molecule; solid state; solid state nuclear magnetic resonance; tool

Description: The long-term objective of this research project is to develop a detailed atomic-resolution structural and functional understanding of protein aggregation and fibrillation relevant to human disease, principally with studies of alpha-synuclein. Wild type alpha-synuclein and its mutants (A30P, E46K and A53T) associated with early-onset Parkinson's disease (PD) will be examined in several structural states, including protofibrils, fibrils and membrane-associatedcomplexes. We propose that the chemical details of these structures determine the natural physiological function of alpha-synuclein and, in the mutants or under environmental stress, contribute to PD pathology. Moreover, we propose that conversion among several structural states is an essential feature of synucleins; this structural plasticity is likely to be required for neuronal development and maintenance. To examine these structures, magic-angle spinning (MAS) solid- state NMR (SSNMR) experiments will be employed, elucidating details of covalent chemistry, conformation and dynamics that are inaccessible to other experimental techniques. Complementary analysis and characterization techniques will include kinetic measurements (by thioflavin T fluorescence, light scattering and sedimentation analysis), high-resolution mass spectrometry, atomic force and electron microscopy, solution NMR and phage display. We will leverage these technologies along with sample preparation and isotopic labeling strategies to improve our understanding of protein aggregation relevant to disease in the specific case of alpha-synuclein, including: (1) the fundamental biophysical principles of alpha-synuclein aggregation and membrane association, (2) specific structural information that will enable rational design strategies for small molecules that modulate alpha-synuclein function and/or serve as diagnostic tools for PD, and (3) elucidation of specific structural interactions between alpha-synuclein and other brain proteins. Relevance to public health: Alpha-synuclein is a central player in Parkinson's disease. The precise relationships between alpha-synuclein structure and this disease are not yet well understood. Our proposed studies aim to advance this fundamental knowledge, which will assist the broader research community in developing improved diagnostic tools and therapies for Parkinson's disease.

描述：该研究项目的长期目标是开发详细的原子分辨率 对与人类疾病相关的蛋白质聚集和纤维颤动的结构和功能的理解， 主要是对α-突触核蛋白的研究。野生型 α-突触核蛋白及其突变体（A30P、E46K 和 A53T）与早发性帕金森病（PD）相关，将在几种结构状态下进行检查， 包括原纤维、原纤维和膜相关复合物。我们建议化学细节 这些结构决定了 α-突触核蛋白的自然生理功能，并且在突变体或突变体中 环境压力，有助于PD病理。此外，我们建议几个之间的转换 结构状态是突触核蛋白的基本特征；这种结构可塑性可能需要 神经元的发育和维持。为了检查这些结构，魔角旋转（MAS）固体 将采用状态核磁共振（SSNMR）实验，阐明共价化学、构象的细节 和其他实验技术无法获得的动力学。补充分析和 表征技术将包括动力学测量（通过硫黄素 T 荧光、光散射 和沉降分析）、高分辨率质谱、原子力和电子显微镜， 溶液核磁共振和噬菌体展示。我们将利用这些技术以及样品制备和 同位素标记策略可提高我们对与疾病相关的蛋白质聚集的理解 α-突触核蛋白的具体案例，包括：（1）α-突触核蛋白的基本生物物理原理 聚集和膜关联，(2) 有助于合理设计的特定结构信息 调节 α-突触核蛋白功能和/或作为诊断工具的小分子策略 PD，以及（3）阐明 α-突触核蛋白和其他脑蛋白之间的特定结构相互作用。 与公共卫生的相关性：α-突触核蛋白是帕金森病的核心参与者。精确的 α-突触核蛋白结构与这种疾病之间的关系尚不清楚。我们提出的 研究旨在推进这一基础知识，这将有助于更广泛的研究界 开发改进的帕金森病诊断工具和疗法。