Biophysical Characterization of Alpha-Synuclein

α-突触核蛋白的生物物理表征

• 批准号: 6781783
• 负责人: ELLEN W DOSS-PEPE
• 金额: $ 11.46万
• 依托单位: UNIV OF MED/DENT NJ-R W JOHNSON MED SCH
• 依托单位国家: 美国
• 财政年份: 2001
• 资助国家: 美国
• 起止时间: 2001-08-15 至 2006-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6781783
• 关键词: Lewy body; Parkinson' s disease; X ray crystallography; alpha synuclein; chemical aggregate; chemical models; chemical stability; chimeric proteins; conformation; electron microscopy; genetic screening; intermolecular interaction; lipids; model design /development; molecular chaperones; nuclear magnetic resonance spectroscopy; peptides; postdoctoral investigator; protein folding; protein sequence; protein structure; structural biology; tissue /cell culture; vesicle /vacuole; yeasts

The long-term research objective of this project is to define how variations in amino acid sequence and surrounding environment of a- synuclein, the major protein component of Lewy bodies in Parkinson Disease affect structure, stability, self-association and/or aggregation, and fibril formation. The long-term goal of the career development plan in the academic setting is to become a fundamental researcher and scientific leader in the field of protein structure, stability, folding and aggregation. In particular, I will concentrate my efforts on the determinants (surrounding environment and amino acid sequence) of folding and misfolding of proteins and the relation of misfolding, aggregation, and precipitation to the neurological disorders which are so prevalent in the aged population. The urgency for biophysical research in this difficult area in which there are more questions than answers is very strong. By the mastering of new techniques, the fine- tuning of others with which I have already had experience, course work, technique workshops, conference presentations, and collaborative interactions throughout the course of the award, I will become complete as a scientist who is competent to address many of the questions and uncertainties which are inherent and continue to evolve in this field. The focus and general approach is to generate peptide models and expression systems for the production and purification of wild-type and variants bearing specific mutations of a-synuclein and (3-synuclein, a protein highly homologous to a-synuclein. In the first aim, the conformations of a-synuclein and (3-synuclein will be compared. The long-term goal of this aim is to determine the factors involved in dictating the native conformations of both a- and (3-synuclein. In the second aim, the hypothesis that (3-synuclein will not aggregate in vin-o will be tested. The long-term goal of this aim is to define the determinants of aggregation (i.e. differences in amino acid sequence, modifications of the protein resulting from aging, environmental factors) and to obtain information on aggregate structure. The third aim will test the hypothesis that the presence of chemical or molecular chaperones, or lipid vesicles that induce structure in a-synuclein in solution will inhibit the formation of insoluble fibrillar structures. The long-term goal of this aim is to provide a mechanism for the formation of Lewy bodies.

这个项目的长期研究目标是确定a-突触核蛋白的氨基酸序列和周围环境的变化如何影响结构、稳定性、自结合和/或聚集以及纤维形成。a-突触核蛋白是帕金森病路易体的主要蛋白质成分。学术背景下的职业发展计划的长期目标是成为蛋白质结构、稳定性、折叠和聚集领域的基础研究人员和科学领先者。特别是，我将集中精力研究蛋白质折叠和错误折叠的决定因素(周围环境和氨基酸序列)，以及错误折叠、聚集和沉淀与老年人普遍存在的神经疾病的关系。在这个问题多于答案的困难领域，生物物理研究的紧迫性非常强烈。通过掌握新的技术，对我已经有经验的其他人进行微调，课程工作，技术研讨会，会议演讲，以及在整个奖项过程中的协作互动，我将成为一个完整的科学家，有能力解决该领域固有的并继续演变的许多问题和不确定性。重点和一般方法是建立多肽模型和表达系统，用于生产和纯化携带α-突触核蛋白和(3-突触核蛋白)的特定突变的野生型和变异体，3-突触核蛋白是一种与a-突触核蛋白高度同源的蛋白质。第一个目的是比较α-突触核蛋白和(3-突触核蛋白)的构象。这一目标的长期目标是确定决定α-和(3-突触核蛋白)天然构象的因素。在第二个目标中，将检验(3-突触核蛋白不会在Vin-O中聚集的假设)。这一目标的长期目标是确定聚集的决定因素(即氨基酸序列的差异、老化引起的蛋白质修饰、环境因素)，并获得关于聚集结构的信息。第三个目标将检验这样一种假设，即溶液中存在化学或分子伴侣，或诱导a-突触核蛋白结构的脂泡，将抑制不溶纤维结构的形成。这一目标的长期目标是为路易体的形成提供一个机制。