NMR Based Studies of Alpha-Synuclein Aggregation and Inhibition

 描述（由申请人提供）：许多实验室已经研究了神经退行性疾病中的聚集机制，尽管该过程仍然难以捉摸，但在定义该过程的不同阶段方面已经取得了重大进展。为了对抗这些使人衰弱的疾病，不仅要描述聚集过程，而且要了解聚集抑制的机制。N-末端乙酰化α-突触核蛋白（Ac-aS）是一种小的神经元内在无序蛋白（IDP），其在病理性神经元损伤中自缔合。 帕金森病患者大脑中的低聚物和纤维的方法。相比之下，与Ac-aS共定位的高度同源的IDP蛋白，乙酰化β-突触核蛋白（Ac-β S），不自缔合成原纤维，但可以在体内充当Ac-aS毒性的神经保护剂以抑制病理性Ac-aS原纤维化。鉴于神经退行性疾病令人难以置信的重要性 在老龄化人口中，疾病及其对全球健康的威胁，详细了解抑制的分子机制至关重要。在这个阶段，对这些过程以及指导抑制与聚集的特定分子间相互作用知之甚少。这项建议的目标是利用核磁共振和其他生物物理技术，结合细胞毒性研究，提供第一个分子描述的复合物 存在于不同的抑制阶段。在此补助金中，我们将：（1）表征不同的aS/β S序列或结构单元对天然抑制剂β S及其毒性突变体加速/抑制原纤维形成的潜力的影响;（2）绘制确定抑制的最早阶段的相互作用区域;和（3）进行aS、β S和杂aS/β S寡聚体的分子表征和毒性研究。拟议工作的结果将是推进我们对aS/ßS抑制的“结构-毒性”关系的基本理解，并解决有关抑制相互作用的位置、性质和特异性的更多全球性问题。这些研究将提供一个框架，以确定特定的相互作用，这是重要的破坏有毒的寡聚体，并提出新的目标，帕金森氏病的治疗干预。本文开发的方法可以更广泛地应用于其他交叉淀粉样蛋白相互作用，这些相互作用已被证明在神经退行性疾病中起关键作用，例如aS和淀粉样蛋白-β-蛋白或aS和tau之间的相互作用。

 DESCRIPTION (provided by applicant): The mechanism of aggregation in neurodegenerative disease has been investigated by numerous laboratories and although the process is still elusive, significant progress has been made in defining different stages of the process. To fight these debilitating illnesses it is not only critical to characterize aggregation processes but alsoto understand the mechanisms of aggregation inhibition. N-terminally acetylated a-synuclein (Ac-aS) is a small neuronal intrinsically disordered protein (IDP) that self-associates in pathological ways to oligomers and fibrils in the brains of patients with Parkinson's disease. In contrast, a highly homologous IDP protein, acetylated ß- synuclein (Ac-ßS) that co-localizes with Ac-aS, does not self-associate into fibrils but can act as a neuro-protector of Ac-aS toxicity in vivo to inhibit pathological Ac-aS fibrillation. In light of the incredible importance of neurodegenerative diseases and their threat to global health in the ageing population, a detailed understanding of the molecular mechanism of inhibition is critical. At this stage very little known about these processes and about the specific inter-molecular interactions that guide inhibition versus aggregation. The goal of this proposal is to use NMR and other biophysical techniques, in conjunction with cell toxicity studies, to provide the first molecular description of the complexes that exist at the different stages of inhibition. In this grant we will: (1) characterize the influnce of different aS/ßS sequences, or structural units, on the potential to accelerate/inhibit fibril formation by the natural inhibitor ßS and its toxic mutants; (2) map interactive regions that define the earliest stages of inhibition; and (3) undertake the molecular characterization and toxicity studies of aS, ßS and hetero aS/ßS oligomers. The outcome of the proposed work will be to advance our fundamental understanding of the 'structure-toxicity' relationship of aS/ßS inhibition and to address more global questions about the location, nature and specificity of inhibitory interactions. These studies will provide a framework for identifying specific interactios that are important for disrupting toxic oligomers and for proposing novel targets for therapeutic intervention in Parkinson's disease. The approaches developed here can be more widely applied to other cross amyloid interactions that have been shown to play a critical role in neurodegenerative disease, such as the interaction between aS and amyloid-ß-protein, or aS and tau.