NMR Based Studies of Alpha-Synuclein Aggregation and Inhibition

 描述(申请人提供)：神经退行性疾病的聚集机制已经被许多实验室研究过，尽管这一过程仍然难以捉摸，但在确定这一过程的不同阶段方面已经取得了重大进展。为了对抗这些令人衰弱的疾病，不仅要确定聚集过程的特征，而且要了解聚集抑制的机制。N-末端乙酰化的α-突触核蛋白(ac-as)是一种小的神经元固有无序蛋白(Idp)，在病理过程中自相关。 帕金森氏病患者脑内寡聚体和纤维形成的途径。相比之下，与Ac-AS共定位的高度同源的IdP蛋白-乙酰化-突触核蛋白(Ac-？S)不会自我结合成纤维，但可以在体内作为Ac-As毒性的神经保护器来抑制病理性Ac-As纤颤。鉴于神经退行性变令人难以置信的重要性 在老龄化人口中，对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，人们对这些过程以及指导抑制和聚集的特定分子间相互作用知之甚少。这项提议的目标是使用核磁共振和其他生物物理技术，结合细胞毒性研究，提供第一个对络合物的分子描述。 存在于抑制的不同阶段。在这项资助中，我们将：(1)表征不同的As/？S序列或结构单元对天然抑制剂？S及其有毒突变体加速/抑制纤维形成的潜力的影响；(2)绘制定义抑制的最早阶段的相互作用区域图；以及(3)进行As、？S和杂As/？S低聚物的分子表征和毒性研究。拟议工作的结果将促进我们对AS/？S抑制物的‘结构-毒性’关系的基本理解，并解决关于抑制物相互作用的位置、性质和特异性的更多全球性问题。这些研究将为确定特定的相互作用提供一个框架，这些相互作用对于破坏有毒低聚物很重要，并为帕金森氏病的治疗干预提出新的靶点。这里开发的方法可以更广泛地应用于其他已被证明在神经退行性疾病中发挥关键作用的交叉淀粉样蛋白相互作用，例如AS与淀粉样蛋白之间的相互作用，或AS与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.