Structure and Function of Alpha-Synuclein

描述(由申请人提供)：蛋白α-突触核蛋白(AS)与家族性和散发性帕金森病(PD)的病因有关。AS的正常功能与其病理性聚集之间的相互作用尚不清楚，但膜结合形式的AS被认为介导了其生理功能，而聚集形式被认为介导了蛋白质的毒性。在结构上，AS具有高度的延展性，在溶液中自由时采用高度无序的构象系综，当结合到磷脂膜上时采用高度螺旋结构，当聚集成淀粉样纤维时采用丰富的b-折叠构象。防止AS聚集成淀粉样蛋白纤维或潜在的有毒低聚物是治疗帕金森病的一种有前途的策略。这项研究的主要目标是详细了解突触核蛋白的结构性质和转变如何调节突触核蛋白的功能和毒性，并确定AS的特定构象状态，以促进具有潜在治疗价值的突触核蛋白相互作用试剂的设计。目前的提议旨在填补我们在理解AS结构方面出现的新的空白，这些空白是由a)发现一个新的PD连接的AS突变E46K；b)发现膜结合的AS可以采用两种不同的拓扑结构，即延伸的螺旋和断裂的螺旋，并提出这两种构象如何影响突触核蛋白功能的假说；c)发现新的AS相互作用伙伴被认为调节突触核蛋白的功能。为了填补这些空白并解决新出现的假说，我们制定了以下具体目标：1.为了确定最近发现的PD连锁突变E46K对AS游离型和膜结合型结构的影响。2.在高分辨率下阐明膜结合型AS的延伸螺旋结构。3.验证AS可以利用其先前已阐明的断裂螺旋结构来调节不同组成的不同拓扑结构的膜之间的相互作用。4.研究AARP16/19结合对膜结合AS结构的影响。这些目标的动机是有机会澄清AS序列变化如何影响蛋白质的结构和聚集，以及我们相信AS的单体膜结合构象更有序度，可能更适合与潜在的治疗药物形成特定的相互作用。这项工作将促进我们对AS的结构、功能和聚集的理解，并将为未来设计和鉴定能够稳定单体AS并防止其齐聚和聚集的试剂提供结构基础。此外，所获得的结果可能对解决其他与年龄相关的运动障碍和痴呆的蛋白质聚集的策略具有普遍意义。

DESCRIPTION (provided by applicant): The protein alpha-synuclein (aS) is implicated in the etiology of both familial and sporadic Parkinson's disease (PD). The interplay between the normal function of aS and its pathological aggregation is poorly understood, but membrane-bound forms of aS are thought to mediate its physiological function, while aggregated forms are thought to mediate the toxicity of the protein. Structurally, aS is highly malleable, adopting a highly disordered conformational ensemble when free in solution, highly helical structures when bound to phospholipids membranes and b-sheet rich conformations when aggregated into amyloid fibrils. Preventing the aggregation of aS into amyloid fibrils or potentially toxic oligomeric species is a promising strategy for the treatment of PD. The overarching goal of this research is to achieve a detailed understanding of how synuclein structural properties and transitions modulate synuclein function and toxicity and to identify specific conformational states of aS that could facilitate the design of synuclein-interacting reagents with potential therapeutic value. The current proposal is aimed at filling a newly emerged gaps in our understanding of aS structure that were created by a) the discovery of a new PD-linked aS mutation, E46K; b) the discovery that membrane-bound aS can adopt two different topologies, an extended helix and a broken helix, and the formulation of a hypothesis regarding how these two conformations may influence synuclein function; c) the discovery of new aS interaction partners thought to modulate synuclein function. To fill these gaps and to address emerging hypotheses we have developed the following specific aims: 1. To determine the effects of the most recently discovered PD-linked mutation, E46K, on structure in the free and membrane-bound forms of aS. 2. To elucidate at high resolution the extended-helix structure of membrane-bound aS. 3. To test the hypothesis that aS can mediate interactions between topologically distinct membranes of different compositions using its previously elucidated broken-helix structure. 4. To determine the effects of AARP16/19 binding on the structure of membrane-associated aS. These aims are motivated by the opportunity to clarify how aS sequence variations influence the structure and aggregation of the protein, and by our belief that monomeric membrane-bound conformations of aS, which are more highly ordered, may be better suited to form specific interactions with potential therapeutics. This work will advance our understanding of aS structure, function, and aggregation and will provide a structural basis for the future design and identification of reagents that can stabilize monomeric aS and prevent its oligomerization and aggregation. Furthermore, the results obtained may have general implications for strategies to address protein aggregation in other age-related motor disorders and dementias.