Modeling synaptic vesicles: how does alpha-Synuclein inhibit fusion?

描述（由申请人提供）：我们的目标是发展对单体α-突触核蛋白（alphaS）抑制突触囊泡融合的机理理解。该研究将为帕金森病（PD）的新治疗策略奠定基础。越来越多的共识是单体α S是突触囊泡运输的中心调节组分。虽然由α S聚集成不溶性原纤维介导的路易体的形成通常与PD相关，但高水平的α S也显示出破坏正常囊泡运输并显著抑制神经递质释放而不形成α S聚集体。我们的方法将涉及突触囊泡膜的生物物理和机械性能的定量研究，我们将联合收割机粗粒度的分子动力学模拟与一个小组的互补生物物理实验。单体α S和突触囊泡膜之间的天然相互作用的精确理解将使我们能够评估蛋白质在与PD相关的囊泡运输缺陷中的作用。有限的生物制药数据已经产生了相互矛盾的观点，在没有原纤维形成的情况下，α S过度表达如何抑制囊泡运输和融合。在从酵母到啮齿动物的多个模型系统中，过量的α S已被证明会阻止质膜上适当的突触囊泡循环。一种观点认为，这种病理可能是由α S和其他突触或质膜蛋白（例如SNARES）之间的相互作用驱动的。我们提出了另一种观点，基于我们的实验室和其他人最近的工作。根据这一观点，α S可以直接改变膜内脂质的物理性质-即膜刚性和相。这是在与其他蛋白质没有特异性相互作用的情况下实现的。因此，我们推断α S可能具有控制突触囊泡融合的内在能力。这种假设是 基于我们的初步数据和来自生物物理实验的已发表结果，这些结果表明α S降低膜的刚性，可以改变膜曲率，并且可以抑制合成（否则无蛋白质）脂质囊泡的融合。我们提出的研究将计算建模与实验X射线散射和原子力显微镜紧密结合，以弥合理解alphaS如何为囊泡融合创造物理屏障的关键差距。拟议的研究途径将提供有关PD相关的贩运缺陷的关键信息。最终，我们的工作将导致更好地理解alphaS的正常和异常功能，并使社区能够开发利用蛋白质天然状态的新治疗策略（即，恢复正常的囊泡运输。

DESCRIPTION (provided by applicant): Our goal is to develop a mechanistic understanding of the inhibition of synaptic vesicle fusion by monomeric alpha-Synuclein (alphaS). This research will establish the foundation for new therapeutic strategies in the treatment of Parkinson's disease (PD). There is growing consensus that monomeric alphaS is a central regulatory component of synaptic vesicle trafficking. Although the formation of Lewy bodies, mediated by the aggregation of alphaS into insoluble fibrils, is commonly associated with PD, high levels of alphaS have also been shown to disrupt normal vesicle trafficking and markedly inhibit neurotransmitter release without the formation of alphaS aggregates. Our approach will involve quantitative studies of the biophysical and mechanical properties of synaptic vesicle membranes for which we will combine coarse-grained molecular dynamics simulations with a panel of complementary biophysical experiments. A precise understanding of the native interactions between monomeric alphaS and synaptic vesicle membranes will position us to evaluate the protein's role in vesicle trafficking defects as they relate to PD. Limited biophysica data have yielded conflicting views on how alphaS over-expression inhibits vesicle trafficking and fusion in the absence of fibril formation. In multiple model systems from yeast to rodents, an overabundance of alphaS has been shown to stall proper synaptic vesicle cycling at the plasma membrane. One view is that this pathology may be driven by interactions between alphaS and other synaptic or plasma membrane proteins (e.g. SNARES). We propose an alternate view based on recent work both from our labs and others. According to this view, alphaS can directly alter the physical properties of lipids within membranes - namely membrane rigidity and phase. This is achieved in the absence of specific interactions with other proteins. We therefore reason that alphaS might have an intrinsic capacity to control synaptic vesicle fusion. This hypothesis is motivated by our preliminary data and published result from biophysical experiments, which show that alphaS reduces a membrane's rigidity, can alter membrane curvature, and can inhibit fusion of synthetic (otherwise protein-free) lipid vesicles. Our proposed research intimately combines computational modeling with experimental x-ray scattering and atomic force microscopy to bridge a critical gap in understanding how alphaS creates physical barriers to vesicle fusion. The proposed research avenue will provide critical information about PD associated trafficking defects. Ultimately, our work will lead to better understanding of normal and abnormal functions of alphaS and position the community to develop new therapeutic strategies that exploit the native state of the protein (i.e., restoring proper vesicle trafficking.