A Single Molecule Approach to neurodegeneration in AD

治疗 AD 神经变性的单分子方法

• 批准号: 7142101
• 负责人: ARI GAFNI
• 金额: $ 18.76万
• 依托单位: UNIVERSITY OF MICHIGAN AT ANN ARBOR
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-07-01 至 2008-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7142101
• 关键词: Alzheimer' s disease; amyloid proteins; chemical kinetics; fluorescence resonance energy transfer; high performance liquid chromatography; liposomes; mass spectrometry; membrane activity; membrane biogenesis; membrane permeability; molecular pathology; nanotechnology; neural degeneration; phospholipids; protein biosynthesis; protein protein interaction; protein structure function; time resolved data

DESCRIPTION (provided by applicant): Understanding the origin of neurotoxicity that leads to neurodegeneration in Alzheimers Disease (AD) is critical to the development of an intervention. Recent studies point to early oligomers of the amyloid beta peptide, A-beta, as critical players in the etiology of AD. Applying traditional approaches to study these oligomers can be challenging because ensemble averaging masks low amounts of transient intermediates and hinders the resolution of species heterogeneity. We propose to apply recently developed approaches, based on single molecule spectroscopy (SMS), that are uniquely suited for these studies to develop the following specific aims: Aim 1: To apply SMS to follow the time evolution of A-beta oligomer formation in solution and to identify those transient oligomers that develop into ordered structures (such as pre- protofibrils) and/or insoluble fibrils. There is clear evidence that during the early stages of A-beta association a highly heterogeneous mix of oligomers develops complicating the identification of the pathological species. We will use SMS of fluorescently labeled A-beta peptides to follow the formation of oligomers as a function of time and to examine the basis for the apparent increased propensity for aggregation of A-beta1-42. The specific hypothesis to be tested is that soluble oligomers form by multiple reaction pathways and that A- beta1-42 more readily forms the initial nucleus for protofibril/fibril formation thus reducing the critical concentration for aggregation in the A-beta 1-42/1-40 mix. Aim 2: To determine the reaction sequence for A- beta oligomer/protofibril formation on the surface of membrane liposomes, to detect membrane permeabilization, to make initial characterization of the pores created and to identify oligomers that affect permeabilization. Recent observations suggest that part of A-beta's neurotoxicity may be associated with membrane binding of pre-fibril structures. We will initiate SMS experiments to test the hypothesis that membranes facilitate the formation of oligomers and to study key aspects of their formation. We will also determine whether a sub-class of the bound A-beta oligomers leads to membrane permeabilization and, if so, at which point this happens and is there a unique species or a multitude of permeabilizing oligomers. Long-term we expect to develop a better understanding of the biomolecular mechanisms and interactions that underlie the evolution of A-beta oligomers, and examine their role in fibril formation and neurotoxicity.

描述（由申请人提供）：了解导致阿尔茨海默病（AD）神经变性的神经毒性的起源对于干预措施的开发至关重要。最近的研究指出，β 淀粉样肽 A-β 的早期寡聚物是 AD 病因学的关键参与者。应用传统方法研究这些低聚物可能具有挑战性，因为整体平均掩盖了少量的瞬时中间体并阻碍了物种异质性的解决。我们建议应用最近开发的基于单分子光谱 (SMS) 的方法，这些方法特别适合这些研究，以实现以下具体目标： 目标 1：应用 SMS 跟踪溶液中 A-β 低聚物形成的时间演化，并识别那些发育成有序结构（例如前原纤维）和/或不溶性原纤维的瞬时低聚物。有明确的证据表明，在 A-β 结合的早期阶段，低聚物的高度异质混合物的发展使病理物种的鉴定变得复杂。我们将使用荧光标记的 A-β 肽的 SMS 来跟踪寡聚体的形成随时间的变化，并检查 A-β1-42 聚集倾向明显增加的基础。待测试的具体假设是可溶性低聚物通过多种反应途径形成，并且A-β1-42更容易形成原纤维/原纤维形成的初始核，从而降低A-β1-42/1-40混合物中聚集的临界浓度。目标 2：确定膜脂质体表面 A-β 寡聚物/原纤维形成的反应顺序，检测膜透化作用，对所产生的孔进行初步表征，并识别影响透化作用的寡聚物。最近的观察表明，A-β 的部分神经毒性可能与前原纤维结构的膜结合有关。我们将启动 SMS 实验来测试膜促进低聚物形成的假设，并研究其形成的关键方面。我们还将确定结合的 A-β 寡聚物的子类是否会导致膜透化，如果是，则在何时发生这种情况，以及是否存在独特的物种或大量的透化寡聚物。从长远来看，我们期望更好地了解 A-β 寡聚物进化的生物分子机制和相互作用，并检查它们在原纤维形成和神经毒性中的作用。