The structural basis of apoE4's role in Alzheimer's Disease

描述（由申请人提供）：载脂蛋白E（apoE）是一种299个氨基酸（~34 kD）的蛋白质，在脂质转运和代谢中起核心作用。与apoE 3和apoE 2不同，apoE 4是阿尔茨海默病（AD）的既定危险因素。然而，这些异构体特异性作用的分子基础在很大程度上是未知的，最重要的是，尚未在结构和功能方面进行系统的探索。大多数注意力都集中在apoE对大脑中A肽加工的影响上。 具体目标。 目的1：鉴定apoE 4中β结构的起始。 目的2：表征apoE亚型与A肽的相互作用。 这些目标将阐明实现apoE 4如何参与AD和神经退行性变的一般机制的理解所需的最重要的细节。在我们（包括合作者）对功能系统的专业知识的指导下，我们有经验和技术，通过应用最有可能揭示AD中apoE亚型效应基础的结构生物学工具来独特地解决这个问题。这些工具包括荧光光谱，电子显微镜，表面等离子体共振和FTIR光谱，虽然我们的主要方法将利用电子顺磁（EPR）光谱的定点自旋标记。 意义由于EPR能够报告溶液中样品的局部结构和空间关系，这项工作可能会转化为候选药物筛选的有效工具。可能的方法包括使用自旋标记的侧链来评估β链阻断剂，这些阻断剂旨在靶向一个已识别的结构域或稳定apoE内不稳定区域的带电化学分子伴侣。由于apoE 3也可能经历不稳定的构象，尽管频率低得多，这种治疗也可能有助于减缓E3携带者的AD进展。

DESCRIPTION (provided by applicant): Apolipoprotein E (apoE) is a 299 amino acid (~34 kD) protein that plays a central role in lipid transport and metabolism. Unlike apoE3 and apoE2, apoE4 is an established risk factor for Alzheimer's disease (AD). However, the molecular basis of these isoform-specific effects is largely unknown and, most importantly, has not been explored systematically in terms of structure and function. Most attention has been focused on the influence of apoE on A¿ peptide processing in the brain. Specific Aims. Aim 1: Identify the initiation of beta structure in apoE4. Aim 2: Characterize the interaction of apoE isoforms with the A¿ peptide. These aims will illuminate the most important details needed for achieving a mechanistic understanding of how apoE4 participates in AD and neurodegeneration in general. Guided by our (including collaborators) expertise on the functional system, we have the experience and technology to uniquely contribute to this problem by applying the structural biology tools most likely to uncover the basis for the apoE isoform effect in AD. These tools include fluorescence spectroscopy, electron microscopy, surface plasmon resonance, and FTIR spectroscopy, though our primary method will utilize electron paramagnetic (EPR) spectroscopy of site-directed spin labels. Significance. Because of the ability of EPR to report on local structure and spatial relationships from the sample in solution, this work may translate into an effective tool for drug candidate screening. Possibilities include use of spin-labeled side chains to evaluate beta-strand blockers designed to target an identified domain or charged chemical chaperones that stabilize a labile region within apoE. Since apoE3 may also experience destabilized conformations, though at a much lower frequency, such treatments may be helpful in slowing the progression of AD in E3 carriers as well.