Physical Biochemistry and Biology of Amyloid Beta-Protein

描述(申请人提供)：阿尔茨海默病(AD)是一种毁灭性的老年人疾病。两种淀粉样蛋白与AD有关，即淀粉样蛋白(A？)还有陶。最近的证据支持一种假说，即“淀粉样级联假说”，该假说认为A？寡聚体是阿尔茨海默病的主要神经致病因子。这项建议的总体目标是了解Aβ及其片段的结构生物学，并建立正式的结构-活性关系。从长远来看，我们寻求获得由A？形成的最接近的神经毒素结构的原子分辨率测定，并通过这样做，第一次使治疗疾病的AD疗法的发展成为可能。将采用多学科战略，采用互补的实验和计算方法。这一策略在过去被非常成功地使用，为A？系统。提出了三个具体目标，以系统和逻辑地在体外生物物理研究中取得进展。在体外和体内研究选定的此类组件的生物活性(目标2)，以确定选定的组件对神经元差异基因表达的影响(目标3)。综上所述，这些研究将为后续的治疗化合物开发和临床试验提供理论和实验基础。目的1.测定A？和tau组件。A.用扫描酪氨酸诱变技术阐明A？齐聚作用。B.确定分子内转角形成的动力学及其对A？集合。C.确定一级结构变化对生物相关和理论上重要的A？多肽。目的2.测定A？装配。A.确定A？培养神经元细胞系和原代神经元上的组装体。B.确定A的效果？果蝇眼睛发育、运动和寿命的组件。目的3.利用生物信息学方法识别和验证AD相关基因。组装结构、神经元类型和神经元衰老。

DESCRIPTION (provided by applicant): Alzheimer's disease (AD) is a devastating disease of the aged. Two amyloid-forming proteins are associated with AD, the amyloid ?-protein (A?) and tau. Recent evidence supports an hypothesis, the "amyloid cascade hypothesis," that posits that A? oligomers are the seminal neuropathogenetic agents in AD. The overall goal of this proposal is to understand the structural biology of A?, and fragments thereof, and to establish formal structure-activity relationships. In the long run, we seek to obtain an atomic-resolution determination of the structure of the proximate neurotoxins formed by A?, and in doing so, enable the development, for the first time, of disease-modifying AD treatments. A multidisciplinary strategy, employing complementary experimental and computational approaches, will be employed. This strategy has been used very successfully in the past, providing novel insights into the A? system. Three specific aims are proposed that systematically and logically progress from in vitro biophysical studies of A? and its oligomeric assemblies (Aim 1), to in vitro and in vivo studies of the biological activity of selected such assemblies (Aim 2), to determination of the effects of selected assemblies on differential gene expression in neurons (Aim 3). Taken together, these studies will provide the theoretical and experimental foundation for subsequent therapeutic compound development and clinical testing in humans. Aim 1. To determine the structural dynamics of A? and tau assembly. a. To use scanning Tyr mutagenesis to elucidate mechanisms of A? oligomerization. b. To determine the dynamics of intramolecular turn formation and its effects on A? assembly. c. To determine the effects of primary structure changes on the conformations and assembly dynamics of biologically relevant and theoretically important A? peptides. Aim 2. To determine the biological effects of A? assemblies. a. To determine the cytotoxic effects of A? assemblies on cultured neuronal cell lines and primary neurons. b. To determine the effects of A? assemblies on Drosophila eye development, locomotion, and longevity. Aim 3. To identify and validate AD-relevant genes using a bioinformatics approach that considers A? assembly structure, neuron type, and neuron senescence.