PHYSICAL BIOCHEMISTRY AND BIOLOGY OF AMYLOID B-PROTEIN

在阿尔茨海默病(AD)中，淀粉样β蛋白(Abeta)纤维的进行性沉积，即“淀粉样斑块， 发生在大脑中。然而，神经元功能障碍可能发生在斑块形成之前。结构-神经毒性 研究表明，包括原纤维在内的毒性组件越来越小， 核旁和ADDLS。在啮齿动物中，Aβ寡聚体可以抑制长时程增强(LTP)，这是一种 学习和记忆。在AD患者中，Abeta寡聚体的存在水平显著高于老年组 普通人。因此，生物物理学、细胞培养、动物和人体研究都支持 假设Abeta寡聚是AD发病机制中的一个关键事件。如果是这样的话，那么 治疗策略取决于对病理性蛋白质折叠机制的阐明(S)， 齐聚和高阶组装。我们实验室继续努力了解最早的 Abeta组装、Abeta单体折叠和齐聚的步骤揭示了关键的结构特征。 这些包括非结构Abeta单体的Val24-Lys28区域的转角形成和相互作用 在中心疏水簇(Leu17-Ala21)中，N-端和C-端。四个目标 构成本申请的目的是检验从这些观测中产生的机械假说。 目的1.确定Abeta基因Val24-Lys28区域转弯的形成机制。 A.确定疏水相互作用的作用。 B.确定静电相互作用的作用。 C.确定氨基酸转位倾向的作用。 目的2.确定分子内折叠和早期Aβ齐聚的机制。 A.确定中心疏水簇(CHC)-C末端相互作用的结构动力学。 B.确定CHC-N-末端相互作用的结构动力学。 C.确定不同的转角构象对Aβ单体结构和齐聚的影响。 目的3.利用O？&N酰基迁移化学实现一种新的准同步反应体系 Abeta42折叠和自组装的研究。 A.合成26-O-酰基-异阿贝塔42(26-AIAbeta42)并研究其多肽随时间的变化 引发O？&N酰基迁移后的二级和四元结构。 B.使用准弹性光散射光谱来确定动力学和热力学参数 Abeta42的自组装。 C.使用离子迁移率光谱-质谱仪监测Abeta自组装中的早期齐聚事件。 D.合成并研究钠保护的26-AIAbeta42的生物物理和生物学行为。 目的4.确定结构特征如何被证明在控制Abeta折叠和 寡聚作用影响多肽的神经毒性。

In Alzheimer's disease (AD), progressive deposition of amyloid beta-protein (Abeta) fibrils, "amyloid plaques," occurs in the brain. However, neuronal dysfunction may occur prior to plaque formation. Structure-neurotoxicity studies have revealed progressively smaller toxic assemblies, including protofibrils, paranuclei, and ADDLs. In rodents, Abeta oligomers can inhibit long-term potentiation (LTP), a model for learning and memory. In AD patients, Abeta oligomers are present in levels significantly greater than in agematched normal individuals. Biophysical, cell culture, animal, and human studies thus all support the hypothesis that oligpmerization of Abeta is a key event in AD pathogenesis. If so, then the development of therapeutic strategies depends on elucidation of the mechanism(s) of pathologic protein folding, oligomerization, and higher-order assembly. Continuing efforts in our laboratory to understand the earliest steps in Abeta assembly, Abeta monomer folding and oligomerization, have revealed key structural features. These include turn formation in the Val24-Lys28 region of the unstructured Abeta monomer and interactions among the central hydrophobic cluster (Leu17-Ala21), N-terminus, and C-terminus. The four aims comprising this application seek to test mechanistic hypotheses emanating from these observations. Aim 1. To determine the mechanisms of turn formation in the Val24-Lys28 region of Abeta. a. To determine the role of hydrophobic interactions. b. To determine the role of electrostatic interactions. c. To determine the role of amino-acid turn propensity. Aim 2. To determine the mechanisms of intramolecular folding and early Abeta oligomerization. a. To determine the structural dynamics of central hydrophobic cluster (CHC)-C-terminus interactions. b. To determine the structural dynamics of CHC-N-terminus interactions. c. To determine the effects of alternative turn conformations on Abeta monomer structure and oligomerization. Aim 3. To use O?>N acyl migration chemistry to implement a new, quasisynchronous system for studies of Abeta42 folding and self-assembly. a. To synthesize 26-O-acyl-isoAbeta42 (26-AIAbeta42) and study the time-dependent changes in peptide secondary and quaternary structure following initiation of O?>N acyl migration. b. To use quasielastic light scattering spectroscopy to determine kinetic and thermodynamic parameters of Abeta42 self-assembly. c. To use ion mobility spectroscopy-mass spectrometry to monitor early oligomerization events in Abeta self-assembly. d. To synthesize and study the biophysical and biological behavior of Na-protected 26-AIAbeta42. Aim 4. To determine how structural features shown to be critical in controlling Abeta folding and oligomerization affect peptide neurotoxicity.