PREPARATION AND CHARACTERIZATION OF AB-DERIVED ACTIVE LIGANDS

该计划项目旨在测试非纤维组织的假说 淀粉样β蛋白(AB)的聚集集合体，我们有 β-衍生活性配体(ADAL)负责触发 导致神经胶质细胞激活、神经元可塑性的过程 阿尔茨海默病的功能障碍、突触变性和细胞死亡 疾病。扩展这一假设，有人提出了特定的神经胶质 蛋白质调节竞争的Abeta组装过程-以增强 阿达尔队形。在这个计划项目中，我们将准备和 描述ADL的特征，定义促进其 队形，我们将确定由 神经元、神经胶质细胞、混合培养物和器官型脑片中的Adals。 这些研究将与组织化学研究相协调，以 建立ADALS与AD病理的相关性。的关注点 项目1是ADALS和ADLS的制备和表征 对它们形成的条件进行调查。我们会 确定可重复形成的最佳体外条件 ADALS，准备足够数量的结构，生化和 生物物理特性研究，以及对它们的研究 与项目2和项目3合作的生物反应。 将研究腺体形成的机制，包括 强调选定的胶质辅助因子蛋白所起的作用 与AD的Abeta相关。我们将扩大我们的研究范围，包括 原子力显微镜(AFM)提供更详细的分析 腺体的形态和分布，我们将使用诸如 腺苷的场流动分离和天然凝胶电泳法 净化。我们将使用激光光散射，分析 超速离心和荧光偏振与能量传递 研究ADAL组装机制的技术。少校 这个项目要解决的问题是：什么是试验性的 条件有利于副词的形成，以及什么技术能够 腺体结构均一制剂的分离？ 什么因素支配着不同的腺体结构的形成，以及 不活跃的纤维蛋白或可溶性Abeta组件？什么是 阿达斯的生化和生物物理特性？具体是什么 Abeta1-42的一级结构特征对 组装成不同的腺体和纤维结构？什么氨基 酸性残基对Adals的生物活性很重要 神经元？在神经胶质里？将这些研究与广泛的 该计划项目中的生物能力将定义 导致炎症的分子和细胞过程 病理、突触变性和神经细胞死亡 阿尔茨海默氏症。

This Program Project seeks to test the hypothese that non-fibrillar aggregated assemblies of amyloid beta (AB), which we have term Abeta-derived active ligands (ADALs), are responsible for triggering processes leading to the glial activation, neuronal plasticity malfunction, synaptic degeneration and cell death in Alzheimer's disease. Extending this hypothesis, it is proposed that particular glial proteins modulate competing Abeta assembly process-to enhance ADAL formation. In this program project we will prepare and characterize ADALs, definign conditions that promote their formation, and we will identify the specific responses activated by ADALs in neurons, glia, mixed cultures and organotypic brain slices. These studies will be coordinated with histochemical studies to establish the relevance of ADALs to AD pathology. The focus of project 1 is the preparation and characterization of ADALs and investigation of conditions reponsible for their formation. We will define optimal in vitro conditions for reproducible formation of ADALs,preparing sufficient quantities for structural, biochemical and biophysical characterization studies, and for investigations of their biological responses in collaboration with Projects 2 and 3. The mechanisms governing ADAL formation will be investigated, with an emphasis on the role played by selected glial co-factor proteins associated wit Abeta in AD. We will extend our studies involving atomic force microscopy (AFM) to provide more detailed analysis of ADAL morphology and distribution, and we will use techniques such as field flow fractionation and native gel electrophoresis for ADAL purification. We will employ laser light scattering, analytical ultracentrifugation and fluroescence polarization and energy transfer techniques to investigate ADAL assembly mechanisms. The major questions to be addressed in this project are: What experimental conditions favor the formation of ADALs and what techniques enable the isolation of homogeneous preparation of ADAL structures? What factors govern the formation of different ADAL structures, and inactive fibrillar or soluble Abeta assemblies? What are the biochemical and biophysical characteristics of ADALs? What specific features of the Abeta 1-42 primary structure are important for assembly into different ADAL and fibrillar structures? What amino acid residues are important for the biological activity of ADALs in neurons? in glia? The integration of these studies with the extensive biological capabilities within this Program Project will define the molecular and cellular processes that lead to the inflammatory pathology, synaptic degeneration and neuronal cell death that occurs in Alzheimer's disease.