Structure/function analysis of A-beta fibril assembly

描述（由申请人提供）：多种实验方法表明，A β肽在皮质中的有序聚集是阿尔茨海默病特征性神经元功能进行性丧失的关键步骤。目前的想法有利于快速形成的聚集体，称为原纤维，在残基41至43结束的A β肽的毒性作用。然而，也有可能由这些肽组成的成熟淀粉样蛋白原纤维，以及终止于39或40位的较短版本，也可能影响疾病，作为毒性或保护性实体。虽然我们对Abeta淀粉样蛋白结构的了解目前只有适度的分辨率，但在过去四年中，在Abeta聚集体的结构和结构能量学分析方面取得了很大进展，并开发了用于聚集体结构分析的新工具。该建议建立在以前的工作，重点是（a）的结构和稳定的A β（1-42）淀粉样蛋白;（B）的结构和稳定的A β（1-42）原纤维;和（c）的结构和性质的不同构象类型的A β淀粉样蛋白原纤维最近报道。将丙氨酸和脯氨酸突变体聚集，并测定和分析由这些突变体形成的淀粉样蛋白原纤维和原纤维的稳定性变化。将对1-42个突变体进行单和双半胱氨酸突变体交联分析，以确定原纤维的内部结构。这些和其他方法也将被应用于原纤维的替代构象的结构和稳定性的分析（Abeta原纤维的改变的构象形式可能影响其生物活性，如酵母朊病毒的淀粉样蛋白形式的情况）。涉及与几个分子建模组的合作，该提案将使用实验数据来测试原纤维和原纤维结构和组装的当前模型，并将允许设计额外的实验来测试结构模型。这些研究应有助于了解Abeta淀粉样蛋白原纤维的结构，装配，和淀粉样蛋白原纤维结构稳定的基本基础，这将是重要的开发结构模型的淀粉样蛋白原纤维的Abeta和其他肽负责神经退行性疾病。此外，这些研究应该继续推动我们对Abeta原纤维及其在疾病中的作用的认识的前沿。这些数据将增强我们对AD分子基础的了解，并在此过程中有助于药物设计分子靶点的开发。

DESCRIPTION (provided by applicant): A variety of experimental approaches implicate the ordered aggregation of the Abeta peptide in the cortex as a key step in the progressive loss of neuronal function characteristic of Alzheimer's disease. Current thinking favors a toxic role for rapidly formed aggregates, called protofibrils, of Abeta peptides ending at residue 41 to 43. However, it is also possible that mature amyloid fibrils composed of these peptides, as well as shorter versions ending at positions 39 or 40, may also influence disease, as either toxic or protective entities. Although our knowledge of Abeta amyloid structure is at present only at modest resolution, great strides have been made in the last four years in the analysis of the structure and structural energetics of Abeta aggregates, and new tools for structural analysis of aggregates have been developed. This proposal builds on previous work by focusing on (a) the structure and stabilization of Abeta (1-42) amyloid; (b) the structure and stabilization of Abeta (1-42) protofibrils; and (c) the structure and properties of different conformational types of Abeta amyloid fibrils recently reported. Alanine and proline mutants will be aggregated, and the change in stability of amyloid fibrils and protofibrils formed from these mutants will be determined and analyzed. Single and double cysteine mutant crosslinking analysis will be conducted on 1-42 mutants, to determine the internal structure of the fibrils. These and other methods will also be applied to the analysis of the structure and stability of alternative conformations of the fibrils (altered conformational forms of Abeta fibrils may have influenced their biological activity, as is the case for amyloid forms of yeast prions). Involving collaborations with several molecular modeling groups, this proposal will use experimental data to test current models for fibril and protofibril structure and assembly, and will allow for design of additional experiments to test structural models as they are developed. These studies should contribute to the understanding of Abeta amyloid fibril structure, assembly, and the fundamental basis of amyloid fibril structural stabilization, which will be important in developing structural models for amyloid fibrils of Abeta and other peptides responsible for neurodegenerative diseases. In addition, these studies should continue to push the frontiers of our knowledge of Abeta protofibrils and their role in disease. This data will enhance our knowledge of the molecular basis of AD and in the process contribute to the development of molecular targets for drug design.