Structural Analysis of IAPP Fibril Formation and Membrane Interaction

项目概述：IAPP（胰岛淀粉样蛋白多肽）的错误折叠被认为在II型糖尿病中起重要作用，与其他与年龄相关的淀粉样蛋白疾病（包括阿尔茨海默病和帕金森病）中涉及的其他蛋白质类似。然而，这种错误折叠过程的结构细节很难获得。最近的工作，主要来自我的小组，证明了位点定向自旋标记（SDSL）是研究淀粉样蛋白结构的有力方法。在这项拟议的研究中，我们将使用SDSL提供IAPP错误折叠所涉及的定义构象状态的详细结构信息，并且我们将尝试确定小分子抑制剂如何阻止这些结构的形成。Specific Aim 1旨在生成IAPP淀粉样原纤维的三维模型。淀粉样蛋白原纤维是淀粉样蛋白疾病的病理标志，是一个逐步错误折叠过程的最终产物。如果不详细了解淀粉样蛋白的纤维结构，就不可能理解淀粉样蛋白错误折叠的分子机制。在具体目标2中，我们建议对？为了对我们之前的发现提供一个机制的理解，这种膜相互作用可以催化IAPP的错误折叠。在Specific Aim 3中，我们建议提供IAPP非纤维细胞毒性低聚物的详细结构信息。重要的是，这些定义明确的低聚物已经在体内被鉴定出来，并且被认为在淀粉样蛋白疾病（包括II型糖尿病）中发挥重要作用。在Specific Aim 4中，我们将利用我们的SDSL方法，结合Specific Aims 1-3的结构信息，研究小分子抑制剂如何与IAPP相互作用并防止错误折叠。相关性：从Specific Aims 1-4中获得的结构和机制信息将极大地促进治疗2型糖尿病和其他淀粉样蛋白疾病（包括阿尔茨海默病和帕金森病）的药物的开发。此外，我们的研究应该使设计有选择性地改变错误折叠的突变体成为可能。这些突变体将为研究错误折叠淀粉样蛋白的细胞靶点和体内毒性机制提供有力的工具。

DESCRIPTION (provided by applicant): Project Summary: The misfolding of IAPP (islet amyloid polypeptide) is thought to play an important role in type II diabetes and is analogous to that of other proteins involved in other age-related amyloid diseases, including Alzheimer and Parkinson disease. However, structural details of this misfolding process have been difficult to obtain. Recent work, largely from my group, demonstrates that site-directed spin labeling (SDSL) is a powerful approach for investigating the structures of amyloidogenic proteins. In this proposed study, we will use SDSL to provide detailed structural information on defined conformational states involved in IAPP misfolding, and we will try to determine how small molecule inhibitors can prevent those structures from forming. Specific Aim 1 is designed to generate a three-dimensional model of IAPP amyloid fibrils. Amyloid fibrils are the pathological hallmarks of amyloid diseases and represent the end product of a stepwise misfolding process. Understanding the molecular mechanism of amyloid protein misfolding will not be possible without detailed knowledge of the fibrillar structures. In Specific Aim 2, we propose to perform structural studies on ?-helical, membrane-bound IAPP in order to provide a mechanistic understanding of our previous finding that such membrane interactions can catalyze the misfolding of IAPP. In Specific Aim 3, we propose to provide detailed structural information on non-fibrillar, cytotoxic oligomers of IAPP. Importantly, these well-defined oligomers have been identified in vivo, and are thought to play an important role in amyloid diseases, including type II diabetes. In Specific Aim 4, we will utilize our SDSL approach, combined with the structural information from Specific Aims 1-3, to study how small molecule inhibitors interact with IAPP and prevent misfolding. Relevance: The structural and mechanistic information obtained from Specific Aims 1-4 should greatly facilitate the development of therapeutic agents for the treatment of type II diabetes and other amyloid diseases, including Alzheimer and Parkinson disease. In addition, our studies should make it possible to design mutants that selectively alter misfolding. Such mutants would provide powerful tools for studying the cellular targets and mechanisms of toxicity of misfolded amyloid proteins in vivo.