Amyloid ion channels to design therapeutics for neurodegenerative diseases

描述(申请人提供)：蛋白质构象疾病，包括神经退行性疾病(阿尔茨海默病和帕金森病)，II型糖尿病和囊性纤维化由蛋白质错误折叠引起，其3D构象从天然(可溶)改变为非天然(不可溶)折叠结构，称为淀粉样蛋白。理解这种错误折叠和导致病理生理细胞活动和退化的3D构象一直是最重要但也是最具挑战性的研究领域之一。最初的观点认为，错误折叠诱导的纤维结构通过自由基氧化应激和/或非特异性膜泄漏改变细胞膜组成，从而导致功能获得和病理生理学，从而破坏离子稳态。然而，最近的研究表明，球状、非纤维状的大分子复合体足以诱导细胞病理，推测是直接通过它们的膜孔来实现的。事实上，据报道，小分子单体和寡聚肽可以在人工细胞膜和天然细胞膜中诱导离子电导。这些可能的离子通道的三维结构和配体诱导的构象变化还知之甚少。了解类淀粉样蛋白通道的三维结构及其在细胞毒性中的作用，对于设计对抗药物和合理的构象疾病治疗策略是至关重要的。这项提议旨在检验关于蛋白质构象疾病的假设驱动的问题，并设计实现这些目标的技术。我们的总体假设是，蛋白质错误折叠疾病是由形成离子通道和分子的球状(而不是纤维状)构象造成的，其他干预措施可以用于有效的治疗，以调节它们的结构和活性。我们提出的具体目标如下：1：研究脂膜中重组的球状淀粉样蛋白多肽的三维离子通道结构；2：检测各种药物和多肽对通道开闭构象的响应；3：使用AFM-离子电导测量系统和药物相结合的方法将通道开闭构象与通道电导相关联。因此，我们的目标是提供一个独特和可行的模型系统，用于筛选可能的药物分子，并为由于离子通道结构和活性缺陷而导致的一系列退行性疾病设计治疗方案，即通道病。

DESCRIPTION (provided by applicant): Protein conformational diseases, including neurodegenerative (Alzheimer's and Parkinson's diseases), type II diabetes and cystic fibrosis result from protein misfolding that alters their 3D conformations from native (soluble) to non-native (insoluble) folded structures, called amyloids. Understanding such misfolding and the resulting 3D conformations that induce pathophysiological cellular activity and degeneration have been one of the most important and yet challenging areas of research. The original idea suggests that misfolding-induced fibrillar structure results into a gain-of-function and induce pathophysiology by altering cell membrane composition via free radical oxidative stress and/or non-specific membrane leakage and thence destabilizing ionic homeostasis. However, recent studies show that globular, non-fibrillar macromolecular complexes are sufficient to induce cell pathology, presumably directly by their membrane poration. Indeed, small monomeric and oligomeric peptides are reported to induce ionic conductances in artificial as well as native cell membranes. 3D structure and ligand-induced conformational changes of these putative ion channels are poorly understood. An understanding of the 3-D structure of amyloidogenic peptide-channel and its role in cell toxicity is essential for designing counteracting drugs and rational strategies for treatment of conformational diseases. This proposal aims to examine hypothesis-driven questions about protein conformational diseases and also design techniques to accomplish the goals. Our overall hypothesis is that protein misfolding diseases result from their globular (not fibrillar) conformation that forms ion channels and molecules and that other interventions, to modulate their structure and activity, could be used for effective therapy. We propose the following Specific Aims: 1: examine 3D ion-channel structure of globular amyloidogenic peptides reconstituted in lipid membrane; 2: Examine open-closed channel conformations in response to various pharmacological agents and peptides; 3: correlate channel open-close conformations with channel conductance using a combined AFM-ion conductance measurement system and pharmacological agents. Thus we aim to provide a unique and feasible model system for screening putative drug molecules and designing therapeutics for a wide range of degenerative diseases that result from defects in ion channel structure and activity, the "channelopathies".