Solid-state NMR Structural Characterizations of Polymorphic Transthyretin Amyloids

蛋白质错误折叠和淀粉样蛋白形成与许多疾病有关，如淀粉样变性， 普里恩和阿尔茨海默氏症。Prion病的独特之处在于，天然折叠的Prion蛋白形成 具有不同分子构象的聚集体(Pron菌株)，它们是不同疾病的基础 表型1-3病毒株可能编码在蛋白质的初级序列和突变中 诱导不同的菌株，导致不同的疾病表型。最近的研究表明， 菌株假说适用于其他同样表现为多种疾病的淀粉样变性疾病。 表型：1，2，4，5非蛋白淀粉样蛋白表现出广泛的构象多样性，6-10 可能与表型变异有关。然而，人们对其分子基础知之甚少。 不同的错误折叠途径和淀粉样蛋白的结构多样性。淀粉样蛋白的结构研究如下 对了解淀粉样蛋白多样性的分子机制至关重要。病原菌的作用 还应检查错误折叠途径上的突变。系统的生物物理研究已经， 然而，由于限制淀粉样蛋白的形成，对先前研究的淀粉样蛋白来说是具有挑战性的 与不同疾病表型相关的致病突变数量。此外，最多的 广泛研究了与阿尔茨海默氏症和老年痴呆症相关的多肽、β-淀粉样蛋白和α-突触核蛋白 帕金森氏症分别是自然展开的，使得多肽不适用于 初始构象转变(错折)的机理研究。这个研究项目的目的是 在研究天然折叠蛋白转甲状腺蛋白(TTR)的淀粉样蛋白形成机制时，使用 固态核磁共振。转甲状腺素(Ttr)是人类30多种蛋白质中的一种。 构象变化和错误组装成β结构的淀粉样蛋白。野生型淀粉样蛋白的形成和 已知有100多种TTR突变形式可导致各种淀粉样变性，具有巨大的 表型多样性。11这一建议的主要假设是Ttr的致病突变形式 可能有不同的错误折叠途径，采用不同的淀粉样蛋白构象，具有不同的毒性 活动，这可能导致不同的疾病表型和组织选择性沉积。这个 假说将通过野生型和野生型淀粉样蛋白的结构特征进行验证 各种致病突变形式的TTR.特别是，具有创新标记的固态核磁共振 这些方案将为研究淀粉样蛋白的多样性提供有价值的见解。该提案的具体目标是 探索：(1)类淀粉样核心区结构特征。(2)分子的构象变化 淀粉样蛋白形成过程中的环状区域。(3)WT的四级结构和TTR的突变形式 淀粉样蛋白。错误折叠和淀粉样蛋白形成途径的机械理解将是 对于开发有效的TTR淀粉样变性治疗策略至关重要。

Protein misfolding and amyloid formation is implicated in numerous diseases such as amyloidoses, prion and Alzheimer's diseases. Prion disease is unique in that the natively folded prion protein forms aggregates with distinct molecular conformations (prion strains), which underlie different disease phenotypes.1-3 The prion strain may be encoded in the primary sequence and mutations of the protein induce different strains, causing distinct disease phenotypes. Recent studies have suggested the strain hypothesis is applicable to other amyloid diseases that also manifest diverse disease phenotypes.1,2,4,5 Nonprion amyloids were shown to exhibit a wide conformational diversity,6-10 which may be linked to the phenotype variations. However, little is known about molecular basis of the diverse misfolding pathways and structural diversity of amyloid. Structural studies of the amyloid are essential to understanding molecular mechanism of amyloid diversity. Effect of the pathogenic mutations on misfolding pathway should also be examined. The systematic biophysical studies have, however, been challenging for previously investigated amyloidogenic proteins due to the limited number of pathogenic mutations associated with distinct disease phenotypes. In addition, the most extensively studied polypeptides, β-amyloid and α-synuclein associated with Alzheimer's and Parkinson's diseases respectively, are natively unfolded, rendering the polypeptides not amenable for mechanistic studies of the initial conformational transition (misfolding). This research program is aimed at investigating amyloid formation mechanisms of a natively folded protein, transthyretin (TTR), using solid-state NMR. Transthyretin (TTR) is one of more than 30 human proteins that undergo an aberrant conformational change and misassemble into β-structured amyloid. Amyloid formation of wild type and more than 100 mutant forms of TTR are known to cause various amyloidoses with enormous phenotype diversity.11 The main hypothesis of this proposal is that pathogenic mutant forms of TTR may have distinct misfolding pathways, adopting diverse amyloid conformations with different toxic activities, which may result in diverse disease phenotypes and tissue-selective depositions. The hypothesis will be tested through the structural characterization of amyloid derived from wild-type and various pathogenic mutant forms of TTR. In particular, solid-state NMR with innovative labeling schemes will provide valuable insights into amyloid diversity. Specific aims of the proposal are to explore: (1) Native-like structural features of amyloid core regions. (2) Conformational changes of the loop regions during amyloid formation. (3) Quaternary structure of WT and mutant forms of TTR amyloid. Mechanistic understanding of the misfolding and amyloid formation pathways would be critical to developing effective therapeutic strategies for TTR amyloidoses.