New assay method for pinpointing structural features in amyloid oligomer formation

摘要 该项目的目标是为生物医学研究提供一项重要的新技术， 尤其是对于研究与阿尔茨海默氏症进展相关的蛋白质的研究人员来说 疾病(AD)。开发治疗阿尔茨海默病的大多数努力，其中大多数已经 靶向Aβ纤维，失败了。越来越多的证据表明，低聚物形成 作为聚合过程中形成的中间体的β可能是 疾病的发展。然而，为了设计针对有毒低聚物物种的治疗药物 或其他改变的蛋白质形式，我们首先需要这些物种的结构信息。在这里，我们 提出一种新的结构方法来表征易于聚集的低聚中间体 系统，并测试与阿尔茨海默病相关的特定淀粉样多肽的新方法。这 工作将开发一种新的结构分析，它协同结合了局部和全球 基于结构质谱学的方法。羟基自由基溶剂可及性方法 揭示作为构象函数发生的局部残基特有的结构变化 变化或复杂的形成，而天然质谱学方法揭示了全球 低聚物大小和化学计量的结构信息。这些方法将应用于 家族性阿尔茨海默病中淀粉样多肽的研究 这些多肽与野生型不同。新的化验也将被用来表征 淀粉样多肽和p3多肽之间的相互作用，p3多肽的含量是 研究更充分的Aβ，纤化速度比Ab快，但可以减轻Ab的毒性。最后，我们将联系到 这些中间结构形式对毒性有影响。这些结果将具有巨大的治疗作用。 对于AD的价值，并将为未来难以研究的结构调查奠定基础 由聚集或错误折叠引起的一系列疾病中的中间形式 蛋白质。

Summary The goal of this project is to provide a significant new technology for biomedical research, particularly for researchers studying proteins associated with the progression of Alzheimer's Disease (AD). The majority of efforts to develop therapeutics for AD, most of which have targeted Aβ fibrils, have failed. A growing body of evidence now suggests that oligomeric forms of Aβ, which are intermediates that form during the aggregation process, may be the culprits in disease progression. However, in order to design therapeutics targeting toxic oligomeric species or other altered protein forms, we first require structural information on those species. Here, we propose a new structural method to characterize oligomeric intermediates in aggregation-prone systems, and to test the new methodology on specific amyloid peptides implicated in AD. This work will develop a new structural assay which synergistically combines local and global structural mass spectrometry-based methods. The hydroxyl radical solvent accessibility method reveals local residue-specific structural changes that occur as a function of conformational changes or complex formation, while the native mass spectrometry method reveals global structural information on oligomer size and stoichiometry. These methods will be applied to the study of the amyloid peptides in the familial forms of AD to determine how the oligomeric states of these peptides differ from the wildtype. The new assay will also be used to characterize the interaction between amyloid peptides and the p3 peptide, which is twice as abundant as the more well-studied Aβ, fibrilzes faster than Ab, but can mitigate Ab toxicity. Finally, we will relate these intermediate structural forms to toxicity. These results will be of immense therapeutic value for AD, and will lay the groundwork for future structural investigations of difficult-to-study intermediate forms in a wide range of diseases caused by the aggregation or misfolding of proteins.