Inhibiting protein-protein interactions in the early stages of amyloid formation

在这个项目中，我们专注于胰岛相关多肽（IAPP）（II型糖尿病）和A-β（阿尔茨海默氏症）的聚集。越来越多的证据表明，这些蛋白质的内在无序的单体与聚集倾向的低聚物处于动态平衡。尽管IAPP/A-β聚集的表征取得了重大进展，其折叠/聚集机制的机械理解仍然模糊，需要新的基础知识来告知therapy.Objectives的设计：结合天然MS与生化/生物物理测定，我们将：1。探索小分子抑制剂如何影响IAPP/A-β聚集：使用MS，IMS和动力学测定（ThT荧光），我们将使用SER实验室最近发现的“命中”筛选新的小分子聚集抑制剂（Nature Chem（2015）和Nature Chem Biol（2016））。使用ETD的MS图谱（FS：Anal Chem（2016））结合其他基于溶液的测定，我们将阐明哪些物质负责与不同抑制剂的相互作用，并表征抑制剂结合界面。然后我们将使用细胞生物学测定来确定抑制剂是否影响细胞毒性。探索聚集的序列依赖性：使用SER实验室开发的新的聚集筛选（Nature Chem Biol（2016）），我们将选择具有改变的聚集特性的IAPP/A-β序列。然后，我们将选择增强/延缓疾病进展的突变，从而阐明这些突变如何干扰折叠和聚集。新奇/及时：据我们所知，该项目将揭示折叠/聚合的能量景观的第一个原子论见解。通过关注与II型糖尿病/阿尔茨海默氏症相关的淀粉样蛋白的病原体肽，结果也将对当今人类健康产生影响。实验方法：我们将充分利用天然MS，并将其与化学生物学，生物物理学和细胞生物学相结合，为学生提供广泛的技术培训。所有方法都已启动并运行，使学生能够快速进步。

In this project we focus on aggregation of Islet Associated PolyPeptide (IAPP) (type II diabetes) and A-beta (Alzheimer's). Increasing evidence suggests that the intrinsically disordered monomers of these proteins are in dynamic equilibrium with aggregation-prone oligomers. Despite significant progress in the characterization of IAPP/A-beta aggregation, mechanistic understanding of their folding/aggregation mechanisms remains obscure and new fundamental knowledge is needed to inform the design of therapies.Objectives: Combining native MS with biochemical/biophysical assays we will:1. Explore how small molecule inhibitors affect IAPP/A-beta aggregation: using MS, IMS and kinetic assays (ThT fluorescence) we will screen for new small molecule inhibitors of aggregation using 'hits' recently discovered in SER's lab (Nature Chem (2015) and Nature Chem Biol (2016)). Using MS mapping with ETD (FS: Anal Chem (2016)) combined with other solution-based assays we will elucidate which species are responsible for interactions with different inhibitors and characterize inhibitor binding interfaces. We will then use cell biological assays to determine whether the inhibitors affect cytotoxicity.2. Explore the sequence-dependence of aggregation: using a novel screen for aggregation developed in SER's laboratory (Nature Chem Biol (2016)) we will select IAPP/A-beta sequences with altered aggregation properties. We will then select for mutations that enhance/retard disease progression and thus, elucidate how these mutations perturb folding and aggregation. Novelty/Timeliness:To our knowledge the project will reveal the first atomistic insights into the energy landscapes of folding/aggregation. By focusing on peptides that are the causative agent of amyloid associated with type II diabetes/Alzheimer's the results will also have impact on human health today.Experimental Approach:We will utilize native MS to the full for the project, and combine this with chemical biology, biophysics and cell biology, offering the student training in a wide range of techniques. All methods are up and running, enabling the student to make rapid progress.