Structural analysis of SAA-amyloid fibrils by cryo-EM and NMR

通过冷冻电镜和核磁共振对 SAA 淀粉样原纤维进行结构分析

• 批准号: 313913757
• 负责人: Professor Dr. Bernd Reif
• 金额: --
• 依托单位: Professur für Festkörper-NMR-Spektroskopie
• 依托单位国家: 德国
• 项目类别: Research Grants
• 财政年份: 2016
• 资助国家: 德国
• 起止时间: 2015-12-31 至 2021-12-31
• 项目状态: 已结题
• 来源: https://gepris.dfg.de/gepris/projekt/313913757?language=en
• 关键词: Structural; analysis; SAA; amyloid; fibrils

AA amyloidosis presents, among all protein misfolding diseases, unique advantages for studying the structure, formation and propagation of pathogenic amyloids. There are highly relevant in vitro, cell culture and mouse model systems to reproduce crucial steps of amyloid fibril formation. Amyloid deposits are comparatively large-sized and amenable for biochemical analysis. Murine SAA1 is an extracellular protein and small enough to be well accessible for NMR. These features enable us to take an integrative structural biology approach in which we use cryo-electron microscopy (EM) and solid-state NMR spectroscopy to analyze the structure of the fibrils. These fibril structures are relevant for disease because they give rise to tissue damage and underlie the spreading of disease and protein misfolding inside the tissue. The two major techniques employed in this project, cryo-EM and solid-state NMR, are perfectly complementary. Cryo-EM can determine the overall electron density, symmetry and protofilament substructure and is amenable to body-derived fibrils. Solid-state NMR is an ideal tool to characterize dynamic regions within the aggregate. In this project we want to complementarily apply these techniques to study the structure of pathogenically relevant AA amyloid fibrils by examining bona fide fibrils from murine spleen, cell culture derived and in vitro formed fibrils.

在所有蛋白质错误折叠疾病中，AA淀粉样变性在研究致病性淀粉样蛋白的结构、形成和繁殖方面具有独特的优势。存在高度相关的体外、细胞培养和小鼠模型系统来再现淀粉样蛋白原纤维形成的关键步骤。淀粉样蛋白沉积物是相对大的尺寸和适合生化分析。小鼠SAA 1是一种细胞外蛋白，小到足以很好地进行NMR。这些功能使我们能够采取综合结构生物学方法，其中我们使用冷冻电子显微镜（EM）和固态NMR光谱分析原纤维的结构。这些原纤维结构与疾病有关，因为它们引起组织损伤并成为疾病传播和组织内蛋白质错误折叠的基础。在这个项目中采用的两种主要技术，冷冻EM和固态NMR，是完全互补的。Cryo-EM可以确定整体电子密度，对称性和原丝子结构，并适合于身体来源的原纤维。固体核磁共振是表征聚集体内动态区域的理想工具。在这个项目中，我们想补充应用这些技术来研究致病相关的AA淀粉样蛋白纤维的结构，通过检查真正的纤维从小鼠脾脏，细胞培养衍生和体外形成的纤维。