Predicted Loop Regions Promote Aggregation: A Study of Amyloidogenic Domains in the Functional Amyloid FapC.,Journal of Molecular Biology

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Predicted Loop Regions Promote Aggregation: A Study of Amyloidogenic Domains in the Functional Amyloid FapC.
Journal of Molecular Biology ( IF 4.7 ) Pub Date : 2020-02-19 , DOI: 10.1016/j.jmb.2020.01.044
Madhu Nagaraj ₁ , Mumdooh Ahmed ₂ , Jeppe Lyngsø ₃ , Brian Stougaard Vad ₁ , Andreas Bøggild ₁ , Anne Fillipsen ₁ , Jan Skov Pedersen ₃ , Daniel Erik Otzen ₄ , Ümit Akbey ₅

Affiliation

Protein fibrillation is traditionally associated with misfolding, loss of functional phenotype, and gain of toxicity in neurodegenerative diseases. However, many organisms exploit fibrils in the form of functional amyloids (FA), as seen in bacteria, such as E. coli, Salmonella, Bacillus, and Pseudomonas. Here, we provide structural information and mechanistic data for fibrillation of the smallest amyloidogenic truncation unit along with the full-length version (FL) of the major amyloid protein FapC from Pseudomonas, predicted to consist of three β-hairpin-forming imperfect repeats separated by disordered regions. Using a series of truncation mutants, we establish that the putative loops (linkers) increase the rate of aggregation. The minimal aggregation unit consisting of a single repeat with flanking disordered regions (R3C) aggregates in a pathway dominated by secondary nucleation, in contrast to the primary nucleation favored by full-length (FL) FapC. SAXS on FapC FL, R3C, and remaining truncation constructs resolves two major coexisting species in the fibrillation process, namely pre-fibrillar loosely aggregated monomers, and cylindrical, elliptical cross-section fibrils. Solid-state NMR spectra identified rigid parts of the FapC fibril. We assigned Cα-Cβ chemical shifts, indicative of a predominant β-sheet topology with some α-helix or loop chemical shifts. Our work emphasizes the complex nature of FapC fibrillation. In addition, we are able to deduce the importance of non-repeat regions (i.e., predicted loops), which enhance the amyloid protein aggregation and their influence on the polymorphism of the fibril architecture.

中文翻译：

预测的环区域促进聚集：功能性淀粉样蛋白FapC中的淀粉样蛋白形成域的研究。

传统上，蛋白原纤化与神经退行性疾病中的错误折叠，功能表型丧失和毒性增加有关。但是，如细菌（例如大肠杆菌，沙门氏菌，芽孢杆菌和假单胞菌）中所见，许多生物利用功能性淀粉样蛋白（FA）形式的原纤维。在这里，我们提供了最小的淀粉样蛋白生成截短单位的原纤维化的结构信息和力学数据，以及来自假单胞菌的主要淀粉样蛋白FapC的全长版本（FL），预计其由三个由β-发夹形成的不完美重复组成，无序区域。使用一系列的截断突变体，我们确定推定的环（接头）增加了聚集的速率。与全长（FL）FapC促进的初级成核相反，最小的聚集单位由具有侧翼无序区（R3C）的单个重复组成，聚集在次级成核为主的途径中。FapC FL，R3C和剩余的截短构建体上的SAXS解决了原纤化过程中两个主要共存的物种，即原纤维状的松散聚集单体和圆柱形，椭圆形横截面的原纤维。固态NMR光谱确定了FapC原纤维的刚性部分。我们分配了Cα-Cβ化学位移，该化学位移表示具有某些α-螺旋或环化学位移的主要β-折叠拓扑。我们的工作强调了FapC纤颤的复杂性。此外，我们能够推断出非重复区域（即预测循环）的重要性，

更新日期：2020-02-20

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