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首页> 外文期刊>Proceedings of the National Academy of Sciences of the United States of America >The curli nucleator protein, CsgB, contains an amyloidogenic domain that directs CsgA polymerization
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The curli nucleator protein, CsgB, contains an amyloidogenic domain that directs CsgA polymerization

机译：卷曲成核蛋白CsgB包含淀粉样蛋白结构域，可指导CsgA聚合

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摘要

Curli are functional amyloid fibers assembled by enteric bacteria such as Escherichia coli and Salmonella spp. In E. coli, the polymerization of the major curli fiber subunit protein CsgA into an amyloid fiber depends on the minor curli subunit protein, CsgB. The outer membrane-localized CsgB protein shares ≈30% sequence identity with the amyloid-forming protein CsgA, suggesting that CsgB might also have amyloidogenic properties. Here, we characterized the biochemical properties of CsgB and the molecular basis for CsgB-mediated nucleation of CsgA. Deletion analysis revealed that a CsgB molecule missing 19 amino acids from its C terminus (CsgB_(trunc)) was not outer membrane-associated, but secreted away from the cell. CsgB_(trunc) was overexpressed and purified from the extracellular milieu of cells as an SDS-soluble, nonaggregated protein. Soluble CsgB_(trunc) assembled into fibers that bound to the amyloid-specific dyes Congo red and thioflavin-T. CsgB_(trunc) fibers were able to seed soluble CsgA polymerization in vitro. CsgB_(trunc) displayed modest nucleator activity in vivo, as demonstrated by its ability to convert extracellular CsgA into an amyloid fiber. Unlike WT CsgB, CsgB_(trunc) was only able to act as a nucleator when cells were genetically manipulated to secrete higher concentrations of CsgA. This work represents a unique demonstration of functional amyloid nucleation and it suggests an elegant model for how E. coli guides efficient amyloid fiber formation on the cell surface.
机译：Curli是功能性淀粉样蛋白纤维，由肠道细菌（如大肠杆菌和沙门氏菌）组装而成。在大肠杆菌中，主要卷曲蛋白亚基蛋白CsgA聚合成淀粉样纤维取决于次要卷曲蛋白亚基蛋白CsgB。外膜定位的CsgB蛋白与淀粉样蛋白CsgA共有约30％的序列同一性，这表明CsgB也可能具有淀粉样蛋白生成特性。在这里，我们表征了CsgB的生化特性和CsgB介导的CsgA成核的分子基础。缺失分析显示，CsgB分子的C末端缺少19个氨基酸（CsgB_（trunc））与外膜无关，但从细胞中分泌出来。 CsgB_（trunc）作为SDS可溶性非聚集蛋白从细胞的细胞外环境中过表达和纯化。可溶性CsgB_（trunc）组装成与淀粉样特异性染料刚果红和硫黄素T结合的纤维。 CsgB_（trunc）纤维能够在体外播种可溶性CsgA聚合。 CsgB_（trunc）在体内显示出适度的成核剂活性，如其将细胞外CsgA转化为淀粉样纤维的能力所证明。与野生型CsgB不同，CsgB_（trunc）仅在通过基因操作细胞分泌更高浓度的CsgA时才能够充当成核剂。这项工作代表了功能性淀粉样蛋白成核作用的独特展示，它为大肠杆菌如何指导细胞表面上有效的淀粉样蛋白纤维形成提供了一个优雅的模型。

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