文献类型： 外文期刊

第一作者： Lends, Alons

作者： Lends, Alons;Daskalov, Asen;Delamare, Aline;Berbon, Melanie;Grelard, Axelle;Shenoy, Jayakrishna;Dutour, Antoine;Tolchard, James;Noubhani, Abdelmajid;Giraud, Marie-France;Sanchez, Corinne;Habenstein, Birgit;Guichard, Gilles;Compain, Guillaume;Loquet, Antoine;Daskalov, Asen;Saupe, Sven J.;Daskalov, Asen;Maleckis, Ansis;Jaudzems, Kristaps;Morvan, Estelle

作者机构：

期刊名称：COMMUNICATIONS BIOLOGY （ 影响因子：6.548； 五年影响因子：6.816 ）

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年卷期： 2022 年 5 卷 1 期

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收录情况： SCI

摘要： Structural investigations of amyloid fibrils often rely on heterologous bacterial overexpression of the protein of interest. Due to their inherent hydrophobicity and tendency to aggregate as inclusion bodies, many amyloid proteins are challenging to express in bacterial systems. Cell-free protein expression is a promising alternative to classical bacterial expression to produce hydrophobic proteins and introduce NMR-active isotopes that can improve and speed up the NMR analysis. Here we implement the cell-free synthesis of the functional amyloid prion HET-s(218-289). We present an interesting case where HET-s(218-289) directly assembles into infectious fibril in the cell-free expression mixture without the requirement of denaturation procedures and purification. By introducing tailored C-13 and N-15 isotopes or CF3 and (CH2F)-C-13 labels at strategic amino-acid positions, we demonstrate that cell-free synthesized amyloid fibrils are readily amenable to high-resolution magic-angle spinning NMR at sub-milligram quantity. A method for cell-free synthesis of amyloid fibrils is reported that also allows for solid-state NMR analysis to characterize complex and pathological fibrillary assemblies.

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