TurboID 实现活细胞和生物体内高效的邻近标记。
Efficient proximity labeling in living cells and organisms with TurboID.
机构信息
Department of Chemistry, Massachusetts Institute of Technology, Cambridge, Massachusetts, USA.
Departments of Genetics, Stanford University, Stanford, California, USA.
出版信息
Nat Biotechnol. 2018 Oct;36(9):880-887. doi: 10.1038/nbt.4201. Epub 2018 Aug 20.
Protein interaction networks and protein compartmentalization underlie all signaling and regulatory processes in cells. Enzyme-catalyzed proximity labeling (PL) has emerged as a new approach to study the spatial and interaction characteristics of proteins in living cells. However, current PL methods require over 18 h of labeling time or utilize chemicals with limited cell permeability or high toxicity. We used yeast display-based directed evolution to engineer two promiscuous mutants of biotin ligase, TurboID and miniTurbo, which catalyze PL with much greater efficiency than BioID or BioID2, and enable 10-min PL in cells with non-toxic and easily deliverable biotin. Furthermore, TurboID extends biotin-based PL to flies and worms.
蛋白质相互作用网络和蛋白质区室化是细胞中所有信号转导和调节过程的基础。酶催化的邻近标记（PL）已成为研究活细胞中蛋白质空间和相互作用特征的一种新方法。然而，目前的 PL 方法需要超过 18 小时的标记时间，或者使用细胞通透性有限或毒性高的化学物质。我们使用基于酵母展示的定向进化工程，构建了两种具有广泛结合活性的生物素连接酶突变体 TurboID 和 miniTurbo，它们比 BioID 或 BioID2 具有更高的 PL 效率，并能够在非毒性且易于传递的生物素存在下，在 10 分钟内对细胞进行 PL。此外，TurboID 将基于生物素的 PL 扩展到了果蝇和线虫。
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