BioID as a Tool for Protein-Proximity Labeling in Living Cells
BioID as a Tool for Protein-Proximity Labeling in Living Cells
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DOI：
10.1007/978-1-4939-9546-2_15
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发表时间：
2019-01-01
期刊：
影响因子：
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通讯作者：
Roux, Kyle J.
中科院分区：
文献类型：
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作者：
Sears, Rhiannon M.;May, Danielle G.;Roux, Kyle J.
BioID已经成为识别活细胞中候选蛋白质-蛋白质相互作用(PPI)的一种越来越多的工具。这种方法利用了一种被称为BioID的混杂生物素连接酶，融合到一种感兴趣的蛋白质上，当在细胞中表达时，可以诱导在几个小时内相互作用和接近的蛋白质生物素化，从而产生蛋白质结合的历史。这些生物素化的蛋白质随后被提纯，并通过质谱仪进行鉴定。与通常用于筛选强PPI的其他传统方法相比，BioID允许在定义的一段时间内检测相关生物环境中的微弱和瞬时相互作用。在这里，我们简要回顾了BioID技术带来的科学进步，详细介绍了将该方法应用于活细胞中蛋白质的最新方案，并提供了对常见挫折进行故障排除的见解。
BioID has become an increasingly utilized tool for identifying candidate protein-protein interactions (PPIs) in living cells. This method utilizes a promiscuous biotin ligase, called BioID, fused to a protein of interest that when expressed in cells can be induced to biotinylate interacting and proximate proteins over a period of hours, thus generating a history of protein associations. These biotinylated proteins are subsequently purified and identified via mass spectrometry. Compared to other conventional methods typically used to screen strong PPIs, BioID allows for the detection of weak and transient interactions within a relevant biological setting over a defined period of time. Here we briefly review the scientific progress enabled by the BioID technology, detail an updated protocol for applying the method to proteins in living cells, and offer insights for troubleshooting commonly encountered setbacks.