Split-BioID: a proximity biotinylation assay for dimerization-dependent protein interactions
Split-BioID: a proximity biotinylation assay for dimerization-dependent protein interactions
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DOI：
10.1002/1873-3468.12548
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发表时间：
2017-01-01
期刊：
影响因子：
3.5
通讯作者：
Bollen, Mathieu
中科院分区：
文献类型：
--
作者：
De Munter, Sofie;Gornemann, Janina;Bollen, Mathieu
生物素鉴定(BioID)方案使用生物素连接酶BIRA(BIRA*)的突变体与感兴趣的蛋白质融合，使完整细胞中的邻近蛋白质生物素化。在这里，我们证明了BIRA*的两个非活性部分分别与蛋白磷酸酶PP1的催化和调节亚基融合，在磷酸酶亚单位异源二聚的基础上重建了功能上的BIRA*酶。我们还证明了这种被称为Split-BioID的BIRA*片段互补方法可以用于筛选PP1全酶的底物和其他蛋白质相互作用因子。Split-BioID是一种新的、通用的工具，用于识别蛋白质二聚体的(瞬时)相互作用。
The biotin identification (BioID) protocol uses a mutant of the biotin ligase BirA (BirA*) fused to a protein-of-interest to biotinylate proximate proteins in intact cells. Here, we show that two inactive halves of BirA* separately fused to a catalytic and regulatory subunit of protein phosphatase PP1 reconstitute a functional BirA* enzyme upon heterodimerization of the phosphatase subunits. We also demonstrate that this BirA* fragment complementation approach, termed split-BioID, can be used to screen for substrates and other protein interactors of PP1 holoenzymes. Split-BioID is a novel and versatile tool for the identification of (transient) interactors of protein dimers.