An improved smaller biotin ligase for BioID proximity labeling.
An improved smaller biotin ligase for BioID proximity labeling.
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DOI：
10.1091/mbc.e15-12-0844
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发表时间：
2016-04-15
影响因子：
3.3
通讯作者：
Roux KJ
中科院分区：
文献类型：
--
作者：
Kim DI;Jensen SC;Noble KA;Kc B;Roux KH;Motamedchaboki K;Roux KJ
一种被称为BioID2的较小的用于邻近生物素化的混杂生物素连接酶能够更选择性地靶向融合蛋白，需要较少的生物素补充，表现出对邻近蛋白的增强标记，并展示了使用灵活的连接物来调节生物素标记半径的使用。BioID方法使用混杂的生物素连接酶来检测活细胞中的蛋白质-蛋白质结合以及邻近蛋白质。在这里，我们报告了以BioID2为中心的BioID方法的改进，BioID2是一种小得多的混杂生物素连接酶。BioID2能够更有选择性地靶向融合蛋白，需要更少的生物素补充，并表现出对邻近蛋白的增强标记。因此，BioID2提高了筛选蛋白质-蛋白质关联的效率。我们还证明，BioID2的生物素化范围可以使用灵活的连接子进行相当大的调节，从而能够对生物素标记半径进行特定用途的调整。
A smaller promiscuous biotin ligase for proximity biotinylation called BioID2 enables more-selective targeting of fusion proteins, requires less biotin supplementation, exhibits enhanced labeling of proximate proteins, and demonstrates the use of a flexible linker to modulate the biotin-labeling radius. The BioID method uses a promiscuous biotin ligase to detect protein–protein associations as well as proximate proteins in living cells. Here we report improvements to the BioID method centered on BioID2, a substantially smaller promiscuous biotin ligase. BioID2 enables more-selective targeting of fusion proteins, requires less biotin supplementation, and exhibits enhanced labeling of proximate proteins. Thus BioID2 improves the efficiency of screening for protein–protein associations. We also demonstrate that the biotinylation range of BioID2 can be considerably modulated using flexible linkers, thus enabling application-specific adjustment of the biotin-labeling radius.