A promiscuous biotin ligase fusion protein identifies proximal and interacting proteins in mammalian cells.
A promiscuous biotin ligase fusion protein identifies proximal and interacting proteins in mammalian cells.
复制标题
DOI：
10.1083/jcb.201112098
复制
发表时间：
2012-03-19
期刊：
影响因子：
--
通讯作者：
Burke B
中科院分区：
文献类型：
--
作者：
Roux KJ;Kim DI;Raida M;Burke B
邻近依赖性生物素识别（BioID）是一种利用生物素连接酶融合蛋白来识别天然细胞环境中相互作用蛋白和邻近蛋白的新方法。我们已经开发了一种新的技术，在真核细胞中的蛋白质的邻近依赖性标记。命名为BioID的邻近依赖性生物素鉴定，这种方法是基于融合的混杂大肠杆菌生物素蛋白连接酶的靶向蛋白。BioID的特点是融合蛋白的近邻蛋白质的邻近依赖性生物素化。生物素化的蛋白质可以通过亲和捕获分离并通过质谱法鉴定。我们将BioID应用于核纤层蛋白A（LaA），这是一种具有良好特征的中间丝蛋白，是核纤层的组成部分，核被膜（NE）的重要结构元件。我们确定了多种蛋白质，与和/或接近体内的LaA。其中最丰富的包括位于NE的已知LaA相互作用物，以及一种名为SLAP 75的新NE相关蛋白。我们的研究结果表明，BioID是一种有用的和普遍适用的方法来筛选相互作用和相邻的蛋白质在其天然细胞环境。
Proximity-dependent biotin identification (BioID) is a new approach making use of biotin ligase fusion proteins for the identification of both interacting and neighboring proteins in their native cellular environment. We have developed a new technique for proximity-dependent labeling of proteins in eukaryotic cells. Named BioID for proximity-dependent biotin identification, this approach is based on fusion of a promiscuous Escherichia coli biotin protein ligase to a targeting protein. BioID features proximity-dependent biotinylation of proteins that are near-neighbors of the fusion protein. Biotinylated proteins may be isolated by affinity capture and identified by mass spectrometry. We apply BioID to lamin-A (LaA), a well-characterized intermediate filament protein that is a constituent of the nuclear lamina, an important structural element of the nuclear envelope (NE). We identify multiple proteins that associate with and/or are proximate to LaA in vivo. The most abundant of these include known interactors of LaA that are localized to the NE, as well as a new NE-associated protein named SLAP75. Our results suggest BioID is a useful and generally applicable method to screen for both interacting and neighboring proteins in their native cellular environment.
登录
查看更多内容
DOI：
10.1002/ar.a.20334
发表时间：
2006-07-01
期刊：
ANATOMICAL RECORD PART A-DISCOVERIES IN MOLECULAR CELLULAR AND EVOLUTIONARY BIOLOGY
影响因子：
--
作者：
Holaska, James M.;Wilson, Katherine L.
通讯作者：
Wilson, Katherine L.
影响因子：
4
作者：
Gudise, Santhosh;Figueroa, Ricardo A.;Hallberg, Einar
通讯作者：
Hallberg, Einar
影响因子：
5.6
作者：
Cronan, JE
通讯作者：
Cronan, JE
影响因子：
8
作者：
Choi-Rhee, E;Schulman, H;Cronan, JE
通讯作者：
Cronan, JE
影响因子：
3.7
作者：
Montes de Oca R;Shoemaker CJ;Gucek M;Cole RN;Wilson KL
通讯作者：
Wilson KL