A general method for the covalent labeling of fusion proteins with small molecules in vivo
A general method for the covalent labeling of fusion proteins with small molecules in vivo
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DOI：
10.1038/nbt765
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发表时间：
2003-01-01
影响因子：
46.9
通讯作者：
Johnsson, K
中科院分区：
文献类型：
--
作者：
Keppler, A;Gendreizig, S;Johnsson, K
表征活细胞内蛋白质的运动、相互作用和化学微环境对于详细了解其功能至关重要。旨在实现这一目标的大多数策略是基于将感兴趣的蛋白质与监测偶联蛋白质的环境变化的报告蛋白质遗传融合。实例包括与荧光蛋白的融合、酵母双杂交系统和分裂泛素（1-3）。然而，这些技术具有各种限制，并且相当大的努力致力于用能够探测和调节其功能的小合成分子在体内特异性标记蛋白质。这些方法目前基于小分子与蛋白质的非共价结合、双砷化合物与含有半胱氨酸的肽之间形成稳定复合物或使用生物素受体结构域（4-10）。在这里，我们描述了一种通用的方法，在体内的融合蛋白的共价标记，补充现有的方法，非共价标记的蛋白质，并可能开辟新的方法研究活细胞中的蛋白质。
Characterizing the movement, interactions, and chemical microenvironment of a protein inside the living cell is crucial to a detailed understanding of its function. Most strategies aimed at realizing this objective are based on genetically fusing the protein of interest to a reporter protein that monitors changes in the environment of the coupled protein. Examples include fusions with fluorescent proteins, the yeast two-hybrid system, and split ubiquitin(1-3). However, these techniques have various limitations, and considerable effort is being devoted to specific labeling of proteins in vivo with small synthetic molecules capable of probing and modulating their function. These approaches are currently based on the noncovalent binding of a small molecule to a protein, the formation of stable complexes between biarsenical compounds and peptides containing cysteines, or the use of biotin acceptor domains(4-10). Here we describe a general method for the covalent labeling of fusion proteins in vivo that complements existing methods for noncovalent labeling of proteins and that may open up new ways of studying proteins in living cells.