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首页> 外文期刊>Biotechnology and Bioengineering >RepTAGs: Universal tags for isolation and labeling of proteins, for labeling live mammalian cells and for drug discovery
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RepTAGs: Universal tags for isolation and labeling of proteins, for labeling live mammalian cells and for drug discovery

机译：RepTAGs：用于分离和标记蛋白质，标记活哺乳动物细胞和用于药物发现的通用标签

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摘要

Methods for specific immobilization, isolation and labeling of proteins are central to the elucidation of cellular functions. Based on bacterial repressor proteins, which bind to specific target sequences in response to small molecules (macrolide and tetracycline antibiotics) or environmental parameters (temperature), we have developed a set of protein tags (RepTAGs), which enable reversible immobilization of the protein of interest on a solid support for the isolation and quantification as well as for the specific labeling of target proteins with fluorescent dyes for tracking them within a complex protein mixture. Similarly, live mammalian cells were specifically labeled with a fluorescent operator sequence bound to RepTAGs, which were directed towards the cell surface for easy discrimination between transfected and untransfected cell populations. Based on the drug-responsive RepTAG-DNA interactions, it was also possible to quantify or discover antibiotics in environmental samples or compound libraries by means of rapid, sensitive detection methods involving fluorescence polarization and bioluminescence. We believe that the universally applicable RepTAGs will become essential for the analysis and manipulation of proteins in the most diverse areas of protein chemistry and cell biology.
机译：蛋白质特异性固定，分离和标记的方法对于阐明细胞功能至关重要。基于细菌阻遏蛋白，它们可响应小分子（大环内酯和四环素抗生素）或环境参数（温度）而与特定的靶序列结合，我们开发了一套蛋白质标签（RepTAGs），可实现蛋白质的可逆固定化。对固体支持物的分离，定量以及用荧光染料对目标蛋白进行特异性标记以在复杂蛋白混合物中追踪它们的兴趣。类似地，用与RepTAG结合的荧光操纵子序列特异性标记活的哺乳动物细胞，该序列指向细胞表面，以便于区分转染和未转染的细胞群。基于药物反应性RepTAG-DNA的相互作用，还可以通过涉及荧光偏振和生物发光的快速，灵敏的检测方法来定量或发现环境样品或化合物库中的抗生素。我们相信，普遍适用的RepTAGs将在蛋白质化学和细胞生物学最广泛的领域中对蛋白质的分析和操纵必不可少。

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