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用于十二烷基硫酸钠-聚丙烯酰胺凝胶电泳(SDS-PAGE)分析的细胞表面蛋白生物素化

Cell surface protein biotinylation for SDS-PAGE analysis.

作者信息

Elia Giuliano

机构信息

University College Dublin, Belfield, Dublin, Ireland.

出版信息

Methods Mol Biol. 2012;869:361-72. doi: 10.1007/978-1-61779-821-4_29.

Abstract

Cell surface proteins play a very important role in physiology and pathology and are receiving increased attention by the pharmaceutical industry as valuable targets for development of new therapeutics. However, owing to the very nature of this category of proteins, their comprehensive study remains an elusive task. A number of methods have been proposed to enrich and purify cell surface proteins. Among them, usage of biotinylating reagents and exploitation of the strong interaction between biotin and streptavidin for the purification of biotinylated proteins has rapidly gained in popularity and allowed some of the most significant progresses in quantitative proteomics. This chapter focuses on methods for cell surface biotinylation with commercially available reagents, capture by avidin-affinity chromatography and release of the biotinylated surface proteins for downstream analysis by electrophoretic techniques.

摘要

细胞表面蛋白在生理学和病理学中发挥着非常重要的作用,作为新疗法开发的有价值靶点,正受到制药行业越来越多的关注。然而,由于这类蛋白的特性,对其进行全面研究仍然是一项难以捉摸的任务。已经提出了许多方法来富集和纯化细胞表面蛋白。其中,使用生物素化试剂以及利用生物素与链霉亲和素之间的强相互作用来纯化生物素化蛋白的方法迅速受到欢迎,并在定量蛋白质组学中取得了一些最重要的进展。本章重点介绍使用市售试剂进行细胞表面生物素化、通过抗生物素蛋白亲和色谱法捕获以及释放生物素化表面蛋白以通过电泳技术进行下游分析的方法。

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