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Title: Versatile Protein Biotinylation Strategies for Potential High-Throughput Proteomics
Authors: Lue, R.Y.P.
Chen, G.Y.J. 
Hu, Y.
Zhu, Q. 
Yao, S.Q. 
Issue Date: 4-Feb-2004
Citation: Lue, R.Y.P., Chen, G.Y.J., Hu, Y., Zhu, Q., Yao, S.Q. (2004-02-04). Versatile Protein Biotinylation Strategies for Potential High-Throughput Proteomics. Journal of the American Chemical Society 126 (4) : 1055-1062. ScholarBank@NUS Repository.
Abstract: We present intein-mediated approaches for efficient biotinylation of proteins site-specifically. The reactive C-terminal thioester generated from intein-assisted protein splicing (either in vitro or in live cells) served as an attractive and exclusive site for attaching cysteine-containing biotin. Using these novel biotinylation strategies, we were able to efficiently biotinylate many proteins from different biological sources in a potentially high-throughput, high-content fashion. Some of these proteins were subsequently immobilized, in a very simple manner, onto different avidin-functionalized solid surfaces for applications such as protein microarray and surface plasmon resonance (SPR) spectroscopy, highlighting the numerous advantages of using biotin over other tags (e.g., GST, His-tag, etc.) as the method of choice in protein purification/immobilization. In addition, our intein-mediated strategies provided critical advantages over other protein biotinylation strategies in a number of ways. For the first time, we also successfully demonstrated that intein-mediated protein biotinylation proceeded adequately inside both bacterial and mammalian living cells, as well as in a cell-free protein synthesis system. Taken together, our results indicate the versatility of these intein-mediated strategies for potential high-throughput proteomics applications. They may also serve as useful tools for various biochemical and biophysical studies of proteins both in vitro and in vivo.
Source Title: Journal of the American Chemical Society
ISSN: 00027863
DOI: 10.1021/ja037914g
Appears in Collections:Staff Publications

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