The Strep-tag system for one-step purification and high-affinity detection or capturing of proteins

被引:476
作者
Schmidt, Thomas G. M. [2 ]
Skerra, Arne [1 ]
机构
[1] Tech Univ Munich, Lehrstuhl Biol Chem, D-85350 Freising Weihenstephan, Germany
[2] IBA GmbH, D-37079 Gottingen, Germany
关键词
D O I
10.1038/nprot.2007.209
中图分类号
Q5 [生物化学];
学科分类号
071010 ; 081704 ;
摘要
The Strep-tag II is an eight-residue minimal peptide sequence (Trp-Ser-His-Pro-Gln-Phe-Glu-Lys) that exhibits intrinsic affinity toward streptavidin and can be fused to recombinant proteins in various fashions. We describe a protocol that enables quick and mild purification of corresponding Strep-tag II fusion proteins - including their complexes with interacting partners - both from bacterial and eukaryotic cell lysates using affinity chromatography on a matrix carrying an engineered streptavidin (Strep-Tactin), which can be accomplished within 1 h. A high- affinity monoclonal antibody (StrepMAB-Immo) permits stable immobilization of Strep-tag II fusion proteins to solid surfaces, for example, for surface plasmon resonance analysis. Selective and sensitive detection on western blots is achieved with Strep-Tactin/enzyme conjugates or another monoclonal antibody (StrepMAB-Classic). Thus, the Strep-tag II, which is short, biologically inert, proteolytically stable and does not interfere with membrane translocation or protein folding, offers a versatile tool both for the rapid isolation of a functional gene product and for its detection or molecular interaction analysis.
引用
收藏
页码:1528 / 1535
页数:8
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