Heal William P, Wickramasinghe Sasala R, Leatherbarrow Robin J, Tate Edward W
Department of Chemistry, South Kensington Campus, Imperial College, London, UK SW7 2AZ.
Org Biomol Chem. 2008 Jul 7;6(13):2308-15. doi: 10.1039/b803258k. Epub 2008 May 12.
N-Myristoyl transferase-mediated labelling using a substrate modified with an azide or alkyne tag is described as an efficient and site-selective method for the introduction of a bioorthogonal tag at the N-terminus of a recombinant protein. The procedure may be performed in vitro, or in a single over-expression/tagging step in vivo in bacteria; tagged proteins may then be captured using Staudinger-Bertozzi or 'click' chemistry protocols to introduce a secondary label for downstream analysis. The straightforward synthesis of the chemical and molecular biological tools described should enable their use in a wide range of N-terminal labelling applications.
使用经叠氮化物或炔烃标签修饰的底物进行N-肉豆蔻酰基转移酶介导的标记，被描述为在重组蛋白N端引入生物正交标签的一种高效且位点选择性的方法。该过程可在体外进行，或在细菌体内通过单一的过表达/标记步骤完成；然后可以使用施陶丁格-贝托齐反应或“点击”化学方案捕获带标签的蛋白质，以引入用于下游分析的二级标签。所描述的化学和分子生物学工具的直接合成应使其能够用于广泛的N端标记应用。
