Wang Yanxin J, Szantai-Kis D Miklos, Petersson E James
Department of Chemistry, University of Pennsylvania, 231 South 34th Street, Philadelphia, PA 19104, USA.
Org Biomol Chem. 2016 Jul 14;14(26):6262-9. doi: 10.1039/c6ob01020b. Epub 2016 Jun 6.
Thioamides are single-atom substitutions of canonical amide bonds, and have been proven to be versatile and minimally perturbing probes in protein folding studies. Previously, our group showed that thioamides can be incorporated into proteins by native chemical ligation (NCL) with Cys as a ligation handle. In this study, we report the expansion of this strategy into non-Cys ligation sites, utilizing radical initiated desulfurization to "erase" the side chain thiol after ligation. The reaction exhibited high chemoselectivity against thioamides, which can be further enhanced with thioacetamide as a sacrificial scavenger. As a proof-of-concept example, we demonstrated the incorporation of a thioamide probe into a 56 amino acid protein, the B1 domain of Protein G (GB1). Finally, we showed that the method can be extended to β-thiol amino acid analogs and selenocysteine.
硫代酰胺是典型酰胺键的单原子取代物，并且已被证明是蛋白质折叠研究中用途广泛且干扰极小的探针。此前，我们小组表明硫代酰胺可以通过以半胱氨酸作为连接手柄的天然化学连接（NCL）引入蛋白质中。在本研究中，我们报告了将该策略扩展到非半胱氨酸连接位点，利用自由基引发的脱硫反应在连接后 “去除” 侧链硫醇。该反应对硫代酰胺表现出高化学选择性，使用硫代乙酰胺作为牺牲清除剂可进一步提高选择性。作为概念验证的示例，我们展示了将硫代酰胺探针引入一种56个氨基酸的蛋白质 —— 蛋白G（GB1）的B1结构域中。最后，我们表明该方法可以扩展到β-硫醇氨基酸类似物和硒代半胱氨酸。
