Department of Chemistry, Massachusetts Institute of Technology, 16-573a, 77 Massachusetts Avenue, Cambridge, Massachusetts 02139, USA.
J Am Chem Soc. 2012 Jul 4;134(26):10749-52. doi: 10.1021/ja302354v. Epub 2012 Jun 19.
Proteins containing a C-terminal thioester are important intermediates in semisynthesis. Currently there is one main method for the synthesis of protein thioesters that relies upon the use of engineered inteins. Here we report a simple strategy, utilizing sortase A, for routine preparation of recombinant proteins containing a C-terminal (α)thioester. We used our method to prepare two different anthrax toxin cargo proteins: one containing an (α)thioester and another containing a D-polypeptide segment situated between two protein domains. We show that both variants can translocate through protective antigen pore. This new method to synthesize a protein thioester allows for interfacing of sortase-mediated ligation and native chemical ligation.
含有 C 端硫酯的蛋白质是半合成中的重要中间体。目前，合成蛋白质硫酯的主要方法之一依赖于工程化内含肽的使用。在这里，我们报告了一种利用 sortase A 常规制备含有 C 端（α）硫酯的重组蛋白的简单策略。我们使用该方法制备了两种不同的炭疽毒素货物蛋白：一种含有（α）硫酯，另一种含有位于两个蛋白结构域之间的 D 多肤片段。我们表明，这两种变体都可以通过保护性抗原孔转移。这种合成蛋白质硫酯的新方法允许连接酶介导的连接和天然化学连接。
