Department of Chemistry and Skaggs Institute for Chemical Biology , The Scripps Research Institute , 10550 N Torrey Pines Road , La Jolla , California 92037 , United States.
Department of Chemistry , Washington State University , Pullman , Washington 99164-4630 , United States.
ACS Chem Biol. 2018 Mar 16;13(3):578-581. doi: 10.1021/acschembio.7b00998. Epub 2018 Feb 9.
Here, we report the site-specific incorporation of a thioester containing noncanonical amino acid (ncAA) into recombinantly expressed proteins. Specifically, we genetically encoded a thioester-activated aspartic acid (ThioD) in bacteria in good yield and with high fidelity using an orthogonal nonsense suppressor tRNA/aminoacyl-tRNA synthetase (aaRS) pair. To demonstrate the utility of ThioD, we used native chemical ligation to label green fluorescent protein with a fluorophore in good yield.
在这里，我们报告了在重组表达蛋白中特异性掺入含有硫酯的非天然氨基酸（ncAA）。具体来说，我们使用正交的无意义抑制 tRNA/氨酰-tRNA 合成酶（aaRS）对在细菌中以高产率和高保真度遗传编码硫酯激活的天冬氨酸（ThioD）。为了证明 ThioD 的实用性，我们使用天然化学连接以高产率将荧光团标记到绿色荧光蛋白上。
