A Genetically Encoded Acrylamide Functionality

作者：Lee Yan Jiun; Wu Bo; Raymond Jeffrey E; Zeng Yu; Fang Xinqiang; Wooley Karen L; Liu Wenshe R^*

来源：ACS Chemical Biology, 2013, 8(8): 1664-1670.

DOI：10.1021/cb400267m

N-epsilon-Acryloyl-L-lysine, a noncanonical amino acid with an electron deficient olefin, is genetically encoded in Escherichia coli using a pyrrolysyl-tRNA synthetase mutant in coordination with tRNA(CUA)(Pyl). The acrylamide moiety is stable in cells, whereas it is active enough to perform a diverse set of unique reactions for protein modifications in vitro. These reactions include 1,4-addition, radical polymerization, and 1,3-dipolar cycloaddition. We demonstrate that a protein incorporated with N-epsilon-acryloyl-L-lysine is efficiently modified with thiol-containing nucleophiles at slightly alkali conditions, and the acrylamide moiety also allows rapid radical copolymerization of the same protein into a polyacrylamide hydrogel at physiological pH. At physiological conditions, the acrylamide functionality undergoes a fast 1,3-dipolar cycloaddition reaction with diaryl nitrile imine to show turn-on fluorescence. We have used this observation to demonstrate site-specific fluorescent labeling of proteins incorporated with N-epsilon-acryloyl-L-lysine both in vitro and in living cells. This critical development allows easy access to an array of modified proteins for applications where high specificity and reaction efficiency are needed.

• 出版日期2013-8