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Updated: Mar 17, 2026

A Facile Protocol to Generate Site-Specifically Acetylated Proteins in Escherichia Coli
Published on: December 9, 2017
Expanding the genetic code of Escherichia coli with phosphotyrosine
Chenguang Fan1,2, Kevan Ip2, Dieter Söll2,3
1Department of Chemistry and Biochemistry, University of Arkansas, Fayetteville, AR, USA.
Abstract:
Protein phosphorylation is one of the most important post-translational modifications in nature. However, the site-specific incorporation of O-phosphotyrosine into proteins in vivo has not yet been reported. Endogenous phosphatases present in cells can dephosphorylate phosphotyrosine as a free amino acid or as a protein residue. Therefore, we deleted the genes of five phosphatases from the genome of Escherichia coli with the aim of stabilizing phosphotyrosine. Together with an engineered aminoacyl-tRNA synthetase (derived from Methanocaldococcus jannaschii tyrosyl-tRNA synthetase) and an elongation factor Tu variant, we were able to cotranslationally incorporate O-phosphotyrosine into the superfolder green fluorescent protein at a desired position in vivo. This system will facilitate future studies of tyrosine phosphorylation.
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