Translational incorporation of modified phenylalanines and tyrosines during cell-free protein synthesis

Inherent promiscuity of bacterial translation is demonstrated by mass spectrometric quantification of the translational incorporation of ring-substituted phenylalanines and tyrosines bearing fluoro-, hydroxyl-, methyl-, chloro-and nitro-groups in an E. coli-derived cell-free system. Competitive studies using the cell-free system show that the aminoacyl-tRNA synthetases (aaRS) have at least two orders of magnitude higher specificity for the native substrate over these structural analogues, which correlates with studies on the purified synthetase.