Fig. 3: Evolution of chimeric aaRSs. | Nature Communications

Fig. 3: Evolution of chimeric aaRSs.

From: Directed-evolution of translation system for efficient unnatural amino acids incorporation and generalizable synthetic auxotroph construction

Fig. 3

a Directed evolution of the chPheRS for efficient genetic code expansion. The evolution trajectory of representative clones shows increased amber suppression activity compared to the progenitor chPheRS. Mutations in these representative clones are shown. b Structure of C-terminal domain of human mitochondrial PheRS, in which mutation residues in clones 12D4 are shown in sticks and phenylalanyl-adenylate in the amino acid binding pocket is highlighted in magenta. c Amber suppression efficiency of the evolved 12D4- or 13E3-AzFRS-1/chPheT pairs is tested by the GFP reporter assay. d Amber suppression efficiency of the evolved 12D4-AzFRS-2/chPheT pairs is tested, in which the 12D4-AzFRS-2 is under the control of oxb20 or glns promoter, respectively. e Mass spectrometry characterization of the fidelity of AzF incorporation into GFP with the 12D4-AzFRS-2/3C11-chPheT pair. The expected molecular weight (MW) 27796 Da; observed 27797 Da. Error bars represents ±standard error of the mean from three biologically independent experiments. Source data are provided as a Source Data file.

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