Figure 1.

Genetic code expansion in Vmax X2. (A) Vmax X2 cells were transformed with pET-S2TAGsfGFP⁶⁷ and either pEVOL-CNF⁶⁶ or pEVOL-mmPyl to induce the expression of sfGFP bearing an ncAA at the second position of sfGFP. Cells were induced and incubated for 4 h at 37 °C in the presence of 0.5 mM p-azido-l-phenylalanine (pAzF) (pCNFRS) or 10 mM l-Boc-lysine (BocK) (MmPylRS). (B) SDS-PAGE gels illustrate proteins produced in Vmax X2 cells transformed with pET-S2TAGsfGFP and either pEVOL-CNF or pEVOL-mmPyl. L = lysate; S = supernatant; F = flow-through; W = wash; E = elution. (C) Intact protein mass spectra of 2TAG sfGFP variants purified from Vmax X2 cells coexpressing pCNFRS (top) or MmPylRS (bottom) in the presence of pAzF or BocK, respectively.