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. 2013 Jul 2;8(7):e68363. doi: 10.1371/journal.pone.0068363

Figure 5. Site-specific incorporation of pIPhe into GFP in a cell-free expression system.

Figure 5

(A) Western blot visualization of WT GFP and pIPhe-incorporated GFP Y39TAG. Co-translational incorporation of pIPhe was achieved by addition of M. jannaschii IPheRS (100 µg/mL), different types of suppressor tRNA (480 µg/mL), and pIPhe (1 mM) to the reaction medium. tRNA denotes synthetic MjtRNACUA, Opt – tRNACUA Opt. (B) Annotated MS/MS spectrum of the FSVSGEGEGDATY*GK peptide from GFP Y39IPhe. The mass shift between pIPhe and tyrosine was calculated to be 109.9 Da, corresponding to the observed “y” ions.