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. 2022 Mar 16;11:e76941. doi: 10.7554/eLife.76941

Figure 5. Characterization of amino acid incorporation by quadruplet tRNAs (qtRNAs).

Figure 5.

(A) We characterized the amino acid incorporated during translation by co-expressing a qtRNA and an sfGFP-151-quad transcript, purifying the resulting GFP, and analyzing the occupancy of residue 151 using mass spectrometry (Supplementary file 3). (B) Results of applying this pipeline to at least one qtRNA based on each of the 20 canonical scaffolds (Materials and methods – ‘Quantification of qtRNA charging using mass spectrometry’). Charging of (*) qtRNAArgTAGA and qtRNATyrTAGA has been previously reported (DeBenedictis et al., 2021). Data represents the mean of one biological replicate. For some qtRNAs, yield was too low to allow for charging characterization even when purified at 1 L scale (Supplementary file 2). Raw spectra have been deposited in the PRIDE database (Perez-Riverol et al., 2022), dataset identifier PXD031925 and 10.6019/PXD031925.