Fig. 2

Peptides utilized for the current study. a Peptide names and their sequences. Sequences with a glutamic acid residue marked in red represent the four isomeric species discussed in the text and shown in b. ABA 4-acetamidobenzoate, IBA 4-iso-butylamide benzoate, and SR 4-mercaptophenylacetic acid (MPAA). b Absolute configuration and backbone connectivity of the ligation site Glu residue in 2 and its isomers 2 ^D, 2γ, and 2 ^D γ. c Representative HPLC chromatogram (detected at 270 nm) obtained during a ligation reaction initiated by 200 μM E and 300 μM N. Note that schemes detailing the ligation reaction steps are given in Fig. 5 and Supplementary Fig. 13. The chromatogram allows identification of all the species forming the reaction network, including the products 2, 2 ^D, 2γ, and 2 ^D γ, the isomeric electrophile molecules (E ^D, Eγ, and E ^D γ), and the intermediate anhydride species (A and A ^D, Supplementary Fig. 1a). The electrophile and product isomers were all analyzed by LCMS, and their elution times were compared to those of the respective originally synthesized compounds (Supplementary Fig. 1b). A and A ^D peptides were identified by LCMS. Note that peptide N could not be well detected, since it lacks ABA labeling, and that two electrophile hydrolysis products, eluted at early times, are not shown (see SI Supplementary Methods)