Glutathione could also be incorporated efficiently, either on Lysine (peptide 5e) or on the N terminus (peptide 5f). In aqueous media, alkenylation of peptide-EBXs occurred by simple addition of the thiol to generate S-VBX products 6b. These physiological conditions can also be used for protein functionalization. As proof of concept, His6-Cys-Ubiquitin was examined as substrate. A quantitative reaction was obtained, although in this case a 1.4:1 mixture of VBX 6b and thioalkyne 5g was observed. b Decarboxylative alkynylation of peptide-EBXs. Full experimental details are provided in Supplementary Information. a Only the S-alkynylation product was isolated, the HPLC UV-ratio of S-alkynylation 5 and S-alkenylation product 6 is indicated in the parenthesis. The yield was determined based on LC-MS. c Ratio of S-VBX:S-alkynylation = 1.4:1. d The dr was determined by C-NMR on the isolated product. e The dr was not determined.