Peptide dithiodiethanol esters for in situ generation of thioesters for use in native ligation |
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Authors: | A. Pernille Tofteng Knud J. Jensen |
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Affiliation: | a Royal Veterinary and Agricultural University, Thorvaldsensvej 40, DK-1871 Frederiksberg, Denmark b Novo Nordisk, Novo Alle 6B2.54, DK-2880 Bagsvaerd, Denmark |
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Abstract: | A new approach is described for the general Fmoc-based solid-phase synthesis of C-terminal peptide (thio)esters. One hydroxy group of 2,2-dithiodiethanol (used in large excess) was anchored on trityl resin, and the remaining hydroxy group was loaded with the first amino acid. Standard chain elongation and TFA-based peptide release yielded peptide C-terminal dithiodiethanol esters in good purities. Under standard conditions of native chemical ligation (excess thiol, neutral pH), the dithiodiethanol function is presumably reduced and rearranged (or equilibrated) to the thioester via a 5-membered intermediate. The resulting thioesters are shown to undergo native chemical ligation with N-terminal cysteine peptides. Notably, hydrolysis of the reduced ester is a major competing reaction, especially in the presence of 6 M guanidinium chloride, which is often required for solubilization of large peptide fragments. |
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Keywords: | Peptide thioester Native chemical ligation Dithiodiethanol linker Hydrolysis |
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