Poster Presentation 8th Modern Solid Phase Peptide Synthesis & Its Applications Symposium 2022

Rapid one-pot iterative diselenide-selenoester ligations using a novel coumarin-based photolabile protecting group. (#115)

Lucas Kambanis 1 , Timothy S. Chisholm 1 , Sameer S. Kulkarni 1 , Richard J. Payne 1
  1. The University of Sydney, Camperdown, NEW SOUTH WALES, Australia

The development and application of chemoselective peptide ligation chemistry has transformed the field of chemical protein synthesis.1-3 We recently reported the development of a novel peptide ligation method called the diselenide–selenoester ligation (DSL) reaction that has been successfully used to synthesis a number of post-translationally modified protein targets.4, 5 Importantly, this method leverages the reactivity of selenoesters and of the selenol side chain of the 21st amino acid selenocysteine (Sec)  and leads to rapid ligation rates. . Importantly, DSL reactions usually proceed to completion within 10 minutes even at sterically hindered junctions and are also competent down at nanomolar fragment concentrations where the gold standard ligation method native chemical ligation (NCL) are inoperable.

We have recently developed an iterative one-pot peptide ligation strategy that capitalizes on the rapid and efficient nature of the diselenide-selenoester ligation reaction (DSL), together with photodeselenization chemistry to avoid intermediary purification steps.6 The development of a novel photolabile protecting group for the side chain of selenocysteine, namely the 7-diethylamino-3-methyl coumarin (DEAMC) moiety was crucial to this one-pot ligation strategy. Deprotection of the DEAMC group was shown to proceed smoothly within minutes under a mild, reagent-free manner using visible light (λ = 450 nm) without deleterious deselenization of selenocysteine residues, thus enabling a subsequent ligation reaction without purification. In this presentation we will also highlight the application of this DEAMC-protected selenocysteine in iterative DSL chemistry through the efficient one-pot assembly of trimeric and tetrameric fragments of the tumour-associated protein mucin-1, as well as an unmodified apolipoprotein CIII in just 2-4 hours with only a single purification step.

  1. Kent, S. B. H., Total chemical synthesis of proteins. Chem. Soc. Rev. 2009, 38 (2), 338-351.
  2. Bondalapati, S.; Jbara, M.; Brik, A., Expanding the chemical toolbox for the synthesis of large and uniquely modified proteins. Nat. Chem. 2016, 8 (5), 407-18.
  3. Kulkarni, S. S.; Sayers, J.; Premdjee, B.; Payne, R. J., Rapid and efficient protein synthesis through expansion of the native chemical ligation concept. Nat. Rev. Chem. 2018, 2 (4), 1-17.
  4. Mitchell, N. J.; Malins, L. R.; Liu, X.; Thompson, R. E.; Chan, B.; Radom, L.; Payne, R. J., Rapid additive-free selenocystine–selenoester peptide ligation. J. Am. Chem. Soc. 2015, 137 (44), 14011-14014.
  5. Kulkarni, S. S.; Watson, E. E.; Premdjee, B.; Conde-Frieboes, K. W.; Payne, R. J., Diselenide–selenoester ligation for chemical protein synthesis. Nat. Protoc. 2019, 14 (7), 2229-2257.
  6. Kambanis, L.; Chisholm, T. S.; Kulkarni, S. S.; Payne, R. J., Rapid one-pot iterative diselenide–selenoester ligation using a novel coumarin-based photolabile protecting group. Chem. Sci. 2021, 12 (29), 10014-10021.