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On-Demand Detachment of Maleimide Derivatives on Cysteine to Facilitate (Semi)Synthesis of Challenging Proteins

submitted on 16.07.2020, 09:17 and posted on 16.07.2020, 12:30 by Ganga B. Vamisetti, satish gandhesiri, Prasad Sulkshane, Guy Mann, Michael H. Glickman, ashraf brik

The maleimide group is a widely used reagent for bio-conjugation of peptides, proteins and oligonucleotides employing Michael addition and Diels-Alder cycloaddition reactions. However, the utility of this functionality in chemical synthesis of peptides and proteins remains unexplored. We report, for the first time that PdII complexes can mediate the efficient removal of various maleimide derivatives in aqueous conditions. Maleimide removal by PdII was applied for the synthesis of two ubiquitin activity-based probes (Ub-ABPs) employing solid phase chemical ligation (SPCL). SPCL was achieved through a sequential three segments ligation on a polymer support via a maleimide anchor. The obtained probes successfully formed the expected covalent complexes with deubiquitinating enzymes (DUBs) USP2 and USP7, highlighting the use of our new method for efficient preparation of unique synthetic proteins. Importantly, we demonstrate the advantages of our newly developed method for the protection and deprotection of native cysteine with a maleimide group in a peptide fragment derived from thioredoxin-1 (Trx-1) obtained via intein based expression to enable ligation/desulfurization and subsequent disulfide bond formation in a one-pot process.


This project has received funding from the European Research Council (ERC) under the European Union’s Horizon 2020 research and innovation programme (grant agreement no. [831783]). A. B. holds The Jordan and Irene Tark Academic Chair. M. H. G. acknowledges NSF-BSF grant MCB1818280.


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Technion-Israel Institute of Technology



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Declaration of Conflict of Interest

No conflict of interest to declare