Potent Trivalent Inhibitors of Thrombin through Hybridization of Salivary Sulfopeptides from Hematophagous Arthropods

S.M. Agten, E.E. Watson, J. Ripoll-Rozada, L.J. Dowman, M.C.L. Wu, I. Alwis, S.P. Jackson, P.J.B. Pereira*, R.J. Payne*

*Corresponding author for this work

Research output: Contribution to journalArticleAcademicpeer-review

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Blood feeding arthropods, such as leeches, ticks, flies and mosquitoes, provide a privileged source of peptidic anticoagulant molecules. These primarily operate through inhibition of the central coagulation protease thrombin by binding to the active site and either exosite I or exosite II. Herein, we describe the rational design of a novel class of trivalent thrombin inhibitors that simultaneously block both exosites as well as the active site. These engineered hybrids were synthesized using tandem diselenide-selenoester ligation (DSL) and native chemical ligation (NCL) reactions in one-pot. The most potent trivalent inhibitors possessed femtomolar inhibition constants against alpha-thrombin and were selective over related coagulation proteases. A lead hybrid inhibitor possessed potent anticoagulant activity, blockade of both thrombin generation and platelet aggregation in vitro and efficacy in a murine thrombosis model at 1 mg kg(-1). The rational engineering approach described here lays the foundation for the development of potent and selective inhibitors for a range of other enzymatic targets that possess multiple sites for the disruption of protein-protein interactions, in addition to an active site.
Original languageEnglish
Pages (from-to)5348-5356
Number of pages9
JournalAngewandte Chemie-International Edition
Issue number10
Publication statusPublished - 1 Mar 2021


  • anticoagulant
  • peptide engineering
  • peptide ligation
  • protein synthesis
  • thrombin
  • Thrombin

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