Ixolaris: A Factor Xa heparin-binding exosite inhibitor

Robson Q. Monteiro, Alireza R. Rezaie, José M.C. Ribeiro, Ivo M.B. Francischetti

Research output: Contribution to journalArticlepeer-review

Abstract

Ixolaris is a two-Kunitz TFPI (tissue factor pathway inhibitor) from the tick salivary gland. In contrast with human TFPI, Ixolaris binds tightly to the zymogen FX (Factor X) and to dansyl-Glu-Gly-Arg-chloromethyl ketone-treated FXa (DEGR-FXa; active-site-blocked FXa), indicating that exosites are involved in the FX(a)-Ixolaris interaction. Here we provide evidence that Ixolaris binds specifically to the FXa HBE (heparin-binding exosite), since (i) it markedly decreases the inhibition of FXa by the antithrombin-heparin but not the antithrombin-pentasaccharide complex, (ii) it impairs FXa binding to Sepharose-immobilized heparin, and (iii) it allosterically modulates the catalytic activity of FXa for small chromogenic substrates (S-2765). By using a series of recombinant FXa mutants in which the HBE is mutated, we have identified the importance of amino acids involved in the enzyme-inhibitor interaction as being in the following order: Arg-93 ≫ Arg-165 ≥ Lys-169 > Lys-236 > Lys-96 > Arg-240 > Arg-125. Ixolaris at appropriate concentrations also inhibits thrombin formation in vitro by the assembled prothrombinase complex, a process that is critically dependent on the FXa HBE. Ixolaris is the first inhibitor characterized to date that binds specifically to the FXa HBE.

Original languageEnglish (US)
Pages (from-to)871-877
Number of pages7
JournalBiochemical Journal
Volume387
Issue number3
DOIs
StatePublished - May 1 2005
Externally publishedYes

Keywords

  • Blood feeding
  • Factor Xa
  • Heparin-binding exosite
  • Prothrombin
  • Tick saliva

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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