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Simplagrin, a salivary collagen-induced platelet aggregation inhibitor from blackfly Simulim nigrimanum displays an unusual elongated form and inhibits carotid thrombus formation in vivo.

Wednesday, November 06, 2013 — Poster Session I

4:00 p.m. – 6:00 p.m.

FAES Academic Center (Upper-Level Terrace)

NIAID

MOLBIO-7

* FARE Award Winner

Authors

  • AC Chagas
  • P McPhie
  • H Sun
  • D Narum
  • FA Tokomasu
  • LC Alves
  • F Brayner
  • JM Ribeiro
  • E Calvo

Abstract

Little is known about the protein composition and function of blackflies saliva. Previous blackfly sialotranscriptomes revealed proteins distantly related to Aegyptin, a mosquito platelet aggregation inhibitor, supporting the idea of a possibly common origin of hematophagy in Diptera. Recombinant Simplagrin (Simulium platelet aggregation inhibitor) was expressed in HEK293 cells and purified by affinity and size exclusion chromatography. SPR identified collagen as the molecular partner of Simplagrin (affinity of 10-15 nM). Thermodynamic analysis of Simplagrin-Collagen shows that this interaction is both entropically (DH -20kJ/mol) and enthalpically (75 J/K*mol) driven with a DG of -40kJ/mol. Simplagrin specifically inhibits vWF interaction with collagen type III under static conditions and completely blocks platelet adhesion to collagen under flow conditions at high shear rates. No binding to GPVI or Ia2B1 on collagen was found. Simplagrin prevents laser-induced carotid thrombus formation in vivo, without significant bleeding in mice suggesting that it could be used as an antithrombotic drug in thrombosis-related disease. Simplagrin is the first collagen-binding protein characterized in the salivary gland of blackflies and support the hypothesis on the convergence of salivary function of distantly related proteins of blood-feeding arthropods in the face of the host’s hemostatic and immune defense.

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