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PLoS Negl Trop Dis. 2014 Jun 12;8(6):e2947. doi: 10.1371/journal.pntd.0002947. eCollection 2014 Jun.

Simplagrin, a platelet aggregation inhibitor from Simulium nigrimanum salivary glands specifically binds to the Von Willebrand factor receptor in collagen and inhibits carotid thrombus formation in vivo.

Author information

1
Laboratory of Malaria and Vector Research, National Institute of Allergy and Infectious Diseases (NIAID), National Institutes of Health (NIH), Rockville, Maryland, United States of America.
2
Physical and Biochemistry Section, National Institute of Diabetes and Digestive and Kidney Diseases, NIH, Bethesda, Maryland, United States of America.
3
Animal Surgery and Resources Core, National Heart Lung and Blood Institute, NIH, Bethesda, Maryland, United States of America.
4
Laboratory of Malaria Immunology and Vaccinology, NIAID, NIH, Bethesda, Maryland, United States of America.
5
Centro de Pesquisas Aggeu Magalhães (CPqAM/FIOCRUZ) and Laboratório de Imunopatologia Keizo Asami. Universidade Federal de Pernambuco, Recife, Pernambuco, Brazil.

Abstract

BACKGROUND:

Among the several challenges faced by bloodsucking arthropods, the vertebrate hemostatic response against blood loss represents an important barrier to efficient blood feeding. Here we report the first inhibitor of collagen-induced platelet aggregation derived from the salivary glands of a black fly (Simulium nigrimanum), named Simplagrin.

METHODS AND FINDINGS:

Simplagrin was expressed in mammalian cells and purified by affinity-and size-exclusion chromatography. Light-scattering studies showed that Simplagrin has an elongated monomeric form with a hydrodynamic radius of 5.6 nm. Simplagrin binds to collagen (type I-VI) with high affinity (2-15 nM), and this interaction does not involve any significant conformational change as determined by circular dichroism spectroscopy. Simplagrin-collagen interaction is both entropically and enthalpically driven with a large negative ΔG, indicating that this interaction is favorable and occurs spontaneously. Simplagrin specifically inhibits von Willebrand factor interaction with collagen type III and completely blocks platelet adhesion to collagen under flow conditions at high shear rates; however, Simplagrin failed to block glycoprotein VI and Iα2β1 interaction to collagen. Simplagrin binds to RGQOGVMGF peptide with an affinity (K(D) 11 nM) similar to that of Simplagrin for collagen. Furthermore, Simplagrin prevents laser-induced carotid thrombus formation in vivo without significant bleeding in mice and could be useful as an antithrombotic agent in thrombosis related disease.

CONCLUSION:

Our results support the orthology of the Aegyptin clade in bloodsucking Nematocera and the hypothesis of a faster evolutionary rate of salivary function of proteins from blood feeding arthropods.

PMID:
24921659
PMCID:
PMC4055580
DOI:
10.1371/journal.pntd.0002947
[Indexed for MEDLINE]
Free PMC Article
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