A Novel Tetrameric Venom Protein, Agglucetin from Agkistrodon acutus , Acts as a Glycoprotein Ib Agonist

Wen-Jeng Wang; Tur-Fu Huang

doi:10.1055/s-0037-1616536

Thrombosis and Haemostasis, Table of Contents

Thromb Haemost 2001; 86(04): 1077-1086
DOI: 10.1055/s-0037-1616536

Special Article

Schattauer GmbH

A Novel Tetrameric Venom Protein, Agglucetin from Agkistrodon acutus , Acts as a Glycoprotein Ib Agonist

Wen-Jeng Wang

¹Department of Pharmacology, College of Medicine, National Taiwan University, Taipei, Taiwan

,

Tur-Fu Huang

¹Department of Pharmacology, College of Medicine, National Taiwan University, Taipei, Taiwan

› Author Affiliations

Abstract

Summary

A novel platelet agglutination inducer, agglucetin, was purified from the Formosan Agkistrodon acutus snake venom. It migrated as a single band with an apparent molecular mass of 58.8 kDa and two distinct bands of 16.2/14.5 kDa under non-reducing and reducing conditions by SDS-PAGE, respectively. Further confirmed by FPLC, electrospray ionization mass spectrometry and 2D-PAGE, native agglucetin exists as a tetramer composed of disulfide-linked α1, α2, β1 and β2 subunits. Partial N-terminal sequence of agglucetin subunit showed a high degree of homology to those of C-type lectin-like glycoprotein (GP) Ib binding proteins. Functional studies showed that agglucetin, in the absence of von Willebrand factor (vWF), dose-dependently induced platelet agglutination and caused a negligible elevation of intracellular Ca⁺² mobilization and thromboxane B₂ formation in human platelet suspensions. Anti-GP Ib monoclonal antibodies (mAbs), AP1 or LJ-Ib1, specifically inhibited agglucetin-induced platelet agglutination in a dose-dependent manner. However, EDTA, arietin (a long chain RGD-containing disintegrin), 7E3 (an anti-GP IIb/IIIa mAb), heparin, hirudin, PGE₁, or indomethacin exhibited no inhibitory effect on agglucetin-induced platelet agglutination. Furthermore, flow cytometric analysis revealed that FITC-agglucetin dose-dependently bound to human formalin-fixed platelets in a saturable manner, and its binding was specifically blocked by anti-GP Ib mAb. It is concluded that agglucetin, acts specifically on an epitope of platelet membrane GP Ib overlapping with that of AP1, causing platelet agglutination in a Ca⁺²- and GP IIb/IIIa-independent manner.

Keywords

C-type lectin-like venom proteins - Agkistrodon acutus - platelet agglutination - von Willebrand factor - glycoprotein Ib agonist

Full Text

References

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