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Biochim Biophys Acta. 2016 Mar;1860(3):542-56. doi: 10.1016/j.bbagen.2015.12.021. Epub 2015 Dec 23.

A novel fibrinolytic metalloproteinase, barnettlysin-I from Bothrops barnetti (Barnett´s pitviper) snake venom with anti-platelet properties.

Author information

1
Research and Development Center, Ezequiel Dias Foundation, 30510-010, Belo Horizonte, MG, Brazil; Faculty of Biological Sciences, Nacional University of San Marcos, Lima-Peru. Electronic address: eladiooswaldo@gmail.com.
2
Research and Development Center, Ezequiel Dias Foundation, 30510-010, Belo Horizonte, MG, Brazil.
3
Department of Cell Biology, Federal University of Parana, Brazil.
4
Faculty of Biological Sciences, Nacional University of San Marcos, Lima-Peru.
5
Institute for Physiological Chemistry and Pathobiochemistry, University of Münster, Germany.
6
Research and Development Center, Ezequiel Dias Foundation, 30510-010, Belo Horizonte, MG, Brazil; Institute for Physiological Chemistry and Pathobiochemistry, University of Münster, Germany.

Abstract

BACKGROUND:

Viperid snake venoms contain active components that interfere with hemostasis. We report a new P-I class snake venom metalloproteinase (SVMP), barnettlysin-I (Bar-I), isolated from the venom of Bothrops barnetti and evaluated its fibrinolytic and antithrombotic potential.

METHODS:

Bar-I was purified using a combination of molecular exclusion and cation-exchange chromatographies. We describe some biochemical features of Bar-I associated with its effects on hemostasis and platelet function.

RESULTS:

Bar-I is a 23.386 kDa single-chain polypeptide with pI of 6.7. Its sequence (202 residues) shows high homology to other members of the SVMPs. The enzymatic activity on dimethylcasein (DMC) is inhibited by metalloproteinase inhibitors e.g. EDTA, and by α2-macroglobulin. Bar-I degrades fibrin and fibrinogen dose- and time-dependently by cleaving their α-chains. Furthermore, it hydrolyses plasma fibronectin but not laminin nor collagen type I. In vitro Bar-I dissolves fibrin clots made either from purified fibrinogen or from whole blood. In contrast to many other P-I SVMPs, Bar-I is devoid of hemorrhagic activity. Also, Bar-I dose- and time-dependently inhibits aggregation of washed human platelets induced by vWF plus ristocetin and collagen (IC50=1.3 and 3.2 μM, respectively), presumably Bar-I cleaves both vWF and GPIb. Thus, it effectively inhibits vWF-induced platelet aggregation. Moreover, this proteinase cleaves the collagen-binding α2-A domain (160 kDa) of α2β1-integrin. This explains why it additionally inhibits collagen-induced platelet activation.

CONCLUSION:

A non-hemorrhagic but fibrinolytic metalloproteinase dissolves fibrin clots in vitro and impairs platelet function.

GENERAL SIGNIFICANCE:

This study provides new opportunities for drug development of a fibrinolytic agent with antithrombotic effect.

KEYWORDS:

Antithrombotic; Direct acting fibrinolytic enzyme; Integrins; Metalloproteinase; Snake venoms; von willebrand factor

PMID:
26723171
DOI:
10.1016/j.bbagen.2015.12.021
[Indexed for MEDLINE]
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