Eladio Flores Sanchez1, Michael Richardson2, Luiza Helena Gremski3, Silvio Sanches Veiga3, Armando Yarleque4, Stephan Niland5, Augusto Martins Lima5, Maria Inácia Estevao-Costa6, Johannes Andreas Eble5. 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.
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.
Authors: Luciana S Oliveira; Maria Inácia Estevão-Costa; Valéria G Alvarenga; Dan E Vivas-Ruiz; Armando Yarleque; Augusto Martins Lima; Ana Cavaco; Johannes A Eble; Eladio F Sanchez Journal: Molecules Date: 2019-09-26 Impact factor: 4.411
Authors: Lorenzo Seneci; Christina N Zdenek; Abhinandan Chowdhury; Caroline F B Rodrigues; Edgar Neri-Castro; Melisa Bénard-Valle; Alejandro Alagón; Bryan G Fry Journal: Front Immunol Date: 2021-03-11 Impact factor: 7.561
Authors: Eladio Flores Sanchez; Michael Richardson; Luiza Helena Gremski; Silvio Sanches Veiga; Armando Yarleque; Stephan Niland; Augusto Martins Lima; Maria Inácia Estevao-Costa; Johannes Andreas Eble Journal: Data Brief Date: 2016-04-30