Molecular identification and activity upon chromogenic substrates of a venombin A from Bothrops atrox Peruvian snake venom

Authors

  • Gustavo A. Sandoval Laboratorio de Biología Mole- cular, Facultad de Ciencias Bio- lógicas, Universidad Nacional Mayor de San Marcos. Ciudad Universitaria, Av. Venezuela Cdra. 34 s/n. Apartado 110058, Lima 11, Perú.
  • Fanny Lazo Laboratorio de Biología Mole- cular, Facultad de Ciencias Bio- lógicas, Universidad Nacional Mayor de San Marcos. Ciudad Universitaria, Av. Venezuela Cdra. 34 s/n. Apartado 110058, Lima 11, Perú.
  • Edith Rodriguez Laboratorio de Biología Mole- cular, Facultad de Ciencias Bio- lógicas, Universidad Nacional Mayor de San Marcos. Ciudad Universitaria, Av. Venezuela Cdra. 34 s/n. Apartado 110058, Lima 11, Perú.
  • Armando Yarlequé Laboratorio de Biología Mole- cular, Facultad de Ciencias Bio- lógicas, Universidad Nacional Mayor de San Marcos. Ciudad Universitaria, Av. Venezuela Cdra. 34 s/n. Apartado 110058, Lima 11, Perú.
  • Russolina B. Zingali Laboratorio de Hemostasia y Venenos. Departamento de Bioquí- mica Médica. Universidad Federal de Rio de Janeiro. Rio de Janeiro – Brazil.

DOI:

https://doi.org/10.15381/rpb.v17i3.12

Keywords:

Bothrops atrox, Thrombin-like enzyme MALDI-TOF, chromogenic substrates.

Abstract

n this work, the thrombin-like enzyme (TLE) from Bothrops atrox has been identified by mass spectrometry and its enzymatic activity evaluated upon several synthetic substrates. The enzyme was purified to homogeneity using three chromatography steps on Sephadex G-75, CM-Sephadex C-50 and Agarose-PAB. Also, molecular weight by PAGE-SDS was determined. For molecular identification of this enzyme, mass spectrometry-based peptide mass fingerprinting was used and later in silico analysis. Enzymatic activities were determined using bovine fibrinogen, BApNA and also upon specific chromogenic substrates such as S-2238, S-2251 y S-2266. As a result of these biochemical and structural procedures, we obtained a TLE from B. atrox venom with a molecular weight of 29,6 kDa. Mass spectrometry analysis of obtained peptides, allow us to identify this enzyme as a venombin A, showing a 75% sequence homology. After recording enzymatic activity, this TLE showed coagulant activity on bovine fibrinogen and upon BApNA, S-2238 y S-2266, being unable to hydrolyze S-2251 substrate. Using this combination of structural and functional approaches, we have identified the main component of B. atrox venom related to its coagulant activity, as well as a detailed evaluation of its enzymatic activity upon several substrates

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Published

12/31/2010

Issue

Vol. 17 No. 3 (2010)

Section

Articles

License

Copyright (c) 2010 Gustavo A. Sandoval, Fanny Lazo, Edith Rodriguez, Armando Yarlequé, Russolina B. Zingali

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How to Cite

Sandoval, Gustavo A., Fanny Lazo, Edith Rodriguez, Armando Yarlequé, and Russolina B. Zingali. 2010. “Molecular Identification and Activity Upon Chromogenic Substrates of a Venombin A from Bothrops Atrox Peruvian Snake Venom”. Revista Peruana De Biología 17 (3): 365-70. https://doi.org/10.15381/rpb.v17i3.12.