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BioMed Research International
Volume 2014 (2014), Article ID 595186, 13 pages
http://dx.doi.org/10.1155/2014/595186
Research Article

Isolation and Biochemical Characterization of a New Thrombin-Like Serine Protease from Bothrops pirajai Snake Venom

1Centro de Estudos de Biomoléculas Aplicadas à Saúde, CEBio, Fundação Oswaldo Cruz, Fiocruz Rondônia e Departamento de Medicina, Universidade Federal de Rondônia, UNIR, Rua da Beira 7176, Bairro Lagoa, 76812-245 Porto Velho, RO, Brazil
2Departmento de Biologia Celular e Molecular, Instituto de Biologia, Universidade Federal Fluminense, 24210-130 Niteroi, RJ, Brazil
3Centro de Ciências Naturais e Humanas, Universidade Federal do ABC, 09210-170 Santo André, SP, Brazil
4Escola de Artes, Ciências e Humanidades, USP, 03828-000 São Paulo, SP, Brazil
5Universidade Federal de São João Del Rei, UFSJ, Campus Alto Paraopeba, 36420-000 Ouro Branco, MG, Brazil
6Institute for Research in Biomedicine (IRB Barcelona), 08028 Barcelona, Spain
7CIBER-BBN, Barcelona Science Park, 08028 Barcelona, Spain
8Proteomic Platform, Barcelona Science Park, 08028 Barcelona, Spain
9Department of Organic Chemistry, University of Barcelona, 08028 Barcelona, Spain
10School of Chemistry, University of KwaZulu Natal, Durban 4001, South Africa

Received 9 July 2013; Revised 20 September 2013; Accepted 1 December 2013; Published 26 February 2014

Academic Editor: Edward G. Rowan

Copyright © 2014 Kayena D. Zaqueo et al. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

Abstract

This paper presents a novel serine protease (SP) isolated from Bothrops pirajai, a venomous snake found solely in Brazil that belongs to the Viperidae family. The identified SP, named BpirSP-39, was isolated by three chromatographic steps (size exclusion, bioaffinity, and reverse phase chromatographies). The molecular mass of BpirSP-39 was estimated by SDS-PAGE and confirmed by mass spectrometry (39,408.32 Da). The protein was able to form fibrin networks, which was not observed in the presence of serine protease inhibitors, such as phenylmethylsulfonyl fluoride (PMSF). Furthermore, BpirSP-39 presented considerable thermal stability and was apparently able to activate factor XIII of the blood coagulation cascade, unlike most serine proteases. BpirSP-39 was capable of hydrolyzing different chromogenic substrates tested (S-2222, S-2302, and S-2238) while Cu2+ significantly diminished BspirSP-39 activity on the three tested substrates. The enzyme promoted platelet aggregation and also exhibited fibrinogenolytic, fibrinolytic, gelatinolytic, and amidolytic activities. The multiple alignment showed high sequence similarity to other thrombin-like enzymes from snake venoms. These results allow us to conclude that a new SP was isolated from Bothrops pirajai snake venom.