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Molecular Characterization of Lys49 and Asp49 Phospholipases A2 from Snake Venom and Their Antiviral Activities against Dengue virus
Laboratory of Biotechnology and Health, Research and Development Center, Ezequiel Dias Foundation, Belo Horizonte 30510-010, MG, Brazil
Laboratory of Biochemistry of Proteins from Animal Venoms, Research and Development Center, Ezequiel Dias Foundation, Belo Horizonte 30510-010, MG, Brazil
Center for Molecular Medicine, Dept. Vascular Matrix Biology, Frankfurt University Hospital, Frankfurt am Main 60590, Germany
Department of Cellular and Molecular Biology, Federal University Fluminense, RJ 24220-008, Brazil
* Author to whom correspondence should be addressed.
Received: 2 August 2013; in revised form: 11 September 2013 / Accepted: 26 September 2013 / Published: 15 October 2013
Abstract: We report the detailed molecular characterization of two PLA2s, Lys49 and Asp49 isolated from Bothrops leucurus venom, and examined their effects against Dengue virus (DENV). The Bl-PLA2s, named BlK-PLA2 and BlD-PLA2, are composed of 121 and 122 amino acids determined by automated sequencing of the native proteins and peptides produced by digestion with trypsin. They contain fourteen cysteines with pIs of 9.05 and 8.18 for BlK- and BlD-PLA2s, and show a high degree of sequence similarity to homologous snake venom PLA2s, but may display different biological effects. Molecular masses of 13,689.220 (Lys49) and 13,978.386 (Asp49) were determined by mass spectrometry. DENV causes a prevalent arboviral disease in humans, and no clinically approved antiviral therapy is currently available to treat DENV infections. The maximum non-toxic concentration of the proteins to LLC-MK2 cells determined by MTT assay was 40 µg/mL for Bl-PLA2s (pool) and 20 µg/mL for each isoform. Antiviral effects of Bl-PLA2s were assessed by quantitative Real-Time PCR. Bl-PLA2s were able to reduce DENV-1, DENV-2, and DENV-3 serotypes in LLC-MK2 cells infection. Our data provide further insight into the structural properties and their antiviral activity against DENV, opening up possibilities for biotechnological applications of these Bl-PLA2s as tools of research.
Keywords: svPLA2s; Bothrops leucurus; Dengue virus; antiviral effect; Real-Time PCR
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MDPI and ACS Style
Cecilio, A.B.; Caldas, S.; Oliveira, R.A.D.; Santos, A.S.B.; Richardson, M.; Naumann, G.B.; Schneider, F.S.; Alvarenga, V.G.; Estevão-Costa, M.I.; Fuly, A.L.; Eble, J.A.; Sanchez, E.F. Molecular Characterization of Lys49 and Asp49 Phospholipases A2 from Snake Venom and Their Antiviral Activities against Dengue virus. Toxins 2013, 5, 1780-1798.
Cecilio AB, Caldas S, Oliveira RAD, Santos ASB, Richardson M, Naumann GB, Schneider FS, Alvarenga VG, Estevão-Costa MI, Fuly AL, Eble JA, Sanchez EF. Molecular Characterization of Lys49 and Asp49 Phospholipases A2 from Snake Venom and Their Antiviral Activities against Dengue virus. Toxins. 2013; 5(10):1780-1798.
Cecilio, Alzira B.; Caldas, Sergio; Oliveira, Raiana A.D.; Santos, Arthur S.B.; Richardson, Michael; Naumann, Gustavo B.; Schneider, Francisco S.; Alvarenga, Valeria G.; Estevão-Costa, Maria I.; Fuly, Andre L.; Eble, Johannes A.; Sanchez, Eladio F. 2013. "Molecular Characterization of Lys49 and Asp49 Phospholipases A2 from Snake Venom and Their Antiviral Activities against Dengue virus." Toxins 5, no. 10: 1780-1798.