HETEROLOGOUS EXPRESSION AND PURIFICATION OF RECOMBINANT CROTOXIN B, THE PHOSPHOLIPASE A2 SUBUNIT OF CROTOXIN

Authors

  • Francisc-Andrei BODA Department of Fundamental Pharmaceutical Sciences (DSFF), Faculty of Pharmacy, University of Medicine and Pharmacy, Târgu Mureș, Romania. Corresponding author: andrei.gaz@umftgm.ro. https://orcid.org/0000-0003-3777-3712
  • Pál SALAMON Department of Bioengineering, Faculty of Economics, Socio-Human Sciences and Engineering, Sapientia Hungarian University of Transylvania, Miercurea Ciuc, Romania. Corresponding author: andrei.gaz@umftgm.ro. https://orcid.org/0000-0002-6803-1212
  • Csongor ORBÁN Department of Bioengineering, Faculty of Economics, Socio-Human Sciences and Engineering, Sapientia Hungarian University of Transylvania, Miercurea Ciuc, Romania. Email: orbancsongor@uni.sapientia.ro. https://orcid.org/0000-0002-7509-748X
  • Lavinia-Anda BERȚA Department of Fundamental Pharmaceutical Sciences (DSFF), Faculty of Pharmacy, University of Medicine and Pharmacy, Târgu Mureș, Romania. Corresponding author: andrei.gaz@umftgm.ro. https://orcid.org/0000-0002-2563-8946
  • Augustin CURTICĂPEAN Department of Fundamental Pharmaceutical Sciences (DSFF), Faculty of Pharmacy, University of Medicine and Pharmacy, Târgu Mureș, Romania. Email: augustin.curticapean@umftgm.ro. https://orcid.org/0000-0002-6770-0429
  • Șerban-Andrei GÂZ Department of Fundamental Pharmaceutical Sciences (DSFF), Faculty of Pharmacy, University of Medicine and Pharmacy, Târgu Mureș, Romania. Email: andrei.gaz@umftgm.ro. https://orcid.org/0000-0002-2486-3391
  • Maria DOGARU Department of Fundamental Pharmaceutical Sciences (DSFF), Faculty of Pharmacy, University of Medicine and Pharmacy, Târgu Mureș, Romania. Email: maria.dogaru@umftgm.ro.

DOI:

https://doi.org/10.24193/subbchem.2018.1.01

Keywords:

Crotoxin, neurotoxin, phospholipase A2, heterologous expression

Abstract

Crotoxin is a heterodimeric β-neurotoxin isolated from the venom of Crotalus durissus terrificus, consisting of two, non-covalently bound subunits, crotapotin (CA) and crotoxin B (CB). Both subunits present four different isoforms, consequently there are up to 16 different crotoxin complexes, each with different activity levels. The biological activities usually associated with crotoxin include neurotoxicity, myotoxicity and cardiotoxicity, however several other important biochemical and pharmacological effects of crotoxin have been observed, such as the antibacterial, antiviral, anti-inflammatory, antitumoral and analgesic activity. In this study we present the production of recombinant crotoxin B (isoform C). Expression of the protein was carried out using E. coli RosettaTM (DE3)pLysS strain, and the obtained protein was separated and purified using Ni2+-affinity chromatography. To the best of our knowledge the developed method represents the first reported method for obtaining the crotoxin B (isoform C) through heterologous expression using an efficient and cost-effective procedure.

References

G. Faure, D. Porowinska, F. Saul “Toxins and Drug Discovery”, Springer Nature, Dordrecht, 2017, Part I, Chapter 1, p. 3.

S.C. Sampaio, S. Hyslop, M.R.M. Fontes, J. Prado-Franceschi, V.O. Zambelli, A.J. Magro, P. Brigatte, V.P. Gutierrez, Y. Cury, Toxicon, 2010, 55, 1045.

C.A.H. Fernandes, W.M. Pazin, M.R.M. Fontes et al., Scientific Reports, 2017, 7, 43885.

L.A. Ponce-Soto, B. Lomonte, L. Rodriques-Simioni, J.C. Novello, S. Marangoni, The Protein Journal, 2007, 26, 221.

D.P. Marchi-Salvador, L.C. Corrêa, A.J. Magro, C.Z. Oliveira, A.M. Soares, M.R. Fontes, Proteins, 2008, 72, 883.

W. Cavalcante, E. Santos-Diz-Filho, M. Gallacci et al., Toxicology Letters, 2007, 172, S226.

O.V. Brazil, M.D. Fontana, N.F. Heluany, Journal of Natural Toxins, 2000, 9, 33.

C. Montecucco, J.M. Gutierrez, B. Lomonte, Cellular and Molecular Life Sciences, 2008, 65, 2897.

T.F. Salvini, A.C. Amaral, E.H. Miyabara, J.A.O. Turri, P.M. Danella, H.S. Selistre de Araujo, Toxicon, 2001, 39, 1141.

J.M. Gutierrez, L.A. Ponce-Soto, S. Marangoni, B. Lomonte, Toxicon, 2008, 51, 80.

E.H. Myiabara, I.L. Baptista, B. Lomonte, H.S. Selistre-de-Araujo, J.M. Gutierrez, A.S. Moriscot, Comparative Biochemistry and Physiology, Part C, 2006, 143, 284.

M. Hernandez, H. Scannone, H.J. Finol, M.E. Pineda, I. Fernandez, A.M. Vargas, M.E. Giron, I. Aguilar, A. Rodriguez-Acosta, Experimental and Toxicologic Pathology, 2007, 59, 129.

P. Zhang, A.S. Lader, M.A. Etcheverry, H.F. Cantiello. Toxicon, 2010, 55, 1236.

D.G. Oliveira, M.H. Toyama, J.C. Novello, L.O.S. Beriam, S. Marangoni, Journal of Protein Chemistry, 2002, 21, 161.

R. Perumal Samy, A. Pachiappan, P. Gopalakrishnakone, M.M. Thwin, Y.E. Hian, V.T.K. Chow, H. Bow, J.T. Weng, BMC Infectious Diseases, 2006, 6, 100.

R. Perumal Samy, P. Gopalakrishnakone, M.M. Thwin, T.K.V. Chow, H. Bow, E.H. Yap, T.W.J. Thong, Journal of Applied Microbiology, 2007, 102, 650.

A.M. Soares, A.C. Mancin, A.L. Cecchini, E.C. Arantes, S.C. Franca, J.M. Gutierrez, J.R. Giglio, International Journal of Biochemistry and Cell Biology, 2001, 33, 877.

V.D.M. Muller, R.R. Russo, A.C.O. Cintra, M.A. Sartim, R.M. Alves-Paiva, L.T.M. Figueiredo, S.V. Sampaio, V.H. Aquino, Toxicon, 2012, 59, 507.

V.D. Muller, R.O. Soares, N.N. dos Santos-Junior, A.C. Trabuco, A.C. Cintra, L.T. Figueiredo, A. Caliri, S.V. Sampaio, V.H. Aquino, PLoS ONE, 2014, 9, e112351, 1.

S.C. Sampaio, A.C. Rangel-Santos, C.M. Peres, R. Curi, Y. Curi, Toxicon, 2005, 45, 671.

D.F. Cardoso, M. Lopes-Ferreira, E.L. Faquim-Mauro, M.S. Macedo, S.H.P. Farsky, Mediators of Inflammation, 2001, 10, 125.

V.O. Zambelli, S.C. Sampaio, L.S. Sudo-Hayashi, K. Greco, L.R.G. Britto, A.S. Alves, B.C. Zychar, L.R.C. Goncalves, D.D. Spaddaci-Morena, R. Otton, M.S. Della-Casa, R. Curi, Y. Cury, Toxicon, 2008, 51, 1357.

J. Wang, X. Qin, Z. Zhang, M. Chen, Y. Wang, B. Gao, Molecular Medicine Report, 2014, 10, 3009.

L.A. Calderon, J.C. Sobrinho, A.M. Soares et al., BioMed Research Internationl, 2014, 2014, 203639, 1.

J.E. Cura, D.P. Blanzaco, C. Brisson, M.A. Cura, R. Cabrol, L. Larrateguy, C. Mendez, J.C. Sechi, J.S. Silveira, E. Theiller, A.R. de Roodt, J.C. Vidal, Clinical Cancer Research, 2002, 8, 1033.

C. Yan, Y. Yang, Z. Qin, Z. Gu, P. Reid, Z. Liang, Acta Pharmacologica Sinica, 2007, 28, 540.

F.S. Nogueira-Neto, R.L. Amorim, P. Brigatte, G. Picolo, W.A. Ferreira Jr., V.P. Gutierrez, I.M. Conceicao, M.S. Della-Casa, R.K. Takahira, J.L.M. Nicoletti, Y. Cury, Pharmacology, Biochemistry and Behavior, 2008, 91, 252.

Q. Zhu, D.C. Wu, X.P. Zhou, S. Gong, B.C. Cheng, Z.H. Qin, P.F. Reid, Q.Z. Yin, X.H. Jiang, Toxicon, 2008, 51, 102.

S. Wang, Y. Zhang, H. Liu, Y. He, J. Yan, Z. Wu, Y. Ding, Applied Microbiology and Biotechnology, 2012, 96, 939.

F. Bugli, R. Graffeo, F.P. Sterbini, R. Torelli, L. Masucci, M. Sali, A. Grasso, S. Rufini, E. Ricci, G. Fadda, M. Pescatori, Toxicon, 2008, 51, 547.

P. Salamon, I. Miklossy, B. Albert, M. Korodi, K. Nagy, I. Bakos, Sz. Lanyi, Cs. Orban, Studia UBB Chemia, 2017, 62, 333.

U.K. Laemmli, Nature, 1970, 227, 680.

STUDIA UBB CHEMIA, Volume 63 (LXIII), No. 1, March 2018

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Published

2018-03-30

How to Cite

BODA, F.-A. ., SALAMON, P. ., ORBÁN, C. ., BERȚA, L.-A. ., CURTICĂPEAN, A. ., GÂZ, Șerban-A. ., & DOGARU, M. . (2018). HETEROLOGOUS EXPRESSION AND PURIFICATION OF RECOMBINANT CROTOXIN B, THE PHOSPHOLIPASE A2 SUBUNIT OF CROTOXIN. Studia Universitatis Babeș-Bolyai Chemia, 63(1), 7–20. https://doi.org/10.24193/subbchem.2018.1.01

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Volume 63, No. 1, 2018

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