Advertisement

Applied Biochemistry and Microbiology

, Volume 53, Issue 6, pp 661–668 | Cite as

Expression of the zebrafish β-defensin 3 mature peptide in Pichia pastoris and its purification and antibacterial activity

  • Y. TaoEmail author
  • C. F. Song
  • W. Li
Article
  • 70 Downloads

Abstract

Defensins are abundant in cells and tissues that are involved in host defense against microbial infections. zfDB3 (zebrafish β-defensin 3) is one of 3 copies of defensin β-like genes from zebrafish (Danio rerio). Here we focus on mzfDB3, which is the gene encoding for the zebrafish β-defensin 3 mature peptide. A codon-optimized mzfDB3 gene with a 6×His-tag at the 3′-end was inserted into the pPICZαA expression vector and transformed into Pichia pastoris X-33 cells. The recombinant zebrafish β-defensin 3 mature peptide (rmzfDB3) was induced with 1.0% methanol at 29°C for 72 h and purified by immobilized metal affinity chromatography. MALDI-TOF/TOF analysis confirmed the expected purified product (rmzfDB3, 5.9 kDa). Fermentation supernatant, which contained rmzfDB3, showed antibacterial activity against Grampositive (i.e., Listeria monocytogenes, Staphylococcus aureus, and Bacillus cereus) and Gram-negative (i.e., Escherchia coli BL21, Vibrio parahaemolyticus, Salmonella lignieres, and Pseudomonas aeruginosa) bacteria.

Keywords

zebrafish β-defensin 3 mature peptide Pichia pastoris recombinant protein expression antibacterial activity 

Abbreviations

zfDB3

zebrafish ß-defensin 3

mzfDB3

zebrafish ß-defensin 3 mature peptide

rmzfDB3

recombinant zebrafish ß-defensin 3 mature peptide

IMAC

immobilized metal affinity chromatography

Preview

Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.

References

  1. 1.
    Ganz, T., Selsted, M.E., Szklarek, D., Harwig, S., Daher, K., Bainton, D.F., and Lehrer, R.I., J. Clin. Invest., 1985, vol. 76, no. 4, pp. 1427–1435.CrossRefPubMedPubMedCentralGoogle Scholar
  2. 2.
    Selsted, M.E., Harwig, S., Ganz, T., Schilling, J., and Lehrer, R., J. Clin. Invest., 1985, vol. 76, no. 4, pp. 1436–1439.CrossRefPubMedPubMedCentralGoogle Scholar
  3. 3.
    Ganz, T., Nat. Rev. Immunol., 2003, vol. 3, no. 9, pp. 710–720.CrossRefPubMedGoogle Scholar
  4. 4.
    Diamond, G., Zasloff, M., Eck, H., Brasseur, M., Maloy, W.L., and Bevins, C.L., Proc. Natl. Acad. Sci., 1991, vol. 88, no. 9, pp. 3952–3956.CrossRefPubMedPubMedCentralGoogle Scholar
  5. 5.
    Selsted, M.E., Tang, Y.Q., Morris, W.L., McGuire, P.A., Novotny, M.J., Smith, W., et al., J. Biol. Chem., 1993, vol. 268, no. 9, pp. 6641–6648.PubMedGoogle Scholar
  6. 6.
    Harder, J., Bartels, J., Christophers, E., and Schröder, J.M., J. Biol. Chem., 2001, vol. 276, no. 8, pp. 5707–5713.CrossRefPubMedGoogle Scholar
  7. 7.
    Yamaguchi, Y., Nagase, T., Makita, R., Fukuhara, S., Tomita, T., Tominaga, T., et al., J. Immunol., 2002, vol. 169, no. 5, pp. 2516–2523.CrossRefPubMedGoogle Scholar
  8. 8.
    Shnitsar, V.M., Lisovskiy, I.L., Soldatkina, M.A., Nespryadko, S.V., Turchak, O.V., Vinnitskaya, A.B., and Pogrebniy, P.V., Exp. Oncol., 2005, vol. 26, no. 5, pp. 328–330.Google Scholar
  9. 9.
    Jang, B.C., Lim, K.J., Choi, I.H., Suh, M.H., Park, J.G., Mun, K.C., et al., Int. J. Mol. Med., 2007, vol. 19, no. 5, pp. 757–763.PubMedGoogle Scholar
  10. 10.
    Beck–Broichsitter, B.E., Dau, H., Moest, T., Jochens, A., Stockmann, P., Wiltfang, J., and Becker, S.T., J. Oral. Pathol. Med., 2015, vol. 44, no. 2, pp. 88–93.CrossRefPubMedGoogle Scholar
  11. 11.
    Chen, Z., Wang, D., Cong, Y., Wang, J., Zhu, J., Yang, J., et al., Appl. Microbiol. Biotechnol., 2011, vol. 89, no. 2, pp. 281–291.CrossRefPubMedGoogle Scholar
  12. 12.
    Peng, Z., Wang, A., Feng, Q., Wang, Z., Ivanova, I.V., He, X., et al., Appl. Microbiol. Biotechnol., 2014, vol. 98, no. 12, pp. 5487–5497.CrossRefPubMedGoogle Scholar
  13. 13.
    Zou, J., Mercier, C., Koussounadis, A., and Secombes, C., Mol. Immunol., 2007, vol. 44, no. 4, pp. 638–647.CrossRefPubMedGoogle Scholar
  14. 14.
    Cuesta, A., Meseguer, J., and Esteban, M.A., Mol. Immunol., 2011, vol. 48, nos. 12–13, pp. 1432–1438.CrossRefPubMedGoogle Scholar
  15. 15.
    Guo, M., Wei, J., Huang, X., Huang, Y., and Qin, Q., Fish Shellfish Immunol., 2012, vol. 32, no. 5, pp. 828–38.CrossRefPubMedGoogle Scholar
  16. 16.
    Wang, G., Li, J., Zou, P., Xie, H., Huang, B., Nie, P., and Chang, M., Fish Shellfish Immunol., 2012, vol. 33, no. 3, pp. 522–531.CrossRefPubMedGoogle Scholar
  17. 17.
    Liang, T., Wang, D.D., Zhang, G.R., Wei, K.J., Wang, W.M., and Zou, G.W., Comp. Biochem. Physiol. B., 2013, vol. 166, no. 1, pp. 91–98.CrossRefPubMedGoogle Scholar
  18. 18.
    Jin, J.Y., Zhou, L., and Gui, J.F., Acta. Hydrobiol. Sin., 2011, vol. 5, no. 5, pp. 739–744.Google Scholar
  19. 19.
    Schägger, H. and Von Jagow, G., Anal. Biochem., 1987, vol. 166, no. 2, pp. 368–379.CrossRefPubMedGoogle Scholar
  20. 20.
    Hsu, K.H., Pei, C., Yeh, J.Y., Shih, C.H., Chung, Y.C., Hung, L.T., and Ou, B.R., J. Gen. Appl. Microbiol., 2009, vol. 55, no. 5, pp. 395–401.CrossRefPubMedGoogle Scholar
  21. 21.
    Wang, A., Wang, S., Shen, M., Chen, F., Zou, Z., Ran, X., et al., Appl. Microbiol. Biotechnol., 2009, vol. 84, no. 5, pp. 877–884.CrossRefPubMedGoogle Scholar
  22. 22.
    Zhang, Y., Teng, D., Mao, R.Y., Wang, X.M., Xi, D., Hu, X.Y., and Wang, J.H., Appl. Microbiol. Biotechnol., 2014, vol. 98, no. 12, pp. 681–694.CrossRefPubMedGoogle Scholar
  23. 23.
    Cao, X.T., Zhang, Y., Mao, R.Y., Teng, D., Wang, X.M., and Wang, J.H., Appl. Microbiol. Biotechnol., 2015, vol. 99, no. 6, pp. 2649–2662.CrossRefPubMedGoogle Scholar
  24. 24.
    Kant, P., Liu, W.Z., and Pauls, K.P., Peptides, 2009, vol. 30, no. 9, pp. 1593–1599.CrossRefPubMedGoogle Scholar

Copyright information

© Pleiades Publishing, Inc. 2017

Authors and Affiliations

  1. 1.College of Food Science and TechnologyShanghai Ocean UniversityShanghaiChina

Personalised recommendations