Skip to main content

Advertisement

Log in

Northeast Red Beans Produce a Thermostable and pH-Stable Defensin-Like Peptide with Potent Antifungal Activity

  • Original Paper
  • Published:
Cell Biochemistry and Biophysics Aims and scope Submit manuscript

Abstract

A 5.4-kDa antifungal peptide was purified from Phaseolus vulgaris L. cv. “northeast red bean” using a protocol that entailed affinity chromatography, ion exchange chromatography, and gel filtration. The molecular mass was determined by matrix-assisted laser desorption ionization time-of-flight. The N-terminal amino acid sequence of the peptide was highly homologous to defensins and defensin-like peptides from several plant species. The peptide impeded the growth of a number of pathogenic fungi, including Mycosphaerella arachidicola Khokhr. (IC50 = 1.7 μM), Setosphaeria turcica Luttr., Fusarium oxysporum Schltdl., and Valsa mali Miyabe & G. Yamada. Antifungal activity of the peptide was fully preserved at temperatures up to 100 °C and pH values from 0 to 12. Congo red deposition at the hyphal tip of M. arachidicola was detected after exposure to the peptide, signifying that the peptide had suppressed hyphal growth. The antifungal peptide did not manifest antiproliferative activity toward human breast cancer MCF7 cells and hepatoma HepG2 cells, in contradiction to the bulk of previously reported plant defensins. The data suggest distinct structural requirements for antifungal and antiproliferative activities.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6

Similar content being viewed by others

References

  1. Fernandez-Caballero, C., Romero, I., Goñi, O., Escribano, M. I., Merodio, C., & Sanchez-Ballesta, M. T. (2009). Characterization of an antifungal and cryoprotective class I chitinase from table grape berries (Vitis vinifera cv. Cardinal). Journal of Agriculture and Food Chemistry, 57, 8893–8900.

    Article  CAS  Google Scholar 

  2. Ye, X. Y., & Ng, T. B. (2001). Isolation of unguilin, a cyclophilin-like protein with anti-mitogenic, antiviral, and antifungal activities, from black-eyed pea. Journal of Protein Chemistry, 20, 353–359.

    Article  PubMed  CAS  Google Scholar 

  3. Sharma, U., & Suresh, C. G. (2011). Purification, crystallization and X-ray characterization of a Kunitz-type trypsin inhibitor protein from the seeds of chickpea (Cicer arietinum). Acta Crystallographica, Section F: Structural Biology and Crystallization Communications, 67, 714–717.

    Article  CAS  Google Scholar 

  4. Banović, B., Surbanovski, N., Konstantinović, M., & Maksimović, V. (2009). Basic RNases of wild almond (Prunus webbii): Cloning and characterization of six new S-RNase and one “non-S RNase” genes. Journal of Plant Physiology, 166, 395–402.

    Article  PubMed  Google Scholar 

  5. Shu, S. H., Xie, G. Z., Guo, X. L., & Wang, M. (2009). Purification and characterization of a novel ribosome-inactivating protein from seeds of Trichosanthes kirilowii Maxim. Protein Expression and Purification, 67, 120–125.

    Article  PubMed  CAS  Google Scholar 

  6. Marzouki, S. M., Almagro, L., Sabater-Jara, A. B., Ros Barceló, A., & Pedreño, M. A. (2010). Kinetic characterization of a basic peroxidase from garlic (Allium sativum L.) cloves. Journal of Food Science, 75, 740–746.

    Article  Google Scholar 

  7. Lin, J., Zhou, X., Wang, J., Jiang, P., & Tang, K. (2010). Purification and characterization of curcin, a toxic lectin from the seed of Jatropha curcas. Preparative Biochemistry and Biotechnology, 40, 107–118.

    Article  PubMed  CAS  Google Scholar 

  8. Gao, A. G., Hakimi, S. M., Mittanck, C. A., Wu, Y., Woerner, B. M., & Stark, D. M. (2000). Fungal pathogen protection in potato by expression of a plant defensin peptide. Nature Biotechnology, 18, 1307–1310.

    Article  PubMed  CAS  Google Scholar 

  9. Ngai, P. H., & Ng, T. B. (2005). Phaseococcin, an antifungal protein with antiproliferative and anti-HIV-1 reverse transcriptase activities from small scarlet runner beans. Biochemistry and Cell Biology, 83, 212–220.

    Article  PubMed  CAS  Google Scholar 

  10. Beer, A., & Vivier, M. A. (2008). Vv-AMP1, a ripening induced peptide from Vitis vinifera shows strong antifungal activity. BMC Plant Biology, 8, 75.

    Article  PubMed  Google Scholar 

  11. Chen, G. H., Hsu, M. P., Tan, C. H., Sung, H. Y., Kuo, C. G., & Fan, M. J. (2005). Cloning and characterization of a plant defensin VaD1 from azuki bean. Journal of Agriculture and Food Chemistry, 53, 982–988.

    Article  CAS  Google Scholar 

  12. Mendez, E., Moreno, A., Colilla, F., Pelaez, F., Limas, G. G., & Mendez, R. (1990). Primary structure and inhibition of protein synthesis in eukaryotic cell-free system of a novel thionin, γ-hordothionin, from barley endosperm. European Journal of Biochemistry, 194, 533–539.

    Article  PubMed  CAS  Google Scholar 

  13. Bloch, C. J., & Richardson, M. (1991). A new family of small (5 kDa) protein inhibitors of insect α-amylases from seeds of sorghum (Sorghum bicolor (L) Moench) have sequence homologies with wheat γ-purothionins. FEBS Letters, 279, 101–104.

    Article  PubMed  CAS  Google Scholar 

  14. Laemmli, U. K., & Favre, M. (1973). Gel electrophoresis of proteins. Journal of Molecular Biology, 80, 575–599.

    Article  PubMed  CAS  Google Scholar 

  15. Silva, J. A., Macedo, M. L., Novello, J. C., & Marangoni, S. (2001). Biochemical characterization and N-terminal sequences of two new trypsin inhibitors from Copaifera langsdorffii seeds. Journal of Protein Chemistry, 20, 1–7.

    Article  PubMed  CAS  Google Scholar 

  16. Fujimura, M., Minami, Y., Watanabe, K., & Tadera, K. (2003). Purification, characterization, and sequencing of a novel type of antimicrobial peptides, Fa-AMP1 and Fa-AMP2, from seeds of buckwheat (Fagopyrum esculentum Moench.). Bioscience, Biotechnology, and Biochemistry, 67, 1636–1642.

    Article  PubMed  CAS  Google Scholar 

  17. Edman, P. (1950). Method for determination of amino acid sequences in peptides. Acta Chemica Scandinavica, 28, 283–293.

    Article  Google Scholar 

  18. Bradford, M. M. (1976). A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Analytical Biochemistry, 7, 248–254.

    Article  Google Scholar 

  19. Wong, J. H., & Ng, T. B. (2003). Gymnin, a potent defensin-like antifungal peptide from the Yunnan bean (Gymnocladus chinensis Baill). Peptides, 24, 963–968.

    Article  PubMed  CAS  Google Scholar 

  20. Nakagawa, R., Yasokawa, D., Ikeda, T., & Nagashima, K. (1996). Purification and characterization of two lectins from callus of Helianthus tuberosus. Bioscience, Biotechnology, and Biochemistry, 60, 259–262.

    Article  PubMed  CAS  Google Scholar 

  21. Moreno, A. B., Martinez Del Pozo, A., & San Segundo, B. (2006). Biotechnologically relevant enzymes and proteins. Antifungal mechanism of the Aspergillus giganteus AFP against the rice blast fungus Magnaporthe grisea. Applied Microbiology and Biotechnology, 72, 883–895.

    Article  PubMed  CAS  Google Scholar 

  22. Wu, X., Sun, J., Zhang, G., Wang, H., & Ng, T. B. (2011). An antifungal defensin from Phaseolus vulgaris cv. ‘Cloud Bean’. Phytomedicine, 18, 104–109.

    Article  PubMed  Google Scholar 

  23. Leung, E. H., Wong, J. H., & Ng, T. B. (2008). Concurrent purification of two defense proteins from French bean seeds: A defensin-like antifungal peptide and a hemagglutinin. Journal of Peptide Science, 14, 349–353.

    Article  PubMed  CAS  Google Scholar 

  24. Yin, F., Pajak, A., Chapman, R., Sharpe, A., Huang, S., & Marsolais, F. (2011). Analysis of common bean expressed sequence tags identifies sulfur metabolic pathways active in seed and sulfur-rich proteins highly expressed in the absence of phaseolin and major lectins. BMC Genomics, 12, 268.

    Article  PubMed  CAS  Google Scholar 

  25. Pelegrini, P. B., Lay, F. T., Murad, A. M., Anderson, M. A., & Franco, O. L. (2008). Novel insights on the mechanism of action of alpha-amylase inhibitors from the plant defensin family. Proteins, 73, 719–729.

    Article  PubMed  CAS  Google Scholar 

  26. Carvalho Ade, O., & Gomes, V. M. (2009). Plant defensins–prospects for the biological functions and biotechnological properties. Peptides, 30, 1007–1020.

    Article  PubMed  Google Scholar 

  27. Lin, P., Wong, J. H., & Ng, T. B. (2009). A defensin with highly potent antipathogenic activities from the seeds of purple pole bean. Bioscience Reports, 30, 101–109.

    Article  PubMed  Google Scholar 

  28. Wong, J. H., Zhang, X. Q., Wang, H. X., & Ng, T. B. (2006). A mitogenic defensin from white cloud beans (Phaseolus vulgaris). Peptides, 27, 2075–2081.

    Article  PubMed  CAS  Google Scholar 

  29. Lee-Huang, S., Kung, H. F., Huang, P. L., Bourinbaiar, A. S., Morell, J. L., Brown, J. H., et al. (1994). Human immunodeficiency virus type 1 (HIV-1) inhibition, DNA-binding, RNA-binding, and ribosome inactivation activities in the N-terminal segments of the plant anti-HIV protein GAP31. Proceedings of the National Academy of Sciences of USA, 91, 12208–12212.

    Article  CAS  Google Scholar 

  30. Shestakov, A., Jenssen, H., Nordström, I., & Eriksson, K. (2012). Lactoferricin but not lactoferrin inhibit herpes simplex virus type 2 infection in mice. Antiviral Research, 93, 340–345.

    Article  PubMed  CAS  Google Scholar 

  31. Farnaud, S., Patel, A., Odell, E. W., & Evans, R. W. (2004). Variation in antimicrobial activity of lactoferricin-derived peptides explained by structure modelling. FEMS Microbiology Letters, 238, 221–226.

    Article  PubMed  CAS  Google Scholar 

  32. Ye, X. Y., Ng, T. B., Tsang, P. W., & Wang, J. (2001). Isolation of a homodimeric lectin with antifungal and antiviral activities from red kidney bean (Phaseolus vulgaris) seeds. Journal of Protein Chemistry, 20, 367–375.

    Article  PubMed  CAS  Google Scholar 

  33. Xia, L., & Ng, T. B. (2005). An antifungal protein from flageolet beans. Peptides, 26, 2397–2403.

    Article  PubMed  CAS  Google Scholar 

  34. He, X. M., Ji, N., Xiang, X. C., Luo, P., & Bao, J. K. (2011). Purification, characterization, and molecular cloning of a novel antifungal lectin from the roots of Ophioglossum pedunculosum. Applied Biochemistry and Biotechnology, 165, 1458–1472.

    Article  PubMed  CAS  Google Scholar 

  35. Kheeree, N., Sangvanich, P., Puthong, S., & Karnchanatat, A. (2010). Antifungal and antiproliferative activities of lectin from the rhizomes of Curcuma amarissima Roscoe. Applied Biochemistry and Biotechnology, 162, 912–925.

    Article  PubMed  CAS  Google Scholar 

  36. Lam, S. K., & Ng, T. B. (2010). Isolation and characterization of a French bean hemagglutinin with antitumor, antifungal, and anti-HIV-1 reverse transcriptase activities and an exceptionally high yield. Phytomedicine, 17, 457–462.

    Article  PubMed  CAS  Google Scholar 

  37. Liu, Y., Chen, Z., Ng, T. B., Zhang, J., Zhou, M., Song, F., et al. (2007). Bacisubin, an antifungal protein with ribonuclease and hemagglutinating activities from Bacillus subtilis strain B-916. Peptides, 28, 553–559.

    Article  PubMed  Google Scholar 

  38. Diao, H., Yu, H. G., Sun, F., Zhang, Y. L., & Tanphaichitr, N. (2011). Rat recombinant β-defensin 22 is a heparin-binding protein with antimicrobial activity. Asian Journal of Andrology, 13, 305–311.

    Article  PubMed  CAS  Google Scholar 

  39. Chen, W., Yang, B., Zhou, H., Sun, L., Dou, J., Qian, H., et al. (2011). Structure–activity relationships of a snake cathelicidin-related peptide, BF-15. Peptides, 32, 2497–2503.

    Article  PubMed  CAS  Google Scholar 

  40. Franco, O. L. (2011). Peptide promiscuity: An evolutionary concept for plant defense. FEBS Letters, 585, 995–1000.

    Article  PubMed  CAS  Google Scholar 

  41. Thevissen, K., Ferket, K. K., Francois, I. E., & Cammue, B. P. (2003). Interactions of antifungal plant defensins with fungal membrane components. Peptides, 24, 1705–1712.

    Article  PubMed  CAS  Google Scholar 

Download references

Author information

Authors and Affiliations

Authors

Corresponding author

Correspondence to Tzi Bun Ng.

Electronic supplementary material

Below is the link to the electronic supplementary material.

Supplementary material 1 (TIFF 2178 kb)

Rights and permissions

Reprints and permissions

About this article

Cite this article

Chan, Y.S., Ng, T.B. Northeast Red Beans Produce a Thermostable and pH-Stable Defensin-Like Peptide with Potent Antifungal Activity. Cell Biochem Biophys 66, 637–648 (2013). https://doi.org/10.1007/s12013-012-9508-1

Download citation

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s12013-012-9508-1

Keywords

Navigation