DOI QR코드

DOI QR Code

Cloning, Characterization and Antifungal Activity of Defensin Tfgd1 from Trigonella foenum-graecum L.

  • Olli, Sudar (Department of Plant Sciences, School of Life Sciences, University of Hyderabad) ;
  • Kirti, P.B. (Department of Plant Sciences, School of Life Sciences, University of Hyderabad)
  • Received : 2005.07.21
  • Accepted : 2006.01.30
  • Published : 2006.05.31

Abstract

Defensins are small cysteine rich peptides with a molecular mass of 5-10 kDa and some of them exhibit potent antifungal activity. We have cloned the coding region of a cDNA of 225 bp cysteine rich defensin, named as Tfgd1, from the legume Trigonella foenum-graecum. The amino acid sequence deduced from the coding region comprised 74 amino acids, of which the N-terminal 27 amino acids constituted the signal peptide and the mature peptide comprised 47 amino acids. The protein is characterized by the presence of eight cysteine resisdues, conserved in the various plant defensins forming four disulphide bridges, which stabilize the mature peptide. The recombinant protein expressed in E coli exhibited antifungal activity against the broad host range fungus, Rhizoctonia solani and the peanut leaf spot fungus, Phaeoisariopsis personata.

Keywords

References

  1. Ai-Guo, G., Hakimi, S. M., Mittnack, C. A., Yonnie, W., Woerner, M., Stark, D. M., Shah, D. M., Liang, J. and Rommens, C. M. T. (2000) Fungal pathogen protection in potato by expression of a plant defensin peptide. Nature Bio/Technology 18, 1307- 1310 https://doi.org/10.1038/82436
  2. Bert, J. C. J., Horst, J. S., Fung, T. L., Marilyn, A. A. and David, J. C. (2003) Structure of petunia hybrida defensin 1, a novel plant defensin with five disulfide bonds. Biochemistry 42, 8214-8222 https://doi.org/10.1021/bi034379o
  3. Bloch, C. Jr. and Richardson, M. (1991) A new family of small (5 kDa) protein inhibitors of insect alpha-amylases from seeds or sorghum (Sorghum bicolor moench) have sequence homologies with wheat ${\gamma}$-purothionins. FEBS Lett. 279, 101-104 https://doi.org/10.1016/0014-5793(91)80261-Z
  4. Broekaert, W. F., Cammue, B. P. A., DeBolle, M. F. C., Thevissen, K., Desamblanx, G. W. and Osborn, R. W. (1997) Antimicrobial peptides from plants. Crit. Rev. Plant Sci. 16, 297-323 https://doi.org/10.1080/07352689709701952
  5. Broekaert, W. F., Terras, F. R. G., Cammue, B. P. A. and Osborn, R. W. (1995) Plant defensins: Novel antimicrobial peptides as components of the host defense system. Plant Physiol. 108, 1353-1358 https://doi.org/10.1104/pp.108.4.1353
  6. Bruix, M. A., Jimenez, J., Santoro, C., Gonzalez, F. J., Colilla, E. and Mendez, M. R. (1993) Solution structure of gamma 1-H and gamma 1-P thionins from barley and wheat endosperm determined by 1H-NMR: a structural motif common to toxic arthropod proteins. Biochemistry 32, 715-724 https://doi.org/10.1021/bi00053a041
  7. Colilla, F. J., Rocher, A. and Mendez, E. (1990) ${\gamma}$-purothionins: amino acid sequence of two polypeptides of a new family of thionins from wheat endosperm. FEBS Lett. 270, 191-194 https://doi.org/10.1016/0014-5793(90)81265-P
  8. Cornet, B., Bonmatin, J. M., Hetru, C., Hoffmann, J. A., Ptak, M. and Vovelle, F. (1995) Refined three-dimensional solution structure of insect defensin A. Structure 3, 435-448 https://doi.org/10.1016/S0969-2126(01)00177-0
  9. Fant, F., Vranken, W., Broekaert, W. and Borremans, F. (1998) Determination of the three-dimensional solution structure of Raphanus sativus antifungal protein by 1H NMR. J. Mol. Biol. 279, 257-270 https://doi.org/10.1006/jmbi.1998.1767
  10. Laemmli, U. K. (1970) Cleavage of structural protein during the assembly of the head of bacteriophage T4. Nature 227, 680- 685 https://doi.org/10.1038/227680a0
  11. Lay, F. T., Schirra, H. J., Scanlon, M. J., Anderson, M. A. and Craik, D. J. (2003) The three-dimensional solution structure of NaD1, a new floral defensin from Nicotiana alata and its application to a homology model of the crop defense protein AlfAFP. J. Mol. Biol. 325, 175-188 https://doi.org/10.1016/S0022-2836(02)01103-8
  12. Melo, F. R., Rigden, D. J., Franco, O. L., Mello, L. V., Ay, M. B., Grossi de Sa, M. F. and Bloch, C. Jr. (2002) Inhibition of trypsin by cowpea thionin: characterization, molecular modeling, and docking. Proteins Struct. Funct. Genet. 48, 311- 319 https://doi.org/10.1002/prot.10142
  13. Mendez, E., Moreno, A., Collila, F., Pelaez, F., Limas, G. G., Mendez, R., Soriano, F., Sainas, M. and DeHaro, C. (1990) Primary structure and inhibition of protein synthesis in eukaryotic cell-free system of a novel thionin, ${\gamma}$-hordothionin, from barley endosperm. Eur. J. Biochem. 194, 533-539 https://doi.org/10.1111/j.1432-1033.1990.tb15649.x
  14. Mendez, E., Rocher, A., Calero, M., Girbes, T., Citores, L. and Soriano, F. (1996) Primary structure of ${\omega}$-hordothionin, a novel member of a family of thionins from barley endosperm, and its inhibition of protein synthesis in eukaryotic and prokaryotic cell-free systems. J. Biochem. 239, 67-73
  15. Osborn, R. W., De Samblanx, G. W., Thevissen, K., Goderis, I., Torrekens, S., Van Leuven, F., Attenborough, S., Rees, S. B. and Broekaert, W. F. (1995) Isolation and characterization of plant defensins from seeds of Asteraceae, Fabaceae, Hippocastanaceae and Saxifragaceae. FEBS Lett. 368, 257- 262 https://doi.org/10.1016/0014-5793(95)00666-W
  16. Parashina, E. V., Serdobinskii, L. A., Kalle, E. G., Lavorova, N. V., Avetisov, V. A., Lunin, V. G. and Naroditskii, B. S. (2000) Genetic engineering of oilseed rape and tomato plants expressing a radish defensin gene. Rus. J. Plant Physiol. 47, 417-423
  17. Penninckx, I. A. M. A., Thomma, B. P. H. J., Buchala, A., Métraux, J. P. and Broekaert, W. F. (1998) Parallel but cooperative activation of jasmonate and ethylene response pathways is required for induction of a plant defensin gene in Arabidopsis. Plant Cell. 10, 2103-2114 https://doi.org/10.1105/tpc.10.12.2103
  18. Sambrook, J., Fritsch, E. F. and Maniatis, T. (1989) Molecular Cloning: A Laboratory Manual, 2nd ed., Cold Spring Harbor Laboratory Press, New York, USA
  19. Spelbrink, R. G., Dilmac, N., Allen, A., Smith, T. J., Shah, D. M. and Hockerman, G. H. (2004) Differential antifungal and calcium channel-blocking activity among structurally related plant defensins. Plant Physiol. 135, 2055-2067 https://doi.org/10.1104/pp.104.040873
  20. Terras, F. R., Eggermont, K., Kovaleva, V., Raikhel, N. V., Osborn, R. W., Kester, A., Rees, S. B., Torrekens, S., Van Leuven, F. and Vanderleyden, J. (1995) Small cysteine-rich antifungal proteins from radish: their role in host defense. Plant Cell. 7, 573-588 https://doi.org/10.1105/tpc.7.5.573
  21. Thevissen, K., Ghazi, A., De Samblanx, G. W., Brownlee, C., Osborn, R. W. and Broekaert, W. F. (1996) Fungal membrane responses induced by plant defensins and thionins. J. Biol. Chem. 271, 15018-15025 https://doi.org/10.1074/jbc.271.25.15018
  22. Thevissen, K., Osborn, R. W., Acland, D. P. and Broekaert, W. F. (2000) Specific binding sites for an antifungal plant defensin from dahlia (Dahlia merckii) on fungal cells are required for antifungal activity. Mol. Plant-Microbe Interact. 13, 54-61 https://doi.org/10.1094/MPMI.2000.13.1.54
  23. Thevissen, K., Warnecke, D. C., François, I. E. J. A., Leipelt, M., Heinz, E., Ott, C., Zähringer, U., Thomma, B. P. H. J., Ferket, K. K. A. and Cammue, B. P. A. (2004) Defensins from insects and plants interact with fungal glucosylceramides. J. Biol. Chem. 279, 3900-3905 https://doi.org/10.1074/jbc.M311165200
  24. Wang, Y., Nowak, G., Culley, D., Hadwiger, L. A. and Fristensky, B. (1999) Constitutive expression of a pea defense gene DRR206 confers resistance to blackleg (Leptosphaeria maculans ) disease in transgenic canola (Brassica napus). Mol. Plant-Microbe Interact. 12, 410-418 https://doi.org/10.1094/MPMI.1999.12.5.410
  25. Wijaya, R., Neumann, G. M., Condron, R., Hughes, A. B. and Polya, G. M. (2000) Defense proteins from seed of Cassia fistula include a lipid transfer protein homologue and a protease inhibitory plant defensin. Plant Sci, 159, 243-255 https://doi.org/10.1016/S0168-9452(00)00348-4
  26. Zhang, N. Y., Jones, B. L. and Tao, P. (1997) Purification and characterization of a new class of insect ${\alpha}$-amylase inhibitors from barley. Cereal Chem. 74, 119-122 https://doi.org/10.1094/CCHEM.1997.74.2.119

Cited by

  1. Novel antifungal defensins from Nigella sativa L. seeds vol.49, pp.2, 2011, https://doi.org/10.1016/j.plaphy.2010.10.008
  2. Identification, characterization and expression of a defensin-like antifungal peptide from the whiteflyBemisia tabaci(Gennadius) (Hemiptera: Aleyrodidae) vol.22, pp.3, 2013, https://doi.org/10.1111/imb.12021
  3. Defensin protein from sweet potato (Ipomoea batatas [L.] Lam ‘Tainong 57’) storage roots exhibits antioxidant activities in vitro and ex vivo vol.135, pp.3, 2012, https://doi.org/10.1016/j.foodchem.2012.05.082
  4. Investigating Therapeutic Potential ofTrigonella foenum-graecumL. as Our Defense Mechanism against Several Human Diseases vol.2016, 2016, https://doi.org/10.1155/2016/1250387
  5. Transgenic tobacco and peanut plants expressing a mustard defensin show resistance to fungal pathogens vol.27, pp.11, 2008, https://doi.org/10.1007/s00299-008-0596-8
  6. Antifungal Immunotherapy and Immunomodulation: A Double-hitter Approach to Deal with Invasive Fungal Infections vol.67, pp.6, 2008, https://doi.org/10.1111/j.1365-3083.2008.02101.x
  7. Prokaryotic expression of a constitutively expressed Tephrosia villosa defensin and its potent antifungal activity vol.80, pp.6, 2008, https://doi.org/10.1007/s00253-008-1648-2
  8. Assessment of Genetic Diversity in Trigonella foenum-graecum Based on Nuclear Ribosomal DNA, Internal Transcribed Spacer and RAPD Analysis vol.29, pp.2, 2011, https://doi.org/10.1007/s11105-010-0233-x
  9. A novel defensin from the lentil Lens culinaris seeds vol.371, pp.4, 2008, https://doi.org/10.1016/j.bbrc.2008.04.161
  10. Evaluation of the developmental toxicity of the aqueous extract from Trigonella foenum-graecum (L.) in mice vol.131, pp.2, 2010, https://doi.org/10.1016/j.jep.2010.06.033
  11. Pharmacological effects ofTrigonella foenum-graecumL. in health and disease vol.52, pp.2, 2014, https://doi.org/10.3109/13880209.2013.826247
  12. Identification and characterization of stress resistance related genes of Brassica rapa vol.34, pp.5, 2012, https://doi.org/10.1007/s10529-012-0860-4
  13. Enhanced resistance to Sclerotinia sclerotiorum in Brassica napus by co-expression of defensin and chimeric chitinase genes vol.57, pp.4, 2016, https://doi.org/10.1007/s13353-016-0340-y
  14. Study on the Coffea arabica/Colletotrichum kahawae pathosystem: Impact of a biological plant protection product vol.116, pp.2, 2009, https://doi.org/10.1007/BF03356290
  15. Antifungal Activity of PvD1 Defensin Involves Plasma Membrane Permeabilization, Inhibition of Medium Acidification, and Induction of ROS in Fungi Cells vol.62, pp.4, 2011, https://doi.org/10.1007/s00284-010-9847-3
  16. Screening of fusion partners for high yield expression and purification of bioactive viscotoxins vol.64, pp.1, 2009, https://doi.org/10.1016/j.pep.2008.10.003
  17. Tandem combination of Trigonella foenum-graecum defensin (Tfgd2) and Raphanus sativus antifungal protein (RsAFP2) generates a more potent antifungal protein vol.13, pp.4, 2013, https://doi.org/10.1007/s10142-013-0334-3
  18. Plant peptides in defense and signaling vol.56, 2014, https://doi.org/10.1016/j.peptides.2014.03.013
  19. In Vitro and In Vivo Activity of Selected Plant Crude Extracts and Fractions Against Penicillium Italicum vol.49, pp.4, 2009, https://doi.org/10.2478/v10045-009-0054-9
  20. Characterization of recombinant plectasin: Solubility, antimicrobial activity and factors that affect its activity vol.46, pp.5, 2011, https://doi.org/10.1016/j.procbio.2011.01.018
  21. Plant defensins—Prospects for the biological functions and biotechnological properties vol.30, pp.5, 2009, https://doi.org/10.1016/j.peptides.2009.01.018
  22. Comparison of silicon nanoparticles and silicate treatments in fenugreek vol.115, 2017, https://doi.org/10.1016/j.plaphy.2017.03.009
  23. Natural products – antifungal agents derived from plants vol.11, pp.7, 2009, https://doi.org/10.1080/10286020902942350
  24. Co-overexpression of Brassica juncea NPR1 (BjNPR1) and Trigonella foenum-graecum defensin (Tfgd) in transgenic peanut provides comprehensive but varied protection against Aspergillus flavus and Cercospora arachidicola vol.35, pp.5, 2016, https://doi.org/10.1007/s00299-016-1945-7
  25. Silicon and Mechanisms of Plant Resistance to Insect Pests vol.7, pp.2, 2018, https://doi.org/10.3390/plants7020033