Journal of Microbiology and Biotechnology

  • 제23권2호
  • /
  • Pages.167-176
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  • 2013
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  • 1017-7825(pISSN)
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  • 1738-8872(eISSN)
한국미생물·생명공학회 (The Korean Society for Microbiology and Biotechnology)
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Purification and Characterization of the Bacteriocin Thuricin Bn1 Produced by Bacillus thuringiensis subsp. kurstaki Bn1 Isolated from a Hazelnut Pest

  • Ugras, Serpil (Department of Biology, Faculty of Science, Karadeniz Technical University) ;
  • Sezen, Kazim (Department of Biology, Faculty of Science, Karadeniz Technical University) ;
  • Kati, Hatice (Department of Biology, Faculty of Arts and Science, Giresun University) ;
  • Demirbag, Zihni (Department of Biology, Faculty of Science, Karadeniz Technical University)
  • 투고 : 2012.09.20
  • 심사 : 2012.10.15
  • 발행 : 2013.02.28
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초록

A novel bioactive molecule produced by Bacillus thuringiensis subsp. kurstaki Bn1 (Bt-Bn1), isolated from a common pest of hazelnut, Balaninus nucum L. (Coleoptera: Curculionidae), was determined, purified, and characterized in this study. The Bt-Bn1 strain was investigated for antibacterial activity with an agar spot assay and well diffusion assay against B. cereus, B. weinhenstephenensis, L. monocytogenes, P. savastanoi, P. syringae, P. lemoignei, and many other B. thuringiensis strains. The production of bioactive molecule was determined at the early logarithmic phase in the growth cycle of strain Bt-Bn1 and its production continued until the beginning of the stationary phase. The mode of action of this molecule displayed bacteriocidal or bacteriolytic effect depending on the concentration. The bioactive molecule was purified 78-fold from the bacteria supernatant with ammonium sulfate precipitation, dialysis, ultrafiltration, gel filtration chromatography, and HPLC, respectively. The molecular mass of this molecule was estimated via SDS-PAGE and confirmed by the ESI-TOFMS as 3,139 Da. The bioactive molecule was also determined to be a heat-stable, pH-stable (range 6-8), and proteinase K sensitive antibacterial peptide, similar to bacteriocins. Based on all characteristics determined in this study, the purified bacteriocin was named as thuricin Bn1 because of the similarities to the previously identified thuricin-like bacteriocin produced by the various B. thuringiensis strains. Plasmid elution studies showed that gene responsible for the production of thuricin Bn1 is located on the chromosome of Bt-Bn1. Therefore, it is a novel bacteriocin and the first recorded one produced by an insect originated bacterium. It has potential usage for the control of many different pathogenic and spoilage bacteria in the food industry, agriculture, and various other areas.

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참고문헌

  1. Abriouel, H., C. M. Franz , N. Ben Omar, and A. Galvez. 2011. Diversity and applications of Bacillus bacteriocins. FEMS Microbiol. Rev. 35: 201-232. https://doi.org/10.1111/j.1574-6976.2010.00244.x
  2. Ahern, M., S Verschueren, and D. V. Sinderen. 2003. Isolation and characterisation of a novel bacteriocin produced by Bacillus thuringiensis strain B439. FEMS Microbiol. Lett. 220: 127-131. https://doi.org/10.1016/S0378-1097(03)00086-7
  3. Anderson, I., A. Sorokin, and V. Kapatral. 2005. Comparative genome analysis of Bacillus cereus group genomes with Bacillus subtilis. FEMS Microbiol. Lett. 250: 175-184. https://doi.org/10.1016/j.femsle.2005.07.008
  4. Barboza-Corona, J. E., H. Vazquez-Acosta, D. K. Bideshi, and R. Salcedo-Hernandez.. 2007. Bacteriocin-like inhibitor substances produced by Mexican strains of Bacillus thuringiensis. Arch. Microbiol. 187: 117-126. https://doi.org/10.1007/s00203-006-0178-5
  5. Barboza-Corona, J. E., N. Fuente-Salcido, N. Alva-Murillo, A. Ochoa-Zarzosa, and J. E. Lopez-Meza. 2009. Activity of bacteriocins synthesized by Bacillus thuringiensis against Staphylococcus aureus isolates associated to bovine mastitis. Vet. Microbiol. 138: 179-183. https://doi.org/10.1016/j.vetmic.2009.03.018
  6. Beegle, C. C. and T. Yamamoto. 1992. History of Bacillus thuringiensis Berliner research and development. Can. Entomol. 124: 587-616. https://doi.org/10.4039/Ent124587-4
  7. Bradford, M. M. 1976. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal. Biochem. 72: 248. https://doi.org/10.1016/0003-2697(76)90527-3
  8. Chehimi, S., F. Delalande S. Sable, M. R. Hajlaoui, A. Van Dorsselaer, F. Limam, and A. M. Pons. 2007. Purification and partial amino acid sequence of thuricin S, a new anti-Listeria bacteriocin from Bacillus thuringiensis. Can. J. Microbiol. 53: 284-290. https://doi.org/10.1139/w06-116
  9. Cherif, A., H. Ouzari, D. Daffonchio, H. Cherif, K. Ben Slama, A. Hassen, et al. 2001. Thuricin 7: A novel bacteriocin produced by Bacillus thuringiensis BMG1.7, a new strain isolated from soil. Lett. Appl. Microbiol. 32: 243-247. https://doi.org/10.1046/j.1472-765X.2001.00898.x
  10. Cherif, A., S. Chehimi, F. Limem, B. M. Hansen, N. B. Hendriksen, D. Daffonchio, and A. Boudabous. 2003. Detection and characterization of the novel bacteriocin entomocin 9, and safety evaluation of its producer, Bacillus thuringiensis subsp. entomocidus HD9. J. Appl. Microbiol. 95: 990-1000. https://doi.org/10.1046/j.1365-2672.2003.02089.x
  11. Cherif, A., W. Rezgui, N. Raddadi, D. Daffonchio, and A. Boudabous. 2008. Characterization and partial purification of entomocin 110, a newly identified bacteriocin from Bacillus thuringiensis subsp. entomocidus HD110. Microbiol. Res. 163: 684-692. https://doi.org/10.1016/j.micres.2006.10.005
  12. Chen, H. and D. G. Hoover. 2003. Bacteriocins and their food applications. Comprehen. Rev. Food Sci. Food Safety 2: 82-100
  13. Cleveland, J., T. J. Montville, I. F. Nes, and M. L. Chikindas. 2001. Bacteriocins: Safe, natural antimicrobials for food preservation. Int. J. Food Microbiol. 71: 1-20. https://doi.org/10.1016/S0168-1605(01)00560-8
  14. Delves-Broughton, J. 1990. Nisin and its uses as food preservative. Food Tech. 44: 100-117.
  15. De la Fuente-Salcido, N., M. G. Alanís-Guzmán, D. K. Bideshi, R. Salcedo-Hernández, M. Bautista-Justo, and J. E. Barboza- Corona. 2008. Enhanced synthesis and antimicrobial activities of bacteriocins produced by Mexican strains of Bacillus thuringiensis. Arch. Microbiol. 190: 633-640. https://doi.org/10.1007/s00203-008-0414-2
  16. DeVuyst, L. and E. J. Vandamme. 1994. Nisin, a lantibiotic produced by Lactococcus lactis subsp. lactis: Properties, biosynthesis, fermentation and applications, pp. 151-221. In L. De Vuyst and E. J. Vandamme (eds.). Bacteriocins of Lactic Acid Bacteria. Chapman and Hall, London.
  17. Favret, M. E. and A. A. Yousten. 1989. Thuricin: The bacteriocin produced by Bacillus thuringiensis. J. Invertebr. Pathol. 53: 206-216. https://doi.org/10.1016/0022-2011(89)90009-8
  18. Feitelson, J. S., J. Payne, and L. Kim. 1992. Bacillus thuringiensis: Insects and beyond. BioTechnology 10: 271-275. https://doi.org/10.1038/nbt0392-271
  19. Ghanbari, M., M. Rezaei, M. Soltani, and G. Shah-Hosseini. 2009. Production of bacteriocin by a novel Bacillus sp. strain RF 140, an intestinal bacterium of Caspian Frisian Roach (Rutilus frisiikutum). Iran. J. Vet. Res. 10: 267-272.
  20. Gray, E. J., K. D. Lee, A. M. Souleimanov, M. R. Di Falco, X. Zhou, A. Ly, et al. 2006. A novel bacteriocin, thuricin 17, produced by plant growth promoting rhizobacteria strain Bacillus thuringiensis NEB17: Isolation and classification. J. Appl. Microbiol. 100: 545-554. https://doi.org/10.1111/j.1365-2672.2006.02822.x
  21. Hardy, K. G. 1993. Plasmids, pp. 138-272. In K. G. Hardy (ed.). The Practical Approach Series, 2nd Ed.
  22. Hyronimus, B., C. Le Marrec, and M. C. Urdaci. 1998. Coagulin, a bacteriocin-like inhibitory substance produced by Bacillus coagulans I4. J. Appl. Microbiol. 85: 42-50. https://doi.org/10.1046/j.1365-2672.1998.00466.x
  23. Jack, R. W., J. R. Tagg, and B. Ray. 1995. Bacteriocins of Gram positive bacteria. Microbiol. Rev. 59: 171-200.
  24. Kamoun, F., H. Mejdoub, H. Aouissaoui, J. Reinbolt, A. Hammami, and S. Jaoua. 2005. Purification, amino acid sequence and characterization of bacthuricin F4, a new bacteriocin produced by Bacillus thuringiensis. J. Appl. Microbiol. 98: 881-888. https://doi.org/10.1111/j.1365-2672.2004.02513.x
  25. Klaenhammer, T. R. 1993. Genetics of bacteriocins produced by lactic acid bacteria. FEMS Microbiol. Rev. 12: 39-86. https://doi.org/10.1111/j.1574-6976.1993.tb00012.x
  26. Klein, C., C. Kaletta, and K. D. Entian. 1993. Biosynthesis of the lantibiotic subtilin is regulated by a histidine kinase/response regulator system. Appl. Environ. Microbiol. 59: 296-303.
  27. Laemmli, U. K. 1970. Cleavage of the structural proteins during the assembly of the head of bacteriophage T4. Nature 227: 680-685. https://doi.org/10.1038/227680a0
  28. Lee, H., J. J. Churey, and R. W. Worobo. 2009a. Biosynthesis and transcriptional analysis of thurincin H, a tandem repeated bacteriocin genetic locus, produced by Bacillus thuringiensis SF361. FEMS Microbiol. Lett. 299: 205-213. https://doi.org/10.1111/j.1574-6968.2009.01749.x
  29. Lee, K. D., E. J. Gray, F. Mabood, W. J. Jung, T. Charles, S. R. Clark, et al. 2009b. The class IId bacteriocin thuricin-17 increases plant growth. Planta 229: 747-755. https://doi.org/10.1007/s00425-008-0870-6
  30. Naclerio, G., E. Ricca, M. Sacco, and M. De Felice. 1993. Antimicrobial activity of a newly identified bacteriocin of Bacillus cereus. Appl. Environ. Microbiol. 59: 4313-4316.
  31. Padilla, C., P. Brevis, O. Lobos, and E. Hubert. 1996. Bacteriocin activity of Pseudomonas sp. on enteropathogenic bacteria in an artificial aquatic system. Lett. Appl. Microbiol. 23: 371-374. https://doi.org/10.1111/j.1472-765X.1996.tb01339.x
  32. Paik, H. D., S. S. Bae, and J. G. Pan. 1997. Identification and partial characterization of tochicin, a bacteriocin produced by Bacillus thuringiensis subsp. tochigiensis. J. Ind. Microbiol. Biotechnol. 19: 294-298. https://doi.org/10.1038/sj.jim.2900462
  33. Ross, R. P., S. Morgan, and C. Hill. 2002. Preservation and fermentation: Past, present and future. Int. J. Food Microbiol. 79: 3-16. https://doi.org/10.1016/S0168-1605(02)00174-5
  34. Sezen, K. and Z. Demirbag. 1999. Isolation and insecticidal activity of some bacteria from the hazelnut beetle (Balaninus nucum L.). Appl. Entomol. Zool. 34: 85-89. https://doi.org/10.1303/aez.34.85

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