Journal of Animal Science and Technology

Korean Society of Animal Sciences and Technology (한국축산학회)
권호 표지
DOI QR코드

DOI QR Code

Isolation of Bacillus amyloliquefaciens ATC6 Producing Acidic Cellulase

산성 Cellulase를 분비하는 Bacillus amyloliquefaciens ATC6의 분리

  • Lee, Se-Hyung (Department of Animal Resources Science, College of Bio-Resources Science, Dankook University) ;
  • Chae, Jong-Pyo (Department of Animal Resources Science, College of Bio-Resources Science, Dankook University) ;
  • Kim, Min-Jeong (Department of Animal Resources Science, College of Bio-Resources Science, Dankook University) ;
  • Kang, Dae-Kyung (Department of Animal Resources Science, College of Bio-Resources Science, Dankook University)
  • 이세형 (단국대학교 생명자원과학대학 동물자원학과) ;
  • 채종표 (단국대학교 생명자원과학대학 동물자원학과) ;
  • 김민정 (단국대학교 생명자원과학대학 동물자원학과) ;
  • 강대경 (단국대학교 생명자원과학대학 동물자원학과)
  • Received : 2009.08.14
  • Accepted : 2010.02.03
  • Published : 2010.02.28
Download PDF
⟨ Previous Next ⟩

Abstract

A bacterium producing acidic cellulase was isolated from pig feces. The isolate, ATC6 strain, was found to be Gram-positive, non-motile, catalase-positive, and spore-forming stain. Under an electron microscope, the cells were observed to be rod-shaped. The isolate was identified as Bacillus amyloliquefaciens ATC6 on the basis of morphological and biochemical properties as well as 16S rRNA gene sequences. Optimum pH and temperature for the cellulase activity of the culture supernatant of B. amyloliquefaciens ATC6 were found to be pH 4.5 and $55^{\circ}C$, respectively. More than 80% of its maximum activity was maintained at pH 4.0. The cellulase activity was maintained at temperatures ranging from 35 to $55^{\circ}C$ after 2 h incubation at pH 4.5, whereas it's activity decreased rapidly at $65^{\circ}C$.

사료곡물을 효율적으로 이용하기 위해, 산성 cellulase를 분비하는 ATC6 균주를 선발하였다. 선발된 ATC6 균주의 형태, 당 이용성 및 16S rDNA 서열분석 등을 통해 동정한 결과 Bacillus amyloliquefaciens에 속하는 것으로 나타났으므로, B. amyloliquefaciens ATC6로 명명하였다. B. amyloliquefaciens ATC6가 분비하는 cellulase 효소의 활성은 대수생장기 중반부터 급격히 증가하였고, 균의 생장이 정체기에 이르면 효소의 활성도 더 이상 증가하지 않는 것으로 나타났다. 효소의 최적 pH를 조사한 결과, pH 4.5에서 최대의 효소 활성을 나타내었으며, pH 4.0에서도 최고 활성의 80% 이상을 유지하였다. 또한 pH 4.0~pH 8.0까지 상대적으로 넓은 범위에서 최대 활성의 60% 이상을 유지하였다. Cellulase 활성의 최적 온도는 $55^{\circ}C$ 이었으며, $75^{\circ}C$의 고온에서도 최대 활성의 50% 정도의 활성을 나타내었다. 효소의 열 안정성을 조사한 결과, $45^{\circ}C$ 및 $55^{\circ}C$에서는 2시간 후에도 효소의 활성이 안정하게 유지되었으나, $65^{\circ}C$에서는 시간의 경과와 함께 효소 활성이 급격히 감소하는 것으로 나타났다.

Keywords

References

  1. Beguin, P. and Aubert, J. P. 1994. The biological degradation of cellulase. FEMS Microbiol. Rev. 13:25-58. https://doi.org/10.1111/j.1574-6976.1994.tb00033.x
  2. Campbell, G. L. and Bedford, M. R. 1992. Enzyme applications for monogastric feeds: A review. Can. J. Anim. Sci. 72:449-453. https://doi.org/10.4141/cjas92-058
  3. Castanon, J. I. R., Flores, M. P. and Pettersson, D. 1997. Mode of degradation of non-starch polysaccharides by feed enzyme preparations. Anim. Feed Sci. Technol. 68:361-365. https://doi.org/10.1016/S0377-8401(97)00046-1
  4. Chiang, C. -C., Bi Y. and Peter, W. -S. C. 2005. Effects of xylanase supplementation to wheat-based diet on the performance and nutrient availability of broiler chickens. Asian-Aust. J. Anim. Sci. 18(88):1141-1146. https://doi.org/10.5713/ajas.2005.1141
  5. Emtizi, G. and Nahvi, I. 2004. Production of thermostable ${\alpha}$-amylase and cellulase from Cellulomonas sp. J. Microbiol. Biotechnol. 14:1196-1199.
  6. Grigorevski-Lima, A. L, Da Vinha, F. N., Souza, D. T., Bispo, A. S., Bon, E. P., Coelho. R. R. and Nascimento, R. P. 2009. Aspergillus fumigatus thermophilic and acidophilic endoglucanases. Appl. Biochem. Biotechnol. 155:321-329.
  7. Jeong, W. H., Yang, S. Y., Song, M. D., Ha, J. G. and Kim, C. W. 2003. Isolation of Bacillus sp. producing xylanase and cellulase and optimization of medium conditions for its production. Kor. J. Microbiol. Biotechnol. 31(4):383-388.
  8. Kiarie, E., Nyachoti, C. M., Slominski, B. A. and Blank, G. 2007. Growth performance, gastrointestinal microbial activity, and nutrient digestibility in early-weaned pigs fed diets containing flaxseed and carbohydrase enzyme. J. Anim Sci. 85(11):2982-2993. https://doi.org/10.2527/jas.2006-481
  9. Kim, C. -H. and Lee, S. -S. 2003. Isolation and characterization of cellulolytic anaerobic fungi from the guts of the Hanwoo cattle and the Korean native goat. J. Anim. Sci. & Tech. (Kor). 45(6):1007-1018. https://doi.org/10.5187/JAST.2003.45.6.1007
  10. Kim, I. H., Park, J. S., Hancock, J. D., Hines, R. H., Cobb, C., Cao, H., Hong, J. W. and Kwon, O. S. 2003. Effects of amylase and cellulase supplementation in sorghum-based diets for finishing pigs. Asian-Aust. J. Anim. Sci. 16(11):70-76. https://doi.org/10.5713/ajas.2003.70
  11. Kim, J. -Y, Hur, S. -H. and Hong, J. -H. 2004. Isolation and characterization of an alkaline cellulase produced by alkalophilic Bacillus sp. HSH-810. Kor. J. Microbiol. 40(2):139-146.
  12. Kim, M. J., Lim, S. J. and Kang, D.-K. 2008. Isolation of a Bacillus licheniformis DK42 producing cellulase and xylanase, and properties of the enzymes. J. Anim. Sci. & Technol. (Kor). 50(3):1-10. https://doi.org/10.5187/JAST.2008.50.1.001
  13. Kim, T. -I., Han, J. -D., Jeon, B. -S., Ha, S.-W., Yang, C. -B. and Kim, M.-K. 1999. Isolation and characterization of Bacillus subtilis CH-10 secreting cellulase from cattle manure. Kor. J. Microbiol. 35:277-282.
  14. Miller, G. L., Blum, R., Glennon, W. E. and Burton, A. L. 1960. Measurement of carboxymethyl cellulase activity. Anal. Biochem. 2:127-132.
  15. Oh, S. -H, Moo Sung Kim, Sung So, and Hyung Joo Suh. 2003. Studies on the Production of Cellulase by Trichoderma sp. SO-571 and the Enzyme Treatment for Cellulosic Fabrics. J. Microbiol. Biotechnol. 31(3):42-45.
  16. Park, J. -K., Chan Sung Jeong, Do Yeun Park, Hyun Cheol Kim, Seung Cheol Lee and Chang-Hyun Kim. 2009. Effects of Increasing Inclusion Levels of Rumen Cellulolytic Bacteria Culture on In vivo Ruminal Fermentation Patterns in Hanwoo Heifers. J. Anim. Sci. & Technol. (Kor). 51(1):45-52. https://doi.org/10.5187/JAST.2009.51.1.045
  17. Sohn, J. H., Lee, J. H., Yi, H., Chun, J., Bae, K. S., Ahn, T. Y. and Kim, S. J. 2004. Kordia algicida gen. nov., sp. nov., an algicidal bacterium isolated from redtide. Int. J. Syst. Evol. Microbiol. 54:675-680. https://doi.org/10.1099/ijs.0.02689-0
  18. Sohail M, Siddiqi R, Ahmad A, Khan SA. 2009. Cellulase production from Aspergillus niger MS82: effect of temperature and pH. New Biotechnol. [Epub ahead of print].