Journal of Life Science (생명과학회지)

Korean Society of Life Science (한국생명과학회)
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Characterization of Agarase from an Isolated Marine Bacterium, Simiduia sp. SH-1

해양성 Simiduia sp. SH-1 균주의 분리 및 한천분해효소의 특성조사

  • Lee, Sol-Ji (Department of Bioscience, Graduate School, Silla University) ;
  • Oh, Soo-Jeong (Department of Pharmaceutical Engineering, Silla University) ;
  • Lee, Dong-Geun (Department of Bioscience, Graduate School, Silla University) ;
  • Lee, Sang-Hyeon (Department of Bioscience, Graduate School, Silla University)
  • 이솔지 (신라대학교 일반대학원 바이오과학과) ;
  • 오수정 (신라대학교 제약공학과) ;
  • 이동근 (신라대학교 일반대학원 바이오과학과) ;
  • 이상현 (신라대학교 일반대학원 바이오과학과)
  • Received : 2015.07.23
  • Accepted : 2015.09.20
  • Published : 2015.11.30
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Abstract

Agarase from a novel agar-degrading bacterium isolated from seawater in Namhae at Gyeongsangnamdo province of Korea was characterized. The SH-1 strain was selected from thousands of colonies on Marine agar 2216 media. Almost full 16S rRNA gene sequence of the agarolytic SH-1 strain showed 99% similarity with that of bacteria of Simiduia genus and named as Simiduia sp. SH-1. Agarase production was growth related, and activity was declined from stationary phase. Secreted agarase was prepared from culture media and characterized. It showed maximum activity of 698.6 units/L at pH 7.0 and 30℃ in 20 mM Tris-HCl buffer. Agarase activity decreased as the temperature increased from an optimum of 30℃, with 90% and 75% activity at 40℃ and 50℃, respectively. Agarase was not heat resistant. Slightly lower agarase activity was observed at pH 6.0 than at pH 7.0, without statistical difference, and 80% and 75% activity were observed at pH 5.0 and 8.0, respectively. Neoagarotetraose and neoagarobiose were the main final products of agarose, indicating that it is β-agarase. Simiduia sp. SH-1 and its β-agarase would be useful for the industrial production of neoagarotetraose and neoagarobiose, which have a whitening effect on skin, delaying starch degradation, and inhibiting bacterial growth.

경상남도 남해군 연안의 해수를 이용하여 신규의 한천분해 해양성세균을 분리하고 한천분해효소의 특징을 분석하였다. Marine agar 2216 배지에서 분리한 SH-1 균주는 16S rRNA 유전자 염기서열분석을 통해 Simiduia 속 세균과 99% 유사하여 Simiduia sp. SH-1으로 명명하였다. Simiduia sp. SH-1 균주가 생성하는 한천분해효소는 성장의존성 산물로 판단되었으며 정체기부터 효소활성이 감소되었다. 한천분해효소는 pH 7.0(20 mM Tris-HCl buffer), 30℃에서 최대활성(698.6 units/l)을 나타내었다. 효소의 활성은 30℃에서 최적이었고 이후 온도가 증가함에 따라 활성이 감소하였으며, 40℃와 50℃에서 각각 약 90%와 75%의 상대활성을 보였으나 내열성은 보이지 않았다. 최적 pH인 pH 7.0에 비해 pH 6.0에서는 오차범위 내에서 조금 낮은 활성을 보였으며 pH 5.0과 pH 8.0에서는 각각 80%와 75% 정도의 상대활성을 보였다. TLC 분석을 통하여 Simiduia sp. SH-1 균주가 생성하는 한천분해효소는 agarose를 분해하여 피부의 미백활성, 전분노화의 방지 및 세균성장의 억제 등의 유용한 효과를 가지는 기능성 한천올리고당인 neoagarotetraose와 neoagarobiose를 최종적으로 생성하는 β-agarase로 확인되었다. 따라서, Simiduia sp. SH-1 균주와 이 균주가 생산하는 β-agarase는 산업적 생산에 유용하게 활용될 수 있을 것으로 기대된다.

Keywords

References

  1. Cha, J. A., Kim, Y. J., Seo, Y. B. and Yoon, M. H. 2009. Isolation of an agarolytic bacteria, Cellvibrio mixtus SC-22 and the enzymatic properties. J. Appl. Biol. Chem. 52, 157-162. https://doi.org/10.3839/jabc.2009.027
  2. Chi, W. J., Lim, J. H., Park, D. Y., Kim, M. C., Kim, C. J., Chang, Y. K. and Hong, S. K. 2013. Isolation and characterization of a novel agar degrading bacterium, Alteromonas macleodii subsp. GNUM08120, from red macroalgae. Kor. J. Microbiol. Biotechnol. 41, 8-16. https://doi.org/10.4014/kjmb.1208.08001
  3. Fu, X. T., Pan, C. H., Lin, H. and Kim, S. M. 2009. Gene cloning, expression, and characterization of a beta-agarase, agaB34, from Agarivorans albus YKW-34. J. Microbiol. Biotechnol. 19, 257-264.
  4. Hong, J. H., Lee, J. J., Choi, H. S., Hur, S. H. and Kong, J. Y. 2000. Antibacterial activity of agarooligosaccharides produced by β-agarase from Bacillus cereus ASK 202. J. Fd. Hyg. Safety 15, 277-281.
  5. Jang, H. J., Lee, D. G., Lee, S. W., Jeon, M. J., Chun, W. J., Kwon, K. K., Lee, H. S. and Lee, S. H. 2011. Isolation of a marine-derived Flammeovirga sp. mbrc-1 strain and characterization of its agarase. Kor. Soc. Biotechnol. Bioeng. J. 26, 552-556.
  6. Jeon, M. J., Kim, A. R., Lee, D. G. and Lee, S. H. 2012. Cloning, expression, and characterization of a novel GH-16 β-agarase from Agarivorans sp. JA-1. J. Life Sci. 22, 1545-1551. https://doi.org/10.5352/JLS.2012.22.11.1545
  7. Jung, I. S. 2007. Purification and characterization of agarase produced from marine bacterium, Sphingomonas paucimobilis AS-1 and Aligibacter lectus AS-3. M.S. dissertation. Silla University, Busan, Korea.
  8. Kim, B. J., Song, C. M., Ha, S. D., Hwang, S. H., Kim, H. J., Bae, S. K. and Kong, J. Y. 2000, Physicochemical properties of agarooligosaccharides for using as food stuffs. Kor. J. Food Sci. Technol. 32, 284-290.
  9. Kim, H. T., Lee, S., Lee, D., Kim, H. S., Bang, W. G., Kim, K. H. and Choi, I. G. 2010. Overexpression and molecular characterization of Aga50D from Saccharophagus degradans 2-40: an exo-type beta-agarase producing neoagarobiose. Appl. Microbiol. Bioechnol. 86, 227-234. https://doi.org/10.1007/s00253-009-2256-5
  10. Kim, Y. J. 2010. Properties of the agarase and its gene isolated from a marine bacterium, Tamlana agarivorans. M.S. dissertation. Chungnam National University, Taejon, Korea.
  11. Kim, Y. N. 2011. Isolation and purification of new agarase from marine bacterium. M.S. dissertation. Gyeongsang National University, Jinju, Korea.
  12. Lee, D. G., Jang, M. K., Lee, O. H., Kim, N. Y., Ju, S. A. and Lee, S. H. 2008. Over-production of a glycoside hydrolase family 50 β-agarase from Agarivorans sp. JA-1 in Bacillus subtilis and the whitening effect of its product. Biotechnol. Lett. 30, 911-918. https://doi.org/10.1007/s10529-008-9634-4
  13. Lee, D. G., Kim, N. Y., Jang, M. K., Lee, O. H. and Lee, S. H. 2007. Isolation and characterization of a marine bacterium Thalassomonas sp. SL-5 producing β-agarase. J. Life Sci. 17, 70-75. https://doi.org/10.5352/JLS.2007.17.1.070
  14. Lee, D. G., Lee, O. H., Jang, H. J., Jang, M. K., Yoo, K. H. and Lee, S. H. 2008. Isolation and characterization of a marine derived bacterium Glaciecola sp. SL-12 producing β-agarase. J. Life Sci. 18, 58-62. https://doi.org/10.5352/JLS.2008.18.1.058
  15. Lee, J. H. and Lee, S. Y. 2014. Isolation and characterization of marine bacterial strain SH-1 producing agar-degrading enzymes. Kor. J. Microbiol. Biotechnol. 42, 324-330. https://doi.org/10.4014/kjmb.1410.10009
  16. Lim, D. J., Kim, B. J., Bae, S. K., Kim, J. D. and Kong, J. Y. 1999. Immobilization of agarase for the agarooligosaccharide production. Kor. J. Appl. Microbiol. Biotechnol. 27, 208-214.
  17. Liu, N., Mao, X., Du, Z., Mu, B. and Wei, D. 2014. Cloning and characterization of a novel neoagarotetraose-forming-β-agarase, AgWH50A from Agarivorans gilvus WH0801. Carbo. Res. 388, 147-151. https://doi.org/10.1016/j.carres.2014.02.019
  18. Oh, C., Nikapitiya, C., Lee, Y., Whang, I., Kim, S. J., Kang, D. H. and Lee, J. 2010. Cloning, purification and biochemical characterization of beta agarase from the marine bacterium Pseudoalteromonas sp. AG4. J. Ind. Microbiol. Biotechnol. 37, 483-494. https://doi.org/10.1007/s10295-010-0694-9
  19. Rhee, Y. J., Han, C. R., Kim, W. C., Jun, D. Y., Rhee, I. H. and Kim, Y. H. 2010. Isolation of a novel freshwater agarolytic Cellvibrio sp. KY-YJ-3 and characterization of its extracellular beta-agarase. J. Microbiol. Biotechnol. 20, 1378-1385. https://doi.org/10.4014/jmb.1007.07010
  20. Seok, J. H., Park, H. G., Lee, S. H., Nam, S. W., Jeon, S. J., Kim, J. H. and Kim, Y. H. 2010. High-level secretory expression of recombinant β-agarase from Zobellia galactanivorans in Pichia pastoris. Kor. J. Microbiol. Biotechnol. 38, 40-45.
  21. Shi, Y. L., Lu, X. Z. and Yu, W. G. 2008. A new β-agarase from marine bacterium Janthinobacterium sp. SY12. World J. Microbiol. Biotechnol. 24, 2659-2664. https://doi.org/10.1007/s11274-008-9792-5
  22. Vera, J., Alvarez, R., Murano, E., Slebe, J. C. and Leon, O. 1998. Identification of a marine agarolytic Pseudoalteromonas isolate and characterization of its extracellular agarase. Appl. Environ. Microbiol. 64, 4378-4383.
  23. Yang, J. I., Chen, L. C., Shih, Y. Y., Hsieh, C., Chen, C. Y., Chen, W. M. and Chen, C. C. 2011. Cloning and characterization of β-agarase AgaYT from Flammeovirga yaeyamensis strain YT. J. Biosci. Bioeng. 112, 225-232. https://doi.org/10.1016/j.jbiosc.2011.05.016