KSBB Journal

  • 제19권4호
  • /
  • Pages.301-307
  • /
  • 2004
  • /
  • 1225-7117(pISSN)
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  • 2288-8268(eISSN)
한국생물공학회 (The Korean Society for Biotechnology and Bioengineering)
권호 표지

시종 누룩사상균, Aspergillus coreanus NR 15-1의 a-Amylase의 효소학적 특성

Characteristics of a-Amylase of, a New Species, Aspergillus coreanus NR 15-1

  • 이상훈 (계명대학교 미생물학과) ;
  • 정혁준 (계명대학교 미생물학과) ;
  • 여수환 (계명대학교 전통미생물 자원 개발 및 산업화 연구센터) ;
  • 김현수 (계명대학교 미생물학과) ;
  • 유대식 (계명대학교 미생물학과)
  • 발행 : 2004.08.01
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초록

한국 전통누룩으로부터 분리한 신종 Aspergillus coreanus NR 15-1가 생산하는 a-amylase의 효소학적 특성을 조사했다. 공시균주가 생산하는 a-amylase는 황산암모늄을 이용한 분별 침전, CM-cellulose, DEAE-cellulose, Sephadex G-100, hydroxyapatite column chromatography를 통하여 8.7%의 수율을 보이며, 78배로 정제되었다. 공시균주의 a-amylase의 분자량은 Sephadex G-100 겔 여과에 의해 49 kDa으로 나타났으며, SDS-PAGE에 의하여 51 kDa으로 측정되어 본 효소는 monomer로 추정할 수 있었다. 정제효소는 pH 4.0∼11.0 사이에서 안정하였으며, 반응최적 pH는 5.0이었고, 5$0^{\circ}C$ 이하의 온도에서 비교적 안정하며, 반응최적온도는 45$^{\circ}C$로 나타났다. 정제효소는 금속이온에 의해 효소활성에 영향을 받지 않았으나, N-bromosuccinimide에 의해서는 효소활성이 완전히 저해되어 본 공시균주의 a-amylase의 활성부위에는 tryptophan 잔기가 관여한다고 추정할 수 있었다. 정제효소의 전분분해물은 maltose, maltotriose 등의 oligosaccharide를 형성하므로 a-amylase임을 확인할 수 있었다. 신종 누룩시상균인 Aspergillus coreanus NR 15-1의 a-amylase는 5$0^{\circ}C$ 이하의 온도와 pH 4.0∼11.0사이에서 안정하여 온도와 pH의 안정성이 우수하여 누룩제조용 사상균으로 사용이 가능함을 알 수 있었다.

The characteristics of the a-amylase of Aspergillus coreanus NR 15-1 isolated from traditional Korean Nuruk have been carried out. The a-amylase of A. coreanus NR 15-1 was purified by ammonium sulfate precipitation followed by column chromatographies on CM-cellulose, DEAE-cellulose, Sephadex G-100 gel filtration and hydroxyapatite. The a-amylase was purified 78-fold with a yield of 8.7%. The molecular weight of the a-amylase was estimated to be 49 kDa by Sephadex G-100 gel filtration and 51 kDa by SDS-polyacrylamide gel eletrophoresis. These experimental results suggested that the purified enzyme might be monomer. The enzyme was stable between pH 4 and 11. The optimum pH was 5.0. The optimum temperature for enzyme was 45$^{\circ}C$ and the enzyme was stable up to 50$^{\circ}C$. The enzyme was significantly inhibited by 1 mM N-bromosuccinimide. These results suggested that tryptophan residue was involved in the active site of a-amylase. The enzyme was identified as a-amylase because the reaction products of soluble starch hydrolyzed by the purified enzyme was oligosaccharide by thin layer chromatography.

키워드

참고문헌

  1. Yakugaku Zasshi v.277 Report of examination of Corean "Koji" (I) Ueno, K.
  2. J. Brew. Soc. Papan. v.85 Absidia sp. in the Chinese starter(Nuruk) in Korean, microorganisms in Chinese starter from Asia (part 3) Uchimura, T.;S. Takaki;K. Watanabe;M. Kozaki https://doi.org/10.6013/jbrewsocjapan1988.85.888
  3. Kor. J. Appl. Microbiol. Bioeng. v.13 Studies on digestion of raw starch by Rhizopus oryzae Kim, C. J.;M. J. Oh;J. S. Lee
  4. Kor. Biochem. J. v.13 Study on the ${\alpha}$-amylase activity in Aspergillus oryzae Kim, T. Y.;S. J. Kim
  5. Kor. J. Food Sci. Technol. v.18 Purification and characterization of raw starch digesting enzyme from Rhizopus oryzae Kim, C. J.;M. J. Oh;J. S. Lee
  6. Kor. J. Appl. Microbiol. Biotechnol. v.18 Studies on the development and the characteristics of the powerful raw starch digesting enzyme Chung, M. J.;W. N. Hou;J. H. Jeong;H. Taniguchi
  7. J. Microbiol. v.37 Purification and characteristics of glucoamylase in Aspergillus oryzae NR 3-6 isolated traditional Korean Nuruk Yu, T. S.;T. H. Kim;C. Y. Joo
  8. J. Microbiol. Biotechnol. v.14 Characteristics of two forms of glucoamylase from traditional Korean Nuruk fungi, Aspergillus coreanus NR 15-1 Han, Y. J.;T. S. Yu
  9. J. Microbiol. Biotechnol. v.14 A new species of Hyphomycetes, Aspergillus coreanus sp. nov., isolated from traditional Korean Nuruk Yu, T. S.;S. H. Yeo;H. S. Kim
  10. The annotation of the official method of analysis of the National Tax Administration Agency(4th ed.) The Brewing Society of Japan
  11. Anal. Biochem. v.72 A rapid and sensitive method for the quantitation of protein utilizing the principle of protein-dye binding Bradford, M. M. https://doi.org/10.1016/0003-2697(76)90527-3
  12. J. Biochem. v.91 Estimation of the molecular weights of proteins by Sephadex gel filtration Andrews, P. https://doi.org/10.1042/bj0910222
  13. Nature(London) v.227 Cleavage of structural proteins during the assembly of the head of bacteriophage $T_4$ Laemmli, U. K. https://doi.org/10.1038/227680a0
  14. J. Chromator. v.105 Thin-layer chromatographic method for identification of oligosaccharides in starch hydrolysates Hansen, S. A. https://doi.org/10.1016/S0021-9673(01)82270-6
  15. Can. J. Microbiol. v.31 Purification and some properties of the extracellular ${\alpha}$-amylase from Aspergillus awamori Bhella, R. S.;I. Altosaar https://doi.org/10.1139/m85-029
  16. Agric. Biol. Chem. v.32 Acid stable ${\alpha}$-amylase of black Aspergilli. Part IV. Some physicochemical properties Arai, M.;T. Koyano;H. Ozawa;K. Yamada https://doi.org/10.1271/bbb1961.32.507
  17. Kor. J. Mycol. v.17 Purification and characterization of acid-stable ${\alpha}$-amylase of Aspergillus niger K-25 Cho, M. H.
  18. Kor. J. Appl. Microbiol. Biotechnol. v.27 Purification and characteristics of amylase from Haloarcular sp. EH-1 Jung, M. J.;H. S. Park
  19. Kor. J. Appl. Microbiol. Biotechnol. v.25 Purification and characteristics of raw starch hydrolyzing enzyme from Aspergillus niger Jung, M. J.
  20. Kor J. Appl. Microbiol. Biotechnol. v.21 The purification and properties of Schwannipmyces castellii CBS 2863 Park, J. C.;S. Bai;S. Y. Lim;H. Lee;S. B. Chun
  21. Agric. Biol. Chem. v.46 Characterization of a potato starch-digesting bacterium and its production of amylase Taniguch, H.;F. Ogashima;M. Igarashi;Y. Maruyama;M. Nakamura https://doi.org/10.1271/bbb1961.46.2107