Microbiology and Biotechnology Letters (한국미생물·생명공학회지)

The Korean Society for Microbiology and Biotechnology (한국미생물·생명공학회)
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Properties of a Novel Clostridiclm thermocellum Endo-$\beta$-1,4-glucanase Expressed in Escherichia coli

대장균에서 발현되는 Clostridium thermocellum의 섬유소 분해 효소의 특성

  • 정경화 (한국과학기술원 생물공학과) ;
  • 이진호 (한국과학기술원 생물공학과) ;
  • 이용택 (한국과학기술원 생물공학과) ;
  • 김하근 (배재대학교 유전공학과) ;
  • 박무영 (한국과학기술원 생물공학과)
  • Published : 1992.10.01
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Abstract

An endo-$\beta$-1,4-glucanase gene of Clostridium thermocellum was cloned in Escherichia coli and was considered as a novel gene by comparison with the restriction patterns of the C. thermocellum cellulase genes so far reported. The endoglucanase from recombinant E. coli was purified by column chromatography after heat treatment. The purified enzyme was a monomer having molecular weight of 40,000. The enzyme hydrolyzed CMC to glucose and cello-oligosaccharides at :naximum activities at pH 5.0 and $65^{\circ}C$. One of the endproducts, glucose, showed no inhibitory effect on the enzyme activity, while the other endproduct, cellobiose, inhibited slightly. The values of $K_{m}$ and $V_{max}$ of the enzyme for CMC were 0.39% (w/v) and 268 Ulmg protein, respectively.

고온성 혐기성 세균인 Clostridium thermocellum의 섬유소 분해 효소 유전자를 pUC9 플라스미드를 이용하여 대장균에 클로닝하였고, 지금까지 클로닝 된 C.thermocellum의 섬유소 분해 유전자들과 제한효소 양상을 비교하여 새로운 유전자임을 알 수 있었다. 대장균에서 섬유소 분해 효소를 열처리와 column chromatography에 의해서 정제를 하였고, 분자량은 40, 000이었다. 이 효소는 pH 5.0과 $65^{\circ}C$에서 CMC에 대해서 최대 활성을 보였고 최종 산물인 포도당과 cellobiose에 의한 활성의 저해는 크게 나타나지 않았다. CMC에 대한 이 효소의 $K_{m}$ 과 $V_{max}$값은 각각 0.39(w/v)와 268 U/mg protein이었다.

Keywords

References

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