Characterization of Carboxymethylcellulase(CMCase) Produced by Recombinant E. coli Containing CMCase Gene for Cellulomonas sp. YE-5

  • Park, Sung-Won (Dept. of Food and Biotechnology, College of Engineering and Bioproducts Research Center, Yonsei University) ;
  • Her, Nam-Yun (Dept. of Foodculinar, Osan Junior College) ;
  • Kim, Dong-Seob (Dept. of Food Science, Miryang National Univeristy) ;
  • Park, Sun-Jin (Dept. of ood and Biotechnology, College of Engineering and Bioproducts Research Center, Yonsei University) ;
  • Lee, Han-Seung (Dept. of Food and Biotechnology, College of Engineering and Bioproducts Research Center, Yonsei University) ;
  • Park, Hak-Jong (Dept. of Food and Biotechnology, College of Engineering and Bioproducts Research Center, Yonsei University) ;
  • Yu, Ju-Hyun (Dept. of Food and Biotechnology, College of Engineering and Bioproducts Research Center, Yonsei University)
  • Published : 1997.06.01

Abstract

CMCase produced by recombinant E. coli JM109 (pCEH#4) containing CMCase gene from Cellulomonas sp. YE-5 was purified to 24.3 fold and 2.6% yield by ammoniumsulfate precipitation, DEAE-cellulose column chromatography and gel filtration on Sephadex G-100. The optimum pH and temperature for CMCase activity were pH 7.0 and 5$0^{\circ}C$. The enzyme was stable between pH 5.0 and 10.0, and up to 6$0^{\circ}C$. The molecular weight of he enzyme was estimated to be approximately 40,000 daltons by SDS-PAGE. Analysis of the amino acid composition showed that the enzyme contained many glycines and acidic amino acids. The enzyme was an endo-type CMCase and the final enzyme reaction product from hydrolysis of Cm-cellulose by the enzyme was cellobiose. {TEX}$K_{M}${/TEX} value determined with CM-cellulose was 1.28mM.