Secretory Overexpression of Clostridium Endoglucanase A in Saccharomyces cerevisiae Using GAL10 Promoter and Exoinulinase Signal Sequeice.

Saccharomyces cerevisiae에서 GAL 10 promoter와 exoinulinase 분비 서열을 이용한 Clostridium endoglucanase A의 과발현·분비

  • Lim, Myung-Ye (Department of Biomaterial Control, Dong-Eui University) ;
  • Lee, Jin-Woo (Department of Biotechnology & Bioengineering, Dong-Eui University) ;
  • Lee, Jae-Hyung (Department of Biomaterial Control, Dong-Eui University) ;
  • Kim, Yeon-Hee (Department of Biotechnology & Bioengineering, Dong-Eui University) ;
  • Seo, Jin-Ho (Department of Agricultural Biotechnology, Seoul National University) ;
  • Nam, Soo-Wan (Department of Biomaterial Control, Department of Biotechnology & Bioengineering, Dong-Eui University)
  • 임명예 (동의대학교 바이오물질제어학과) ;
  • 이진우 (동의대학교 생명공학과) ;
  • 이재형 (동의대학교 바이오물질제어학과) ;
  • 김연희 (동의대학교 생명공학과) ;
  • 서진호 (서울대학교 농생명공학부) ;
  • 남수완 (동의대학교 바이오물질제어학과, 생명공학과)
  • Published : 2007.09.30


The secretory overexpression of Clostridium thermocellum endoglucanase A gene (celA) was examined in Saccharomyces cerevisiae using Kluyveromyces marxianus exoinulinase (INU1) signal sequence and GAL10 promoter. The two plasmids, pYEG-CT1 with its own signal sequence, and pYInu-CT1 with INU1 signal sequence were introduced to S. cerevisiae SEY2102 and S. cerevisiae 2805 host strains, respectively, and then each transformant was selected on the synthetic defined media lacking uracil. The expression level and secretion efficiency of endoglucanase A was increased by $18{\sim}22%$ and 11%, respectively, by INU1 signal sequence over celA signal sequence. By considering the high level of expression (361 unit/I), plasmid stability (89%), and secretion efficiency (70%), S. cerevisiae 2805 harboring plasmid pYInu-CT1 was selected as the opti-mal host vector system for the production of cellulose-degrading enzyme and recombinant yeast probiotic. The total expression and secretion efficiency of endoglucanase A was 418 unit/l and 73%, respectively, in the batch fermentation of S. cerevisiae 2805/pYlnu-CT1 on galactose medium. The mo-lecular weight of secreted endoglucanase A was found to be greater than 100 kDa, presumably due to the N-linked glycosylation.


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