Journal of Microbiology and Biotechnology

The Korean Society for Microbiology and Biotechnology (한국미생물·생명공학회)
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Overexpression of Arylsulfatase in E. coli and Its Application to Desulfatation of Agar

  • Lim, Jae-Myung (Department of Biotechnology and Bioengineering, Pukyong National University) ;
  • Jang, Yeon-Hwa (Department of Biomaterial Control, Dong-Eui University) ;
  • Kim, Hyeung-Rak (Division of Food Science and Biotechnology, Pukyong National University) ;
  • Kim, Young-Tae (Department of Microbiology, Pukyong National University) ;
  • Choi, Tae-Jin (Department of Microbiology, Pukyong National University) ;
  • Kim, Joong-Kyun (Department of Biotechnology and Bioengineering, Pukyong National University) ;
  • Nam, Soo-Wan (Department of Biotechnology and Bioengineering, Dong-Eui University)
  • Published : 2004.08.01
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Abstract

The arylsulfatase gene (astA, 984 bp ORF) from the P. carrageenovora genome was amplified by PCR and subcloned into the pET21a vector. When the constructed plasmid pAST-A1 (6.4 kb) was introduced into E. coli BL21(DE3), the transformant on the LB plate containing IPTG showed a hydrolyzing activity for 4-methylumbelliferyl sulfate and p-nitrophenyl sulfate. The highest arylsulfatase activity (2.1 unit/ml) was obtained at 10 mM IPTG. Most arylsulfatase activity was found in the cell lysate, whereas no significant activity was detected in the culture supernatant. The molecular weight of the recombinant enzyme was estimated to be 33.1 kDa by SDS-PAGE. After the reaction of agar with arylsulfatase for 12 h at $40^{\circ}C$, the gel strength of the agar increased by 2-fold, and 73% of the sulfate in the agar had been removed. This result suggests that arylsulfatase expressed in E. coli could be useful in the production of electrophoretic grade agarose.

Keywords

References

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