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KMID : 0545120040140040777
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Journal of Microbiology and Biotechnology
2004 Volume.14 No. 4 p.777 ~ p.782
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Overexpression of Arylsulfatase in E. coli and Its Application to Desulfatation of Agar
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Lim JM
Jang YH/Kim HR/Kim YT/Choi TJ/Kim JK/Nam SW
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Abstract
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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 40oC 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.
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