Journal of Microbiology and Biotechnology

The Korean Society for Microbiology and Biotechnology (한국미생물·생명공학회)
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Screening and Characterization of an Enzyme with ${\beta}-Glucosidase$ Activity from Environmental DNA

  • Kim, Soo-Jin (Microbial Genetics Division, National Institute of Agricultural Biotechnology, Rural Development Administration) ;
  • Lee, Chang-Muk (Microbial Genetics Division, National Institute of Agricultural Biotechnology, Rural Development Administration) ;
  • Kim, Min-Young (Microbial Genetics Division, National Institute of Agricultural Biotechnology, Rural Development Administration) ;
  • Yeo, Yun-Soo (Microbial Genetics Division, National Institute of Agricultural Biotechnology, Rural Development Administration) ;
  • Yoon, Sang-Hong (Microbial Genetics Division, National Institute of Agricultural Biotechnology, Rural Development Administration) ;
  • Kang, Han-Cheol (Cell and Genetics Division, National Institute of Agricultural Biotechnology, Rural Development Administration) ;
  • Koo, Bon-Sung (Microbial Genetics Division, National Institute of Agricultural Biotechnology, Rural Development Administration)
  • Published : 2007.06.30
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Abstract

A novel ${\beta}-glucosidase$ gene, bglA, was isolated from uncultured soil bacteria and characterized. Using genomic libraries constructed from soil DNA, a gene encoding a protein that hydrolyzes a fluorogenic analog of cellulose, 4-methylumbelliferyl ${\beta}-D-cellobioside$ (MUC), was isolated using a microtiter plate assay. The gene, bglA, was sequenced using a shotgun approach, and expressed in E. coli. The deduced 55-kDa amino acid sequence for bglA showed a 56% identity with the family 1 glycosyl hydrolase Chloroflexus aurantiacus. BglA included two conserved family 1 glycosyl hydrolase regions. When using $p-nitrophenyl-{\beta}-D-glucoside$ (pNPG) as the substrate, the maximum activity of the purified ${\beta}-glucosidase$ exhibited at pH 6.5 and $55^{\circ}C$, and was enhanced in the presence of $Mn^{2+}$. The $K_m\;and\;V_{max}$ values for the purified enzyme with pNPG were 0.16 mM and $19.10{\mu}mol/min$, respectively. The purified BglA enzyme hydrolyzed both pNPG and $p-nitrophenyl-{\beta}-D-fucoside$. The enzyme also exhibited substantial glycosyl hydrolase activities with natural glycosyl substrates, such as sophorose, cellobiose, cellotriose, cellotetraose, and cellopentaose, yet low hydrolytic activities with gentiobiose, salicin, and arbutin. Moreover, BglA was able to convert the major ginsenoside $Rb_1$ into the pharmaceutically active minor ginsenoside Rd within 24 h.

Keywords

References

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