Journal of Microbiology and Biotechnology

The Korean Society for Microbiology and Biotechnology (한국미생물·생명공학회)
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Biochemical Characterization of a GDSL-Motif Esterase from Bacillus sp. K91 with a New Putative Catalytic Mechanism

  • Ding, Junmei (Engineering Research Center of Sustainable Development and Utilization of Biomass Energy, Ministry of Education, Yunnan Normal University) ;
  • Yu, Tingting (Engineering Research Center of Sustainable Development and Utilization of Biomass Energy, Ministry of Education, Yunnan Normal University) ;
  • Liang, Lianming (Laboratory for Conservation and Utilization of Bio-Resources, and Key Laboratory for Microbial Resources of the Ministry of Education, Yunnan University) ;
  • Xie, Zhenrong (Engineering Research Center of Sustainable Development and Utilization of Biomass Energy, Ministry of Education, Yunnan Normal University) ;
  • Yang, Yunjuan (Engineering Research Center of Sustainable Development and Utilization of Biomass Energy, Ministry of Education, Yunnan Normal University) ;
  • Zhou, Junpei (Engineering Research Center of Sustainable Development and Utilization of Biomass Energy, Ministry of Education, Yunnan Normal University) ;
  • Xu, Bo (Engineering Research Center of Sustainable Development and Utilization of Biomass Energy, Ministry of Education, Yunnan Normal University) ;
  • Li, Junjun (Engineering Research Center of Sustainable Development and Utilization of Biomass Energy, Ministry of Education, Yunnan Normal University) ;
  • Huang, Zunxi (Engineering Research Center of Sustainable Development and Utilization of Biomass Energy, Ministry of Education, Yunnan Normal University)
  • Received : 2014.06.19
  • Accepted : 2014.07.21
  • Published : 2014.11.28
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Abstract

The esterase gene Est8 from the thermophilic bacterium Bacillus sp. K91 was cloned and expressed in Escherichia coli. The monomeric enzyme exhibited a theoretical molecular mass of 24.5 kDa and an optimal activity around $50^{\circ}C$ at pH 9.0. A model of Est8 was constructed using a hypothetical YxiM precursor structure (2O14_A) from Bacillus subtilis as template. The structure showed an ${\alpha}/{\beta}$-hydrolase fold and indicated the presence of a typical catalytic triad consisting of Ser-11, Asp-182, and His-185, which were investigated by site-directed replacements coupled with kinetic characterization. Asp-182 and His-185 residues were more critical than the Ser-11 residue in the catalytic activity of Est8. A comparison of the amino acid sequence showed that Est8 could be grouped into the GDSL family and further classified as an SGNH hydrolase. Est8 is a new member of the SGNH hydrolase subfamily and may employ a different catalytic mechanism.

Keywords

References

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