Fisheries and Aquatic Sciences

The Korean Society of Fisheries and Aquatic Science (한국수산과학회)
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Gene Cloning, Expression, and Characterization of Glucose-1-Phosphatase from Enterobacter cloacae B11

  • Kim, Young-Ok (Biotechnology Research Division, National Fisheries Research and Development Institute) ;
  • Park, In-Suk (Biotechnology Research Division, National Fisheries Research and Development Institute) ;
  • Nam, Bo-Hye (Biotechnology Research Division, National Fisheries Research and Development Institute) ;
  • Kong, Hee-Jeong (Biotechnology Research Division, National Fisheries Research and Development Institute) ;
  • Kim, Woo-Jin (Biotechnology Research Division, National Fisheries Research and Development Institute) ;
  • Lee, Sang-Jun (Biotechnology Research Division, National Fisheries Research and Development Institute) ;
  • Kim, Kyung-Kil (Biotechnology Research Division, National Fisheries Research and Development Institute)
  • Received : 2010.01.13
  • Accepted : 2010.03.16
  • Published : 2010.03.31
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Abstract

A bacterial strain with phytase and glucose-1-phosphatase activity was isolated from seawater. The colony was identified as an Enterobacter cloacae strain and named E. cloacae B11. A gene, agpEnB11, coding for an intracellular acid glucose phosphatase was cloned from the strain and sequenced. It comprised 1,242 nucleotides and encoded a polypeptide of 413 amino acids. Recombinant glucose-1-phosphatase (AgpEn) was overexpressed in Escherichia coli and purified using Ni-NTA column under native conditions. Purified protein displayed a single band of 47 kDa on SDS-PAGE. AgpEn hydrolyzed a wide variety of phosphorylated compounds, with high activity for glucose-1-phosphate and glucose-6-phosphate. Optimum pH and temperature for enzyme activity were pH 5.0 and $50^{\circ}C$, respectively. Enzyme activity was stimulated by $Ca^{2+}$ and $Co^{2+}$, and inhibited by $Cu^{2+}$.

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References

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