Journal of Microbiology and Biotechnology

The Korean Society for Microbiology and Biotechnology (한국미생물·생명공학회)
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Expression, Purification, and Crystallization of D-Psicose 3-Epimerase from Agrobacterium tumefaciens

  • Kim Kwang-Soo (School of Agricultural Biotechnology and Center for Agricultural Biomaterials, Seoul National University) ;
  • Kim Hye-Jung (Department of Molecular Biotechnology, Konkuk University) ;
  • Oh Deok-Kun (Department of Molecular Biotechnology, Konkuk University) ;
  • Cheong Jong-Joo (School of Agricultural Biotechnology and Center for Agricultural Biomaterials, Seoul National University) ;
  • Rhee Sang-Kee (School of Agricultural Biotechnology and Center for Agricultural Biomaterials, Seoul National University)
  • Published : 2006.04.01
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Abstract

D-Psicose 3-epimerase (DPE) catalyzes the interconversion of D-fructose to D-psicose by epimerizing the carbon-3 position. The DPE from Agrobacterium tumefaciens was cloned and expressed in Escherichia coli. The expressed enzyme was purified by affinity chromatography on an IMAC, gel filtration chromatography on a Sephacryl S-300 HR, and anion-exchange chromatography on a RESOURCE Q. The molecular mass of the purified enzyme was estimated to be about 135 kDa by Superdex 200 gel filtration chromatography, corresponding to a homotetramer. The enzyme produced crystals suitable for X-ray diffraction to a $2.0{\AA}$ resolution at 100 K. The crystals were found to belong to the orthorhombic space group $P2_12_12_1$, with unit-cell parameters a=102.4, b=113.0, and $c=131.8{\AA}$. In addition, the calculated packing parameter $(V_m)$ was $2.79{\AA}^3/Da$, the solvent content was 55.92%, and an asymmetric unit consisted of four monomers.

Keywords

References

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