Cloning and expression of D-glucoside 3-dehydrogenase from Rhizobium sp. S10 in Escherichia coli and its application for D-gulose production.

The isolated Rhizobium sp. S10 was identified as D-glucoside 3-dehydrogenase (G3DH) producing microbes. Therefore, the gene encoding for G3DH from Rhizobium sp. S10 was cloned and overexpressed in Escherichia coli strain JM109 as a soluble enzyme. The yield flavoenzyme contains 1686 bp of a complete open reading frame and encodes for 561 amino acid residues. The flavin adenine dinucleotide binding region locates in the N-terminal region of G3DH. The recombinant G3DH (rG3DH) was purified with specific activity of 38.54 u/mg and the molecular weight was estimated to be 66 kDa by SDS-PAGE. The purified rG3DH showed highest activity at pH 7.0, 40 °C toward cellobiose. It can also oxidize a broad range of mono-disaccharides including saccharide derivatives. The glycosides oxidizing activity combined with chemical reaction, could produce D-gulose from lactitol via 3-ketolactitol.