Characterization of a recombinant cellobiose 2-epimerase from Caldicellulosiruptor saccharolyticus and its application in the production of mannose from glucose.
Chang-Su Park, Jung-Eun Kim, Jin-Geun Choi, Deok-Kun Oh
A putative N-acyl-D-glucosamine 2-epimerase from Caldicellulosiruptor saccharolyticus was cloned and expressed in Escherichia coli. The recombinant enzyme was identified as a cellobiose 2-epimerase by the analysis of the activity for substrates, acid-hydrolyzed products, and amino acid sequence. The cellobiose 2-epimerase was purified with a specific activity of 35 nmol min(-1) mg(-1) for D-glucose with a 47-kDa monomer. The epimerization activity for D-glucose was maximal at pH 7.5 and 75°C. The half-lives of the enzyme at 60°C, 65°C, 70°C, 75°C, and 80°C were 142, 71, 35, 18, and 4.6 h, respectively. The enzyme catalyzed the epimerization reactions of the aldoses harboring hydroxyl groups oriented in the right-hand configuration at the C2 position and the left-hand configuration at the C3 position, such as D-glucose, D-xylose, L-altrose, L-idose, and L-arabinose, to their C2 epimers, such as D-mannose, D-lyxose, L-allose, L-gulose, and L-ribose, respectively. The enzyme catalyzed also the isomerization reactions. The enzyme exhibited the highest activity for mannose among monosaccharides. Thus, mannose at 75 g l(-1) and fructose at 47.5 g l(-1) were produced from 500 g l(-1) glucose at pH 7.5 and 75°C over 3 h by the enzyme.
