Influence of ionic liquid cosolvent on transgalactosylation reactions catalyzed by thermostable β-glycosylhydrolase celB from Pyrococcus furiosus

The synthesis of glycosides by enzymatic transglycosylation is a kinetically controlled reaction performed in the context of a non-favorable thermodynamic equilibrium. An unreactive organic cosolvent which increases the selectivity of the enzyme for glycosyl transfer to the acceptor nucleophile compared with water (K-sel) could improve maximum product yield. Here we report on the effect of the ionic liquid 1,3-dimethylimidazoliummethylsulfate on hydrolase and transferase activities of the hyperthermostable beta-glycosidase COB from the archaeon Pyrococcus furiosus. CelB retained full catalytic efficiency for lactose hydrolysis at 80 degrees C in a 50% (by vol.) solution of ionic liquid in sodium citrate buffer, pH 5.5. It was inactive but not irreversibly denatured a 70% ionic liquid. Using lactose (0.15 M) as galactosyl donor, values of K-sel for a representative series of eight acceptor alcohols were determined in kinetic assays at 80 degrees C and found to increase between 1.3-fold (D-Xylose) and 3.1-fold (glycerol) in 45% ionic liquid. Enhancement of K-sel was dependent on ionic liquid concentration and higher than expected from the decrease in water activity caused by the cosolvent. Experimental molar ratios of D-glucose and D-galactose produced during enzymatic conversion of lactose (75-150 mM) in the presence of D-Xylose (0.5 M) or glycerol (0.5 M) showed excellent agreement with predictions based on K-sel values and confirm a significant, yet moderate effect of 45% ionic liquid on increasing the yield Of D-galactoside product, by <= 10%. (c) 2006 Wiley Periodicals, Inc.