Effects of Structural Difference of Ionic Liquids on the Catalysis of Horseradish Peroxidase

The dependence of the catalytic properties of horseradish peroxidase on the structural changes of ionic liquids was investigated with two water-miscible ionic liquids, N-butyl-3methypyridinium tetraftuoroborate ([$BMP_y$][$BF_4$]) and 1-butyl-3-methylimidazolium methylsulfate ([BMIM][$MeSO_4$]), each of which shares an anion ($BF_4^-$) or a cation ($BMIM^+$) with 1-butyl-3-methylimidazolium tetraftuoroborate ([BMIM][$BF_4$]), respectively. The oxidation of guaiacol (2-methoxyphenol) with $H_2O_2$was used as a model reaction. In order to minimize the effect of solution viscosity on the kinetic constants of the enzymatic catalysis, the enzymatic reactions for the kinetic study were performed in water-ionic liquid mixtures containing 25% (v/v) ionic liquid at maximum. Similarly to the previously reported results for [BMIM][$BF_4$], as the concentration of [$BMP_y$][$BF_4$] increased, the $K_m$value increased with a decrease in the $k_{cat}$value: the $K_m$value increased markedly from 2.8 mM in 100% water to 12.6 mM in 25% (v/v) ionic liquid, indicating that ionic liquid significantly weakens the binding affinity of guaiacol to the enzyme. On the contrary, [BMIM][$MeSO_4$] decreased the Km value to 1.4 mM in 25% (v/v) ionic liquid. [BMIM][$MeSO_4$] also decreased $k_{cat}$more than 3-folds [from 13.8 $s^{-1}$in 100% water to 4.1 $s^{-1}$in 25% (v/v) ionic liquid]. These results indicate that the ionic liquids interact with the enzyme at the molecular level as well as at a macroscopic thermodynamic scale. Specifically, the anionic component of the ionic liquids influenced the catalysis of horseradish peroxidase in different ways.