Biocatalytic action of proteases in ionic liquids: Improvements on their enzymatic activity, thermal stability and kinetic parameters

This study evaluated the effect of the addition of the following ionic liquids (IL): choline chloride (CC), tetramethylammonium bromide (TB) and 1‑ethyl‑3‑methylimidazolium bromide (EM), on some biochemical properties including enzymatic activity and different kinetic parameters of commercial proteases. The enzyme-IL combinations that showed the highest increases in enzyme activities were as follows: CC (0.5 mM) and Neutrase® 0.8 L; CC (5 mM) and Flavourzyme® 500 L; TB (2000 mM) and Alcalase® 2.4 L, with relative increases of 20, 15 and 150% in protease activities, respectively, compared to the control assays. The combination TB and Alcalase® 2.4 L showed a reduction of 50% of the activation energy (Ea), an increase of the relation Vmax/Km of 35% and a 16-fold rise in the values of t1/2, and D. Neutrase® 0.8 L combined with CC showed an increase of 20% in the relation Vmax/Km. The combination Flavourzyme® 500 L and CC presented a 20% higher value of the relation Vmax/Km and a 2-fold increase in the values of t1/2 and D compared to the control assay. In summary, the most positive effects observed in this study included proteases with improved activity and stability properties and a greater affinity for the substrate.