Influence of Diol-Based Deep Eutectic Solvents on Laccase Enzyme Activity and Stability

In pursuit of sustainable alternatives to conventional catalysts, laccase, a multicopper oxidoreductase enzyme, has gained significant attention due to its ability to oxidize a broad range of compounds using molecular oxygen. Its natural occurrence in fungi, plants, bacteria, and insects, along with its applicability in bioremediation, wastewater treatment, organic synthesis, and various industrial processes, makes it a valuable biocatalyst. However, laccase's stability and activity are often compromised under industrial conditions such as high temperature, variable pH, and the presence of inhibitors. Recent advancements have explored enzyme engineering, immobilization, and solvent engineering to address these limitations, but these methods often involve complex, costly, or unsustainable procedures. In this search, deep eutectic solvents (DESs), have emerged as a promising green solvents to enhance laccase performance. Studies indicate that polyol-based DESs significantly improve both the activity and thermal stability of laccase. Despite this, the specific influence of polyol structure particularly the number and position of hydroxyl groups remains unclear. This study investigates 15 betaine- and choline chloride-based DESs containing three different diols to elucidate their effect on laccase activity and stability. The findings indicate that only betaine-derived diol-based deep eutectic solvents (DESs) enhance both the activity and stability of laccase. In particular, DESs with a 1:4 molar ratio significantly improves laccase activity. Among them, Bet:1,2-Pro DES exhibit higher laccase activity than Bet:1,3-Pro DES. This suggests that the number and position of hydroxyl groups in the polyols play a crucial role in modulating laccase functionality. Additionally, molecular docking studies support the idea that hydrogen bonding between DES components and amino acids in the enzyme's catalytic cluster contributes to enhanced activity. Overall, these results provide valuable insights for designing effective DESs to improve laccase biocatalysis and expand its practical applications.