Laccase immobilized in adamantane-modified PEGMA-based microgels: Tuning enzyme activity by beta-cyclodextrin

Intelligent immobilized enzymes can be constructed by designing "smart" carrier materials. In this study, we successfully prepared adamantane-modified PEGMA-based microgel-immobilized laccases (AdM-Lac) by pre-cipitation polymerization in a laccase solution. AdM-Lac-DMCD was obtained by mixing 2,6-di-O-methyl-beta-cyclodextrin (DMCD) and AdM-Lac. By comparison, DLS, AFM, and IR results indicate that AdM-Lac exhibits DMCD-induced volume phase transition behavior. NMR analysis confirms the successful formation of inclusion complexes of DMCDs and adamantyl groups on AdM-Lac. The catalytic tests show that the enzyme activity of AdM-Lac-DMCD at 20 degrees C (24.09 U/L) is ca.70% higher than AdM-Lac (14.20 U/L). The Vmax value of AdM-Lac-DMCD (1.22 mu M min-1) is similar to AdM-Lac (1.24 mu M min-1), Whereas the Km value (52 mu M) decreases by ca. 69%, even close to free laccase (41 mu M), and the Vmax/Km ratio (0.0235 min-1) increases by about 2-fold. These results imply that DMCD initiates the volume swelling of AdM-Lac, resulting in increased enzyme activity and improved kinetic properties. Due to AdM-Lac having better storage stability, we suggest that laccase is stored as AdM-Lac, and applied as AdM-Lac-DMCD. This study successfully achieved the regulation of enzyme activity through the addition of molecules, providing a novel strategy for the design of "smart" carrier materials.