Site-Directed Immobilization of DuraPETase onto PET-Binding PDA@SiO2 for High-Efficiency PET Degradation

Plastic pollution caused by poly(ethylene terephthalate) (PET) highlights the urgent need for efficient biodegradation strategies. However, PET hydrolases such as DuraPETase typically exhibit limited substrate affinity for PET and insufficient operational stability. Although conventional immobilization improves enzyme stability, it often compromises catalytic activity. Here, we design a PET-targeting, orientation-controlled immobilization strategy that overcomes this traditional trade-off and enables efficient PET biodegradation. Guided by rational structural analysis, three Cys variants (R53C, R59C, R224C) were engineered for site-specific covalent attachment to a PDA@SiO2 support with inherent PET-binding capability. The resulting conjugates (DuraR53C-PDA@SiO2, DuraR59C-PDA@SiO2, and DuraR224C-PDA@SiO2) displayed distinct catalytic and stability profiles. Among them, DuraR59C-PDA@SiO2 achieved the optimal balance between activity and stability, retaining kinetic properties comparable to the free enzyme and maintaining 87.6% residual activity after 2 h at 80 °C. Water contact angle measurements confirmed its PET-targeting behavior, as evidenced by the reduction in the PET contact angle from 85° to 45°. In 10-day degradation assays at 50 °C, DuraR59C-PDA@SiO2 released a total of 4865.32 μM degradation products, representing a 2.37-fold increase relative to free DuraPETase. These findings demonstrate an effective strategy for industrial enzymatic PET degradation and recycling.