MPNN-guided redesign of PET hydrolases with enhanced catalytic activity below the PET glass transition temperature

Enzymatic depolymerization of polyethylene terephthalate (PET) presents a sustainable route for plastic circularity, but its industrial viability is disadvantaged by the need for thermostable enzymes active under mild, energy-efficient conditions. While Polyester Hydrolase Leipzig 7 (PHL7, also known as PES-H1) rapidly degrades amorphous PET near the glass transition temperature of this polymer (~65{degrees}C), its poor protein expression, inactivation above 60{degrees}C and slow depolymerization below 60{degrees}C limits its practical application. Here, we employ ProteinMPNN and LigandMPNN, structural and evolutionary information, to redesign the sequence of PHL7 and improve protein expression, thermostability and activity. We identified 2/36 experimentally tested variants (D5, D11) with enhanced PET depolymerization at 50{degrees}C, achieving the same efficiency as PHL7 at 70{degrees}C but with a shifted product profile, favoring mono-(2-hydroxyethyl) terephthalate (MHET) over terephthalate. Molecular dynamics revealed that these redesigns exhibit enhanced flexibility in active site regions, providing a mechanistic understanding of their low-temperature catalysis. These variants enable a potential route to resynthesize virgin PET via MHET polycondensation, offering an efficient circular economy pathway.