Dos Santos_2024_J.Phys.Chem.B_128_7486

The enzyme PETase fromIdeonella sakaiensis (IsPETase) strain 201-F6 can catalyze the hydrolysis of polyethylene terephthalate (PET), mainly converting it into mono(2-hydroxyethyl) terephthalic acid (MHET). In this study, we used quantum mechanics/molecular mechanics (QM/MM) simulations to explore the molecular details of the catalytic reaction mechanism of IsPETase in the formation of MHET. The QM region was described with AM1d/PhoT and M06-2X/6-31+G(d,p) potential. QM/MM simulations unveil the complete enzymatic PET hydrolysis mechanism and identify two possible reaction pathways for acylation and deacylation steps. The barrier obtained at M06-2X/6-31+G(d,p)/MM potential for the deacylation step corresponds to 20.4 kcal/mol, aligning with the experimental value of 18 kcal/mol. Our findings indicate that deacylation is the rate-limiting step of the process. Furthermore, per-residue interaction energy contributions revealed unfavorable contributions to the transition state of amino acids located at positions 200-230, suggesting potential sites for targeted mutations. These results can contribute to the development of more active and selective enzymes for PET depolymerization.