Improving enzymatic degradation of unpretreated poly(ethylene terephthalate)

Although the efficiency of poly(ethylene terephthalate) (PET) degradation has been successfully improved by depolymerase engineering, mostly by using Goodfellow-PET (gf-PET) as a substrate, efforts to degrade unpretreated PET materials with high crystallinity remain insufficient. Here, we endeavored to improve the degradation capability of a WCCG mutant of leaf-branch compost cutinase (LCC) on a unpretreated PET substrate (crystallinity > 40%) by employing iterative saturation mutagenesis. Using this method, we developed a high-throughput screening strategy appropriate for unpretreated substrates. Through extensive screening of residues around the substrate-binding groove, two variants, WCCG-sup1 and WCCG-sup2, showed good depolymerization capabilities with both high-(42%) and low-crystallinity (9%) substrates. The WCCG-sup1 variant completely depolymerized a commercial unpretreated PET product in 36 h at 72 degrees C. In addition to enzyme thermostability and catalytic efficiency, the adsorption of enzymes onto substrates plays an important role in PET degradation. This study provides valuable insights into the structure-function relationship of LCC. (c) 2025, Dalian Institute of Chemical Physics, Chinese Academy of Sciences. Published by Elsevier B.V. All rights reserved.