Comparative Biochemistry of Four Polyester (PET) Hydrolases**

The potential of bioprocessing in a circular plastic economy has strongly stimulated research into the enzymatic degradation of different synthetic polymers. Particular interest has been devoted to the commonly used polyester, poly(ethylene terephthalate) (PET), and a number of PET hydrolases have been described. However, a kinetic framework for comparisons of PET hydrolases (or other plastic-degrading enzymes) acting on the insoluble substrate has not been established. Herein, we propose such a framework, which we have tested against kinetic measurements for four PET hydrolases. The analysis provided values of k(cat) and K-M, as well as an apparent specificity constant in the conventional units of M(-1)s(-1). These parameters, together with experimental values for the number of enzyme attack sites on the PET surface, enabled comparative analyses. A variant of the PET hydrolase from Ideonella sakaiensis was the most efficient enzyme at ambient conditions; it relied on a high k(cat) rather than a low K-M. Moreover, both soluble and insoluble PET fragments were consistently hydrolyzed much faster than intact PET. This suggests that interactions between polymer strands slow down PET degradation, whereas the chemical steps of catalysis and the low accessibility associated with solid substrate were less important for the overall rate. Finally, the investigated enzymes showed a remarkable substrate affinity, and reached half the saturation rate on PET when the concentration of attack sites in the suspension was only about 50 nM. We propose that this is linked to nonspecific adsorption, which promotes the nearness of enzyme and attack sites.

Automated summary

Research on enzymatic degradation of synthetic polymers, particularly PET, has been stimulated by the potential of bioprocessing in a circular plastic economy. A kinetic framework for comparisons of PET hydrolases acting on insoluble substrate has been proposed and tested, revealing that a variant of the PET hydrolase from Ideonella sakaiensis is the most efficient enzyme at ambient conditions. Additionally, both soluble and insoluble PET fragments were hydrolyzed faster than intact PET, suggesting that interactions between polymer strands play a role in slowing down PET degradation.