Enhancing high-temperature degradation of polyethylene terephthalate through a synergistic division of enzyme labour between a solid-degrading thermostable cutinase and a reaction intermediate-degrading thermostable carboxylesterase

Abstract

Cutinases degrade PET (polyethylene terephthalate) into various degradation intermediates (DIs) such as OET (oligoethylene terephthalate), BHET (bis-hydroxyethyl terephthalate), and MHET (mono-hydroxyethyl terephthalate), and eventually into TPA (terephthalic acid), which is the terminal product of degradation. Unlike PET, which is insoluble, TPA and the DIs are sparingly soluble in water. This causes both DIs and TPA to be partitioned into aqueous solution, where DIs accumulate without undergoing significant further degradation, despite being better substrates of cutinase than solid PET. This frustrates the creation of a circular economy involving PET and TPA (since TPA must be separated from DIs before re-condensation into PET). We argue that the non-degradation of DIs owes to cutinase becoming progressively depleted from solution, through binding to solid PET. This creates a conundrum, in that degradation of PET is anticipated to be inversely correlated with degradation of DIs (at least while solid PET remains available to deplete cutinase from solution), causing any improvement of the cutinases PET-binding efficiency to only further ensure non-degradation of released DIs. Here, we propose the deployment of a second DI-degrading enzyme; one that remains in solution, and acts as an assistant to the master PET-invading cutinase acting upon PETs surface. We demonstrate that one such dual-enzyme system, consisting of a thermostable Thermus thermophilus carboxylesterase (TTCE), characterized here for the first time, and the already-used thermostable leaf-branch compost cutinase (LCC), allows complete degradation of all products of PET hydrolysis into TPA in solution, at 60 {degrees}C, even in the presence of residual solid PET.