Most plastic in Europe is landfilled or incinerated, rather than recycled. The main recycling process at large scale for polyethylene terephthalate (PET), via thermomechanical means is limited to clear plastic and results in a loss of mechanical properties each time is recycled. Consequently, de novo synthesis is preferred and PET waste continues to accumulate. In fact, around a half of all PET plastic waste was incinerated in 2018, with around a quarter landfilled and just a quarter recycled, according to Plastics Recyclers Europe 2020. Hence, environment-friendly recycling strategies for solving the ubiquitous pollution of the environment with plastic waste are in high demand.

The general objective of PETzyme is the development of an enzymatic technology for the recycling of PET wastes using free and immobilized PET hydrolases acting synergically and with enhanced thermostability. To achieve this goal, three tasks will be considered.

1. In Task A, different versions of both PETase and MHETase will be cloned individually and expressed in bacteria to obtain free PET hydrolases. Furthermore, the technological platform based on muNS-Mi protein will be used to simultaneously express and encapsulate both individual enzymes, but also the combination of the hydrolases to obtain a chimera nanosphere able to perform the serial reactions for PET depolymerization. The nanoparticles will be purified and characterized. Finally sufficient enzyme will be produced at a bioreactor scale to be used in the next tasks of the project.
2. In Task B the enzymes produced in Task A, both in free form and those immobilized, will be characterized in terms of kinetics, pH and thermal stability, and the possibility of reusing the particles in several cycles. The objective is to obtain enzymes with enhanced thermostability, so this task will be developed in parallel with Task A, and feedback will be provided to improve the cloning and expression system.
3. In Task C the optimal enzymatic cocktail, both free and immobilized, will be applied to decompose different PET materials (different crystallinity percentage, presence of additives, etc). Optimal conditions for the enzymatic transformation will be obtained for three selected materials, and two different reactors will be operated: an enzymatic membrane reactor recovering free PET hydrolases using an ultrafiltration membrane, and a reactor with enzymes immobilized onto NS, where the particles are recovered by centrifugation.

PETzyme presents a multidisciplinary and interdisciplinary research proposal covering the Molecular Biology area (PI2, Prof Jose Martínez Costas) and the Chemical Engineering area (PI1, Dr Gemma Eibes). Both disciplines are necessary to attempt to address the complex and grand challenge of the development of a technology for enzymatic PET waste recycling. The scientific and technical impact of the project is considered high, since a simple and cost-effective process for the simultaneous expression of enzymes with a great interest in the PET value chain will be developed. Finally, finding an effective and environment-friendly strategy for PET waste green recycling will have enormous environmental and social benefits. Increasing the amount of recyclable PET would reduce PET waste ending in landfills. Furthermore, the amount of incinerated PET could be reduced, lowering the spreading of toxic and volatile wastes which are dangerous to the environment and human beings.