The Biological Degradation of Polyethylene Terephthalate: Using Saccharomyces cerevisiae as a Chassis for PET Hydrolase Delivery

Plastic pollution has become a significant global issue, with waste plastic materials documented across various ecosystems. As current recycling efforts continue to fall short of addressing the volume of plastic waste being generated, enzymes capable of breaking down plastic materials into their original building blocks are being discovered at an increasing rate. These enzymes are currently applied to PET plastic through purified systems, which are both costly and time-consuming. A significant portion of research efforts have focused on the application of the novel bacterial enzyme IsPETase. This study focuses on the development of a yeast biofilm-based delivery system for PET hydrolases. The expression and activity of IsPETase in biofilm-forming (Ʃ1278b) and non-biofilm-forming (BY4741) S. cerevisiae strains were assessed. Overall elevated levels of degradation, further verified through AFM and SEM analysis, were observed in Ʃ1278b cultures. Roughness values (Rq) supported the microscopy, with Ʃ1278b samples having an Rq value of 33.6 nm compared to 5.4 nm in the control and 26.7 nm in BY4741. Subsequent optimisation, achieved by reducing glucose concentrations to 0.5%, enhanced biofilm formation, protein secretion, and PET degradation in the non-biofilm-forming strain BY4741. Rq in the 0.5% glucose cultures increased to 50.4 nm. These findings confirmed that increased degradation was associated with higher cell surface adhesion. Beyond IsPETase expression, seven further PET hydrolases were tested, representing thermostable variants and novel plastic-degrading enzymes. Three strains of S. cerevisiae were genetically modified, and activity showed varying results between strain and PET hydrolase. Integration of the LCCICCG, an alternative PET hydrolase, presented increased protein secretion and higher surface disruption, with Rq values in the LCCICCG mutant reaching 120.7 nm. Additionally, two unconventional yeast species, Candida intermedia and Rhodotorula diobovata, were identified as promising expression systems due to their low health risks and biofilm formation capabilities. This study provides a valuable foundation for enhancing PET degradation using biofilm-based systems. With further development, this approach holds potential for creating robust circular recycling systems to mitigate plastic waste.