Expression and engineering of PET-degrading enzymes from Microbispora, Nonomuraea , and Micromonospora .

Low recycling rates coupled with increased production have resulted in an alarming rapid accumulation of a widely used plastic material, polyethylene terephthalate (PET). With the buildup of plastics in our environment, there is an urgent need for more sustainable solutions to process them. Biological methods such as enzyme-catalyzed PET recycling or bioprocessing are potential solutions to this problem. Actinobacteria, known for producing enzymes involved in the degradation of complex organic molecules, are of particular interest due to their potential to produce enzymes that may assimilate plastics. This work expands on the discovery and characterization of new PET-degrading enzymes from the Microbispora, Nonomuraea, and Micromonospora genera. Within the Micromonospora genus, we analyzed enzymes from the polyesterase-lipase-cutinase family with ~60% similarity to leaf-branch compost cutinase (LCC), in which one of the enzymes was found to be capable of breaking down PET and bis(2-hydroxyethyl) terephthalate (BHET) at 45°C-50°C. Moreover, we were able to enhance the enzyme's depolymerization rate through further engineering, resulting in a two-fold increase in activity.IMPORTANCEMismanagement of PET plastic waste significantly threatens human and environmental health. Together with the relentless increase in plastic production, plastic pollution is an issue of rising concern. In response to this challenge, scientists are investigating eco-friendly approaches, such as bioprocessing and microbial factories, to sustainably manage the growing quantity of plastic waste in our ecosystem. Industrial applicability of enzymes capable of degrading PET is limited by numerous factors, including their scarcity in nature. The objective of this study is to enhance our understanding of this group of enzymes by identifying and characterizing novel enzymes that can facilitate the breakdown of PET waste. This data will expand the enzymatic repertoire and provide valuable insights into the prerequisites for successful PET degradation.