Recent advances in microbial and enzymatic engineering for the biodegradation of micro- and nanoplastics

Abstract

This review examines the escalating issue of plastic pollution, specifically highlighting the detrimental effects on the environment and human health caused by microplastics and nanoplastics. The extensive use of synthetic polymers such as polyethylene (PE), polyethylene terephthalate (PET), and polystyrene (PS) has raised significant environmental concerns because of their long-lasting and non-degradable characteristics. This review delves into the role of enzymatic and microbial strategies in breaking down these polymers, showcasing recent advancements in the field. The intricacies of enzymatic degradation are thoroughly examined, including the effectiveness of enzymes such as PETase and MHETase, as well as the contribution of microbial pathways in breaking down resilient polymers into more benign substances. The paper also discusses the impact of chemical composition on plastic degradation kinetics and emphasizes the need for an approach to managing the environmental impact of synthetic polymers. The review highlights the significance of comprehending the physical characteristics and long-term impacts of micro- and nanoplastics in different ecosystems. Furthermore, it points out the environmental and health consequences of these contaminants, such as their ability to cause cancer and interfere with the endocrine system. The paper emphasizes the need for advanced analytical methods and effective strategies for enzymatic degradation, as well as continued research and development in this area. This review highlights the crucial role of enzymatic and microbial strategies in addressing plastic pollution and proposes methods to create effective and environmentally friendly solutions.

Scheme 1. Schematic diagram of microbial and enzymatic degradation and upcycling of microplastic. Microplastics in the environment undergo enzymatic degradation by extracellular enzymes and are then utilized as a carbon source by microorganisms, ultimately leading to complete. Figures generated with BioRender (https://biorender.com/).

Fig. 1. Enzymatic degradation of various types of plastics. Microorganisms inhabiting various surfaces of microplastics release various types of extracellular enzymes for biodegradation and fragmentation. Figures generated with BioRender (https://biorender.com/).

Fig. 2. Schematic diagram of the increase in microplastic decomposition efficiency using microbial engineering. Microorganisms discovered in various waste treatment facilities are ultimately improved for efficient microplastic degradation through various processes such as protein engineering, metagenomic library techniques, and bioreactor design. Figures generated with BioRender (https://biorender.com/).

Fig. 3. Microplastic decomposition using microbial engineering. (a) Construction of LCC-expressing plasmid pHK-LCC and extracellular expression of active LCC in C. thermocellum. Reproduced with permission from ref. . Copyright (2020) John Wiley & Sons, Inc. (b) Schematic of the designer T–E consortium composed of two strains Pp-T and Pp-E. Pp-T specializes in TPA degradation, which was developed by deleting the ped operon and constitutively expressing the genes tpaAa, tpaAb, tpaB, tpaC, and tpaK. Reproduced with permission from ref. . Copyright (2023) Springer Nature. (c) Co-cultivation of Y. lipolytica Po1fP and P. stutzeri TPA3P. (A) OD, glucose consumption, and PHB content. (B) BHET hydrolysis curve. Reproduced with permission from ref. . Copyright (2021) Elsevier. (d) Biofilm formation and viability. Morphotypes of the cells in the mature biofilm on the PE sheet. Fluorescent microscopic images of biofilms, which show cell viability after the 28 day incubation. Reproduced with permission from ref. . Copyright (2014) American Chemical Society.

Fig. 4. Schematic diagram of protein engineering strategies employed in the modification of enzymes involved in plastic degradation. The incorporation of protein engineering in the enhancement of plastic-degrading enzymes can induce alterations in enzyme activity and characteristics via diverse mechanisms. This encompasses the refinement of thermal stability, augmentation of enzyme activity through electrostatic interactions between the enzyme and substrate, and the potentiation of enzyme activity through the integration of accessory.

Fig. 5. Strategies for enhancing thermal stability and catalytic activity of plastic degrading enzymes using protein engineering. (a) The IsPETase mutant, characterized by increased thermal stability and a higher T_m value compared to the wild-type IsPETase, achieves this improvement through the stabilization of the β6–β7 linked loop. Reproduced with permission from ref. . Copyright (2019) American Chemical Society. (b) Enhanced catalytic activity of a PETase-EKn variant with a more open substrate binding pocket resulting from C-terminal fusion of PETase with a zwitterionic polypeptide consisting of glutamic acid (E) and lysine (K) residues. Reproduced with permission from ref. . Copyright (2021) American Chemical Society. (c) PETase with a narrow active site due to the double mutation S238F/W159H shows a higher loss of crystallinity in PET and improved aromatic interaction with the substrate compared to wild-type PETase. Reproduced with permission from ref. . Copyright (2018) PNAS. (d) Synergistic depolymerization efficacy of MHETase-PETase, a chimeric enzyme linking the C terminus of MHETase to the N terminus of PETase, on PET films. Reproduced with permission from ref. . Copyright (2020) PNAS.

Fig. 6. Proposed mechanism for PET degradation in Ideonella sakaiensis. The extracellular PETase enzyme catalyzes the hydrolysis of PET, yielding MHET as a byproduct. MHETase, predicted to be a lipoprotein, further hydrolyzes MHET into TPA and EG. Both TPA and EG can serve as energy sources for Ideonella sakaiensis and other microbes. Figures generated with BioRender (https://biorender.com/).

Fig. 7. Increased PET decomposition efficiency by PETase deformation. (a) The SEM images (up panel) and water contact angle analysis (down panel) of the PET film in a single-enzyme degradation system, two-enzyme degradation system with ΔBsEst, and a two-enzyme degradation system with ΔChryBHETase after 48 h at 60 °C. Reproduced with permission from ref. . Copyright (2023) Springer Nature. (b) HPLC profiles of PETase powder incubation experiments: 2 weeks, 3 weeks and 4 weeks after incubation. Green and red lines indicate CC-124 wild type and CC-124_PETase #11 lysates. Reproduced with permission from ref. . Copyright (2020) Springer Nature. (c) PETase activity of the variants, PET degradation activity of IsPETaseWT and variants, the enzyme activity is the sum of MHET and TPA. Reproduced with permission from ref. . Copyright (2019) American Chemical Society. (d) Enzyme activity for 10 days and heat-inactivation experiment of IsPETaseWT and IsPETaseS121E/D186H/R280A. Reproduced with permission from ref. . Copyright (2019) American Chemical Society.