Unlocking the Potential for Microplastic Degradation: In Silico Mutagenesis andMolecular Dynamics Simulations of PET-PETase and MHET-MHETase Interactions by Reverse Screening
DOI:
https://doi.org/10.54536/ajbb.v5i1.7744Keywords:
Active Site Mutations, Computational Docking, Microplastic Biodegradation, Protein Engineering, Protein-Ligand InteractionAbstract
Plastic pollution remains a growing global concern, requiring efficient and sustainable solutions. Enzymes such as PETase and MHETase from Ideonella sakaiensis offer natural mechanisms for plastic degradation but need further enhancement for large-scale applications. This study aimed to improve the structural stability and substrate-binding performance of PETase and MHETase by engineering their homologous variants through in silico methods. Homologous proteins were identified via multiple sequence alignment, and functionally relevant residues near the active site were selected for site-saturation mutagenesis. The resulting variants were evaluated through molecular docking to assess binding energy and interaction profiles, while molecular dynamics (MD) simulations examined their behavior over time using RMSD, and RMSF. Among the MHETase variants, GLY103ALA (–9.5 ± 0.2 kcal/mol), PRO44PHE (–8.4 ± 0.1 kcal/mol), and GLY165HIS (–8.2 ± 0.1 kcal/mol) showed significantly enhanced ligand-binding affinity compared to wild type (–6.3 ± 0.1 kcal/mol). These variants also showed improved dynamic stability, as indicated by reduced ligand RMSD values (GLY103ALA: 0.22 ± 0.01 nm; PRO44PHE: 0.24 ± 0.01 nm; GLY165HIS: 0.25 ± 0.01 nm) compared to the wild type (0.36 ± 0.02 nm), along with stable backbone RMSF values (~0.10 ± 0.01 nm) relative to the wild type (0.18 ± 0.02 nm). For PETase, the GLY35GLN variant toexhibited the most favorable performance, with a lower binding energy (–6.0 ± 0.1 kcal/mol) than to the wild type (–5.2 ± 0.1 kcal/mol). It also exhibited the lowest ligand RMSD (0.28 ± 0.02 nm) compared to wild type (0.41 ± 0.03 nm), and showed reduced structural fluctuation in RMSF (0.20 ± 0.01 nm) relative to wild type (0.26 ± 0.02 nm), indicating enhanced dynamic stability. These results suggest that even minor mutations can significantly improve enzymatic efficiency in microplastic (MP) degradation. Overall, the results show that in silico mutagenesis and simulation can help identify enzyme variants with improved binding and stability, making them good candidates for future lab testing and potential use in breaking down microplastic waste.
Downloads
References
Chen, S., Su, L., Chen, J., Wu, J., & Chen, G. Q. (2023). Protein engineering of PETase from Ideonella sakaiensis to enhance PET degradation via rational design. Biotechnology Advances, 64, 108108. https://doi.org/10.1016/j.biotechadv.2023.108108
Charupanit, K., Tipmanee, V., Sutthibutpong, T., & Limsakul, P. (2022). In silico identification of potential sites for a plastic-degrading enzyme by a reverse screening through the protein sequence space and molecular dynamics simulations. Molecules, 27(10), 3353. https://doi.org/10.3390/molecules27103353
Danso, D., Schmeisser, C., Chow, J., Zimmermann, W., Wei, R., Leggewie, C., Li, X., Hazen, T., & Streit, W. R. (2018). New insights into the function and global distribution of polyethylene terephthalate (PET)-degrading bacteria and enzymes in marine and terrestrial metagenomes. Applied and Environmental Microbiology, 84(8), e02773-17. https://doi.org/10.1128/AEM.02773-17
Duru, C. E., Duru, I. A., & Enyoh, C. E. (2021). In silico binding affinity analysis of microplastic compounds on PET hydrolase enzyme target of Ideonella sakaiensis. Bulletin of the National Research Centre, 45(1), 35. https://doi.org/10.1186/s42269-021-00563-5
Graf, N., Altenbuchner, J., & Groll, M. (2021). Structural and functional characterization of MHETase variants to improve enzymatic PET degradation. Nature Communications, 12(1), 1–12. https://doi.org/10.1038/s41467-021-21999-2
Jambeck, J. R., Geyer, R., Wilcox, C., Siegler, T. R., Perryman, M., Andrady, A., Narayan, R., & Law, K. L. (2015). Plastic waste inputs from land into the ocean. Science, 347(6223), 768–771. https://doi.org/10.1126/science.1260352
Maity, J. P., Chen, C. Y., & Chen, C. A. (2021). A review of biocatalysts and bioreactors for PET degradation. Environmental Pollution, 273, 116513. https://doi.org/10.1016/j.envpol.2020.116513
Raza, M., Lee, J.-Y., & Cha, J. (2022). Microplastics in soil and freshwater: Understanding sources, distribution, potential impacts, and regulations for management. Science Progress, 105(3), 00368504221126676. https://doi.org/10.1177/00368504221126676
Rorrer, N. A., Kearns, F. L., Silveira, R. L., & Beckham, G. T. (2018). Characterization and engineering of a plastic-degrading aromatic polyesterase. Proceedings of the National Academy of Sciences, 115(19), E4350–E4357. https://doi.org/10.1073/pnas.1718804115
Wei, R., & Zimmermann, W. (2017). Biocatalysis as a green method for polyester waste recycling. Trends in Biotechnology, 35(6), 501–510. https://doi.org/10.1016/j.tibtech.2017.01.009
Yoshida, S., Hiraga, K., Takehana, T., Taniguchi, I., Yamaji, H., Maeda, Y., Toyohara, K., Miyamoto, K., Kimura, Y., & Oda, K. (2016). A bacterium that degrades and assimilates poly(ethylene terephthalate). Science, 351(6278), 1196–1200. https://doi.org/10.1126/science.aad6359
Downloads
Published
Issue
Section
License
Copyright (c) 2026 Mia Joy Inocencio
This work is licensed under a Creative Commons Attribution 4.0 International License.
