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iDEC 2025 | Edinburgh

Development of a High-Throughput Assay for Directed Evolution of Polypropylene-Degrading Enzyme HIS1

The UK produced 197.1 thousand metric tonnes of polypropylene (PP) in 2022; only ~1% of this gets recycled. HIS1 (HPPD Inhibitor Sensitive 1), a Fe(II)/2-oxoglutarate-dependent oxygenase, demonstrated up to 13.55% degradation of PP in optimal conditions, highlighting a potential single-enzyme solution1. We aimed to improve the efficiency of HIS1 through a semi-rational directed evolution approach2, generating a more hydrophobic interface for improved substrate affinity. We tested an array of assays for the detection of PP degradation in the presence of the unmodified HIS1. TBO and Ni-PV assays show robust and reproducible signals; their compatibility with high-throughput workflows was demonstrated through a proof-of-concept positive control test. AFM scans indicated increases in PP surface roughness after purified HIS1 treatment, but additional results provided limited corroborating evidence. Computational analyses indicated seven residues for mutation; library construction was initiated but deferred. By addressing the lack of a high-throughput screen, our research aims to accelerate progress in polymer degradation

  1. TAN, Q. L., CHEN, W. T., LIU, H., YAN, W. D., HUANG, X. & LI, Y. 2024. The programmed sequence-based oxygenase screening for polypropylene degradation. Journal of Hazardous Materials, 465, 9. DOI: 10.1016/j.jhazmat.2023.133173 ↩

  2. DELGADO, J., RADUSKY, L. G., CIANFERONI, D. & SERRANO, L. 2019. FoldX 5.0: working with RNA, small molecules and a new graphical interface. Bioinformatics, 35, 4168-4169. DOI: 10.1093/bioinformatics/btz184 ↩