Genomic and enzymatic analysis of polyethylene biodegradation by Pseudomonas fluorescens JNU01 isolated from landfill environments

• Pseudomonas fluorescens strain JNU01 isolated from landfill shows remarkable growth in PE media. • Comparative genomics revealed a unique the AlkB and BVMO genes in strain JNU01 absent in non-PE-degrading reference strain DR133. • FT-IR and SEM analyses confirmed surface modifications in strain JNU01-treated polyethylene. • GC-MS identified 14 metabolites validating complete PE degradation pathways. • Recombinant Pf AlkB has been confirmed to be essential in initiating PE biodegradation. Polyethylene (PE), the world's most produced plastic, poses significant environmental challenges due to its chemical stability and resistance to natural degradation. Here, we report the first isolation of Pseudomonas fluorescens strain JNU01 from landfill environments that can exclusively grow on PE, reaching OD 600 of 0.9 within 2 days. PE biodegradation was confirmed using Fourier transform infrared spectroscopy (FT-IR), field emission scanning electron microscopy (FE-SEM), and gas chromatography-mass spectrometry (GC-MS). Comparative genomic analysis revealed that strain JNU01 harbors co-localized alkane monooxygenase (AlkB) and Baeyer-Villiger monooxygenase (BVMO) genes within the same genetic locus, which were completely absent in the non-degrading reference strain DR133. Quantitative RT-PCR showed 1.5-fold upregulation of both genes when cells were grown on PE medium compared to control conditions. Moreover, functional validation demonstrated that recombinant AlkB effectively initiates PE biodegradation through polymer surface hydroxylation under mild conditions (37 °C, pH 7.5). These results suggest a multi-step oxidative pathway involving AlkB and BVMO that could degrade PE by producing 14 distinct metabolites including alkanes, alkanols, and acids. Our findings reveal the first genomically-defined mechanism for complete PE biodegradation, demonstrating that comparative genomic analysis can guide the discovery of novel plastic-degrading enzymes and provide a framework for engineering enhanced plastic waste remediation technologies.