Characterization and application of two novel environmental DyP-type peroxidases as sustainable biocatalysts for textile dye treatment

The environmental impact of synthetic textile dyes highlights the urgent need for sustainable wastewater treatment solutions. Here, we report the cloning, heterologous expression, purification, and characterization of two novel bacterial dye-decolorizing peroxidases (DyPs), named PbDyP and MbDyP, selected from the metagenome of the BP8 polyurethane-enriched microbial consortium, isolated from a landfill. Both enzymes showed an acidic pH optimum (4.0-4.5) and a moderate temperature preference (∼40 °C), but they differed in stability and sensitivity to hydrogen peroxide. Spectroscopic analyses confirmed their heme-binding nature, while dynamic light scattering revealed different oligomerization states. Both enzymes achieved efficient biotransformation (>80%) of representative anthraquinone, azo, phthalocyanine, sulfur, and triphenylmethane dyes, with particularly high activity toward anthraquinone Reactive Blue 19. Following enzymatic treatment, FT-IR spectroscopy revealed structural changes in diagnostic functional groups of several dyes, such as Vat Red 10, Disperse Orange 30, Direct Black 22, and Reactive Blue 19, while UPLC-MS analysis identified a lower-mass sodium 2-(3-aminobenzene-1-sulfonyl) ethyl sulfate derivative (m/z 318.31), indicating biotransformation of the parent compound. We also conducted in vitro transformation assays for bisphenols, benzenedithiols, and substituted phenols to thoroughly assess the oxidative capabilities of the DyP-type peroxidases. Toxicity tests using Artemia salina demonstrated that, although enzymatic treatment reduced toxicity in the supernatants, the precipitated fractions remained harmful, underscoring the need for integrated remediation strategies. Overall, these findings position PbDyP and MbDyP as promising candidates for eco-friendly dye degradation, providing a foundation for their potential application in industrial wastewater treatment.