Characterization and application potential of a robust recombinant laccase rTpLcc3 from Trametes polyzona for textile...

Laccases (EC 1.10.3.2) are promising biocatalysts for textile wastewater treatment. Here, we identified a novel laccase, TpLcc3, from Trametes polyzona MUCL 38443, heterologously expressed it in Pichia pastoris, and evaluated its biochemical properties and dye-removal performance in citrate buffer (CB) and a simplified simulated industrial dyebath effluent (DB). The glycosylated recombinant enzyme (rTpLcc3) exhibited maximal activity at 70 °C and pH 3.0, broad pH tolerance (pH 4.0-8.0 at 25-30 °C), and substantial thermal stability with half-lives of 187 min at 50 °C and 19 min at 70 °C. rTpLcc3 showed high catalytic efficiency toward ABTS (k_cat/K_m = 576.76 µM⁻¹ s⁻¹), placing it among high-performing fungal laccases. rTpLcc3 alone effectively decolorized azo dyes-Acid Black 194 (AB194) and Bismarck Brown, with AB194 showing the highest removal in CB (71.39 ± 6.16%) and DB (42.17 ± 5.69%). Mediator supplementation markedly enhanced decolorization, with ABTS enabling rapid, complete removal of AB194, and > 90% removal of Malachite Green in both matrices. Across dye classes, ABTS and syringaldehyde were the most effective mediators in CB (72.4 ± 21.8% and 71.4 ± 28.3% mean decolorization, respectively), while ABTS remained superior in DB (80.1 ± 27.8%). Notably, the TpLcc3/mediator systems significantly reduced cytotoxicity of Malachite Green and Crystal Violet while no increase in cytotoxicity was observed for the remaining dyes under the tested conditions.