Exploration and heterologous expression of laccase genes and pesticide degradation ability of laccases from Cerrena unicolor GC.u01.

Abstract

Laccases are valuable industrial enzymes with applications across various fields. While heterologous expression in Pichia pastoris is a common strategy, current approaches face limitations in yield, stability, and catalytic efficiency against recalcitrant agrochemicals. In this study, we sequenced and annotated the first high-quality genome of Cerrena unicolor strain GC.u01 (30.95 Mb, 8,089 genes), revealing a unique laccase gene family comprising nine members. Structural analysis revealed novel catalytic motifs in Lac2, which was successfully expressed in P. pastoris GS115 through codon optimization, yielding a novel recombinant enzyme (70 kDa) with exceptional pH stability (retaining > 80% activity at pH 3.0-8.0 for 24 h) and thermotolerance (> 60% activity at 40 °C), surpassing most reported fungal laccases. Notably, Lac2 demonstrated unprecedented degradation efficiency for azoxystrobin (96.2) and phoxim (30.7%)-the first report of a Cerrena unicolor laccase degrading these pesticides-achieving significantly higher rates than previously described laccases under similar conditions. This study integrates genome mining, enzyme engineering, and functional validation to establish a new paradigm for developing robust biocatalysts against recalcitrant agrochemicals. These unique characteristics of Lac2 suggest the potential of this enzyme in biotechnological and industrial applications.