High-level expression of a bacterial laccase, CueO from Escherichia coli K12 in Pichia pastoris GS115 and its application on the decolorization of synthetic dyes.

Laccases are oxidoreductase catalyze the oxidation of a wide range of substrates with oxygen as the electron acceptor. This report was aimed to the high-level expression of a laccase, CueO from Escherichia coli K12 in Pichia pastoris GS115 and its application on decolorization of synthetic dyes. The yacK gene coding CueO was cloned into an expression vector of Pichia pastoris, pHBM905BDM and expressed in a secretory form with Pichia pastoris GS115 as the host. The yield of the recombinant protein was 556mg/L with high-density fermentation and the enzyme activity was about 41,000U/L. The recombinant laccase was purified and characterized. Its optimum pH and temperature was 3.0 and 55°C with 2, 2'-azino-bis-(3-ethylbenzothazoline-6-sulfonic acid) (ABTS) as the substrate, respectively. This recombinant protein was thermostable and its half life at 70°C was about 60min. In the presence of natural redox mediator acetosyringone, the purified recombinant laccase decolorized 98.1% and 98.5% of Congo red, malachite green, respectively. It also decolorized 90.03% of Remazol brilliant blue R without this mediator. In addition, this enzyme was applied on the decolorization of wastewater from a textile printing factory and showed an obvious bleaching effect.