Production and synthetic dyes decolourization capacity of a recombinant laccase from Pichia pastoris

Abstract

Aims: To produce and purify a recombinant laccase from Pichia pastoris and to test its ability in decolourization of synthetic dyes. Methods and Results: A cDNA encoding for a laccase was isolated from Pycnoporus sanguineus and was expressed in P. pastoris strain SMD1168H under the control of the alcohol oxidase (AOX1) promoter. The laccase native signal peptide efficiently directed the secretion of the recombinant laccase in an active form. Factors influencing laccase expression, such as cultivation temperature, pH, copper concentration and methanol concentration, were investigated. The recombinant enzyme was purified to electrophoretic homogeneity, and was estimated to have a molecular mass of about 62·8 kDa. The purified enzyme showed a similar behaviour to the native laccase produced by P. sanguineus. Four different synthetic dyes including azo, anthraquinone, triphenylmethane and indigo dyes could be efficiently decolourized by the purified recombinant laccase without the addition of redox mediators. Conclusions: Heterologous production of P. sanguineus laccase in P. pastoris was successfully achieved. The purified recombinant laccase could efficiently decolourize synthetic dyes in the absence of mediators. Significance and Impact of the Study: This study is the first report on the synthetic dye decolourization by the recombinant P. sanguineus laccase. The decolourization capacity of this recombinant enzyme suggested that it could be a useful biocatalyst for the treatment of dye‐containing effluents.

Cloning and characterization of CotA laccase from Bacillus subtilis WD23 decoloring dyes

“…However, 25 mM EDTA showed only a mild inhibitory effect on the laccase from Pycnoporus sanguineus (Lu et al 2007). The inhibitory effect of dithiothreitol on CotA laccase and spore laccase was similar to the laccase from P. sanguineus (Lu et al 2009), B. licheniformis LS04 (Lu et al 2013) and P. variabile (Forootanfar et al 2011). The cell-wall--associated spore laccase showed more resistance to xylene than the soluble CotA laccase, which might be due to the interaction of laccase with other proteins in the cell wall.…”

“…Laccase activity depended on copper, which was supported in bacterial and fungal laccases (Lu et al 2009(Lu et al , 2013, and laccase from F. fomentarius was activated by Cu 2+ (Neifar et al 2010). Special attention should be paid to copper and iron ion concentrations, as they are common in wastewater and may interfere with laccase activity if they exceed a certain level (Baldrian 2006).…”

Biodegradation of Synthetic Dyes—A Review

“…Many of the recalcitrant compounds are major environmental pollutants, such as munitions waste, pesticides, organochlorines, polychlorinated biphenyls, polycyclic aromatic hydrocarbons, wood preservatives, synthetic polymers, and synthetic dyes (Pointing 2001). Most synthetic dyes are toxic and highly resistant to degradation due to their complex chemical structures (Lu et al 2009). …”

Heterologous laccase production and its role in industrial applications

“…59 A potential use of a recombinant Pycnoporus sanguineus laccase expressed in P. pastoris for the treatment of dye-containing effluents has also been suggested by Lu and co-workers in light of its remarkable ability to degrade, at different extents, four synthetic dyes belonging to different classes (azo, anthraquinone, triphenylmethane and indigo). 109 Finally, a POXA1b laccase mutant, selected for its improved stability in a wider pH range, has been successfully applied to dye decolorisation. This variant shows a further enlargement of dye degradation ability with respect to the wild-type, being also able to decolorize a recalcitrant dye with a complex stilbene type structure.…”

“…Recombinant laccases have been widely applied in bioremediation purposes, especially for the treatment of synthetic dyes 59,80,109 and, more recently, of toxic polychlorinated biphenyls (PCBs), 142 and PAHs. 161 The purified recombinant Lcc1 from Trametes trogii, expressed in P. pastoris differently decolorizes several synthetic textile dyes, depending on their chemical structures.…”