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Fungal laccases have shown great potential in industrial and environmental applications. They are generally produced as laccase isoenzymes. Thus, to further study the properties of different laccase isoenzymes and their performance in bio-remediation is essential for a deep understanding of laccase function and application. In this study, three Pleurotus ostreatus HAUCC 162 laccase isoenzymes were heterologously expressed, and the effects of different inhibitors, metal ions, and organic solvents on the activity of recombinant laccases were evaluated. In the dye decolorization test, LACC6 showed the highest ability to remove Malachite green (MG), Remazol Brilliant Blue R (RBBR), Bromophenol blue (BB), and Methyl orange (MO) among the three recombinant laccases. Removal rates within 24 h were 91.5%, 84.9%, 79.1%, and 73.1% for MG (100 mg/L), RBBR (100 mg/L), BB (100 mg/L), and MO (100 mg/L), respectively. The MG and RBBR transformation pathways were proposed by using High Performance Liquid Chromatography-Mass Spectrometry (LC-MS) analysis. Based on the results of this work, the production of recombinant LACC6 or improving the portion of LACC6 in the crude extracellular laccase may advance synthetic dye removal., (Copyright © 2019. Published by Elsevier Ltd.)

Chlorophenols (CPs), nitrophenols (NPs), and sulfonamide antibiotics (SAs) are three types of environmental pollutants that are of great concern because of their prevalence and toxicity. In this study, three laccase isoenzymes obtained from Pleurotus ostreatus HAUCC 162 were heterologously expressed and characterized with respect to their ability to degrade CPs, NPs, and, SAs. The three recombinant laccases can efficiently degrade the three types of considered pollutants using a laccase-mediator system (LMS). Their specific efficiencies for the removal of 2NP, 3NP, 4NP, 4CP, 2,4-dichlorophenol (DCP), 2,6-DCP, sulfadiazine (SDZ), sulfamethazine (SMZ), and sulfamethoxazole (SMX) over 60min were 59.21%, 47.91%, 60.24%, 74.9%, 28.9%, 35.1%, 98.1%, 97.5%, and 97.8%, respectively. Based on the analysis of the oxidation products of the CPs, NPs, and SAs, pollutant removal pathways are proposed, namely, the production of 3-nitromuconate and 3-chloromuconate as the key intermediates of 4-NP and 2, 4-DCP; and oxidative coupling for the transformation of SDZ by LMS. The results of present work indicated the laccases could efficiently remove NPs, CPs, and SAs in LMS, which offers an opportunity to apply P. ostreatus HAUCC 162 laccase in the field of environmental biotechnology., (Copyright © 2017 Elsevier B.V. All rights reserved.)

Laccase is useful for various biotechnological and industrial applications. The white-rot fungus Trametes velutina 5930 and its laccase, isolated from the Shennongjia Nature Reserve in China by our laboratory, has great potential for practical application in environmental biotechnology. However, the original level of laccase produced by Trametes velutina 5930 was relatively low in the absence of any inducer. Therefore, in order to enhance the laccase production by Trametes velutina 5930 and make better use of this fungus in the field of environmental biotechnology, the regulation of laccase production and laccase gene expression in Trametes velutina 5930 were investigated in this study. Different metal ions such as Cu(2+) and Fe(2+) could stimulate the laccase synthesis and laccase gene transcription in Trametes velutina 5930. Some aromatic compounds structurally related to lignin, such as tannic acid, syringic acid, cinnamic acid, gallic acid and guaiacol, could also enhance the level of laccase activity and laccase gene transcription. We also found that there existed a positive synergistic effect of aromatic compound and metal ion on the laccase production and laccase gene transcription in Trametes velutina 5930. Taken together, our study may contribute to the improvement of laccase productivity by Trametes velutina 5930.

Laccase is a copper-containing polyphenol oxidase that has great potential in industrial and biotechnological applications. Previous research has suggested that fungal laccase may be involved in the defense against oxidative stress, but there is little direct evidence supporting this hypothesis, and the mechanism by which laccase protects cells from oxidative stress also remains unclear. Here, we report that the expression of the laccase gene from white rot fungus in Pichia pastoris can significantly enhance the resistance of yeast to H(2)O(2)-mediated oxidative stress. The expression of laccase in yeast was found to confer a strong ability to scavenge intracellular H(2)O(2) and to protect cells from lipid oxidative damage. The mechanism by which laccase gene expression increases resistance to oxidative stress was then investigated further. We found that laccase gene expression in Pichia pastoris could increase the level of glutathione-based antioxidative activity, including the intracellular glutathione levels and the enzymatic activity of glutathione peroxidase, glutathione reductase, and γ-glutamylcysteine synthetase. The transcription of the laccase gene in Pichia pastoris was found to be enhanced by the oxidative stress caused by exogenous H(2)O(2). The stimulation of laccase gene expression in response to exogenous H(2)O(2) stress further contributed to the transcriptional induction of the genes involved in the glutathione-dependent antioxidative system, including PpYAP1, PpGPX1, PpPMP20, PpGLR1, and PpGSH1. Taken together, these results suggest that the expression of the laccase gene in Pichia pastoris can enhance the resistance of yeast to H(2)O(2)-mediated oxidative stress by stimulating the glutathione-based antioxidative system to protect the cell from oxidative damage.

A laccase-producing white-rot fungi strain Ganoderma sp.En3 was newly isolated from the forest of Tzu-chin Mountain in China. Ganoderma sp.En3 had a strong ability of decolorizing four synthetic dyes, two simulated dye bath effluents and the real textile dye effluent. Induction in the activity of laccase during the decolorization process indicated that laccase played an important role in the efficient decolorization of different dyes by this fungus. Phytotoxicity study with respect to Triticum aestivum and Oryza sativa demonstrated that Ganoderma sp.En3 was able to detoxify four synthetic dyes, two simulated dye effluents and the real textile dye effluent. The laccase gene lac-En3-1 and its corresponding full-length cDNA were then cloned and characterized from Ganoderma sp.En3. The deduced protein sequence of LAC-En3-1 contained four copper-binding conserved domains of typical laccase protein. The functionality of lac-En3-1 gene encoding active laccase was verified by expressing this gene in the yeast Pichia pastoris successfully. The recombinant laccase produced by the yeast transformant could decolorize the synthetic dyes, simulated dye effluents and the real textile dye effluent. The ability of decolorizing different dyes was positively related to the laccase activity. In addition, the 5'-flanking sequence upstream of the start codon ATG in lac-En3-1 gene was obtained. Many putative cis-acting responsive elements were predicted in the promoter region of lac-En3-1., (Copyright © 2011 Elsevier B.V. All rights reserved.)

The laccase gene lac48424-1 and its corresponding full-length cDNA were cloned and characterized from a novel white-rot fungi Trametes sp. 48424 which had the high yield of laccase and strong ability for decolorizing different dyes. The 1563 bp full-length cDNA of lac48424-1 encoded a mature laccase protein containing 499 amino acids preceded by a signal peptide of 21 amino acids. The deduced protein sequence of LAC48424-1 showed high similarity with other known fungal laccases and contained four copper-binding conserved domains of typical laccase protein. The functionality of lac48424-1 gene encoding active laccase was verified by expressing the gene in the yeast Pichia pastoris successfully. It was found that the recombinant laccase produced by the yeast transformant could decolorize different dyes. The 5'-flanking sequence upstream of start codon was obtained by Self-Formed Adaptor PCR. Many putative cis-acting responsive elements involved in the transcriptional regulation were identified in the promoter region of lac48424-1., (Copyright © 2010 Elsevier Ltd. All rights reserved.)

Promotion of laccase production has attracted much attention, because of its potential application in environment and industry areas. Metal ions and aromatic compounds are known to be effective in stimulating laccase production in many white-rot fungi. In this study, we examined the effect of several metal ions and aromatic compounds on extracellular laccase activity as well as on the transcription of laccase multi-gene family in Pleurotus ostreatus HAUCC 162. Extracellular laccase activity is enhanced by the addition of Cu 2+ (3.86-fold), Fe 2+ (1.85-fold), Mn 2+ (1.65-fold), Cd 2+ (1.45-fold), ferulic acid (1.83-fold), vanillic acid (1.62-fold), coumaric acid (2.16-fold), and cinnamic acid (1.95-fold) after 10-days induction, and the increased transcript level of laccase multi-gene family was also observed. Synergistic stimulation of metal ions (Fe 2+ , Cu 2+ ) and aromatic compounds (vanillic acid, cinnamic acid, and ferulic acid) further improved the stimulatory effect, compared with single inducer. The results of decolorization test further indicated the laccase produced by P. ostreatus HAUCC 162 had a strong capacity to decolorize different synthetic dyes, which offers an opportunity to apply P. ostreatus HAUCC 162 laccase in the field of environmental biotechnology. [ABSTRACT FROM AUTHOR]