Laccases can catalyze the remediation of hazardous synthetic dyes in an eco-friendly manner, and thermostable laccases are advantageous to treat high-temperature dyeing wastewater. A novel laccase from Geothermobacter hydrogeniphilus (Ghlac) was cloned and expressed in Escherichia coli. Ghlac containing 263 residues was characterized as a functional laccase of the DUF152 family. By structural and biochemical analyses, the conserved residues H78, C119, and H136 were identified to bind with one copper atom to fulfill the laccase activity. In order to make it more suitable for industrial use, Ghlac variant Mut2 with enhanced thermostability was designed. The half-lives of Mut2 at 50 °C and 60 °C were 80.6 h and 9.8 h, respectively. Mut2 was stable at pH values ranging from 4.0 to 8.0 and showed a high tolerance for organic solvents such as ethanol, acetone, and dimethyl sulfoxide. In addition, Mut2 decolorized approximately 100% of 100 mg/L of malachite green dye in 3 h at 70 °C. Furthermore, Mut2 eliminated the toxicity of malachite green to bacteria and Zea mays. In summary, the thermostable laccase Ghlac Mut2 could effectively decolorize and detoxify malachite green at high temperatures, showing great potential to remediate the dyeing wastewater.

Objective: The present study was conducted to examine the gas production, fermentation characteristics, nutrient degradation, and methanogenic community composition of a rumen fluid culture with Broussonetia papyrifera (B. papyrifera) subjected to ensiling or steam explosion (SE) pretreatment.Methods: Fresh B. papyrifera was collected and pretreated by ensiling or SE, which was then fermented with ruminal fluids as ensiled B. papyrifera group, steam-exploded B. papyrifera group, and untreated B. papyrifera group. The gas and methane production, fermentation characteristics, nutrient degradation, and methanogenic community were determined during the fermentation.Results: Cumulative methane production was significantly improved with SE pretreatment compared with ensiled or untreated biomass accompanied with more volatile fatty acids production. After 72 h incubation, SE and ensiling pretreatments decreased the acid detergent fiber contents by 39.4% and 22.9%, and neutral detergent fiber contents by 10.6% and 47.2%, respectively. Changes of methanogenic diversity and abundance of methanogenic archaea corresponded to the variations in fermentation pattern and methane production.Conclusion: Compared with ensiling pretreatment, SE can be a promising technique for the efficient utilization of B. papyrifera, which would contribute to sustainable livestock production systems.