Abstract:The CotA-laccase with good thermal stability and resistance to alkaline environments is known as a green catalyst and widely used in industrial dye degradation and wastewater treatment. The enzymatic properties of CotA-laccase cloned from Bacillus pumilus W3 expressed in E. coli was studied. Its yield was increased through optimizing fermentation medium. The recombinant expression of CotA laccase gene in E. coli was achieved by molecular means. The CotA-laccase fermentation medium was optimized by BP neural network and firefly algorithms, and the dye decolorization of recombined CotA-laccase was investigated. The recombinant CotA-laccase had good thermal stability and alkali tolerance. The optimal condition was pH 3.5 and 80 ℃.Recombinant CotA laccase showed goodaffinity for ABTS with a Km of 0.247 mmol/L and a Kcat of 41.32 s-1. The expression of recombinant CotA laccase was 1915.3 U/L under suitable conditions. The compositions of fermentation medium optimized by BP neural network and firefly algorithms were 1.16% of peptone, 0.428% of yeast powder, 1.06% of NaCl, 0.274 mol/L of CuSO 4 and 0.931% of glycerol. The CotA-laccase activity of the optimized medium reached 3 088.68 U/L, which was 61.26% higher than that before optimization. The significant decolorization effect of recombinant CotA laccase on malachite green and acid blue 129 was observed, reaching 90%decolorization rate. CotA-laccase cloned from Bacillus pumilus W3 has good enzymatic properties, showing promising industrial application prospects.The BP neural network modeling combined with firefly algorithm is a feasible methodto optimize the E. coli fermentation.
