Improving the decolorization activity of Bacillus pumilus W3 CotA-laccase to Congo Red by rational modification

Congo Red (CR) is a typical azo dye with highly toxic and carcinogenic properties. This study aimed to improve the decolorization activity of Bacillus pumilus W3 CotA-laccase for azo dye CR. This work analyzed the interaction between CotA-laccase and CR based on homology modeling and molecular docking. The three amino acids (Gly323, Thr377, Thr418) in the substrate-binding pocket were rationally modified through saturation mutation. Finally, the obtained multi-site mutants T377I/T418G and G323S/T377I/T418G decolorized 76.59% and 59.37% of CR within 24 h at pH 8.0 without a mediator, which were 3.15- and 2.44-fold higher than the wild-type CotA. The catalytic efficiency of the multi-site mutants T377I/T418G and G323S/T377I/T418G to CR were increased by 2.21- and 2.01-fold compared with the wild-type CotA, respectively. The mechanism of activity enhancement of mutants was proposed by structural analysis. This evidence suggests that the mutants T377I/T418G and G323S/T377I/T418G could be used as novel bioremediation tools.