Improvement in lignin peroxidase oxidative stability via surface display by Bacillus subtilis.

Lignin peroxidase (LiP) is a key enzyme involved in lignin degradation. However, the production of natural LiP is limited by the low enzyme-producing capacity of native fungal hosts and the poor stability of the enzyme, hindering its industrialization. To overcome these limitations, we prepared an immobilized enzyme by fusing the LiP gene with the Bacillus subtilis (B. subtilis) spore coat protein CotB and displaying it on the bacterial surface. This approach aims to improve enzymatic stability and industrial applicability. Enzymatic characterization showed that the immobilized LiP exhibited optimal activity at 50 °C and pH 5.0, representing a 10 °C increase over the optimal temperature (40 °C) and a lower pH optimum compared to free LiP (pH 6.0). Under extreme conditions (70 °C, 6 h), the free enzyme retained less than 10% residual activity, whereas CotB-LiP maintained 42.7% ± 1.37% activity. After 1 h incubation in 1 mM HO, CotB-LiP retained 60% relative activity, while free LiP was almost completely inactivated. These findings indicate that spore surface-displayed LiP has significant application potential in environmental remediation and agriculture and provides a reference strategy for low-cost immobilized enzyme production.