School of Life Sciences, Jiangsu University, Zhenjiang, Jiangsu, 212013, China.
Appl Biochem Biotechnol. 2024 Oct;196(10):6849-6863. doi: 10.1007/s12010-024-04869-8. Epub 2024 Feb 27.
Lignin peroxidase (LiP) has a good application prospect in lignin degradation, environmental treatment, straw feed, and other industries. However, its application is constrained by the high price and low stability of enzyme preparation. In this study, the Escherichia coli-Bacillus subtilis (E. coli-B. subtilis) shuttle expression vector pHS-cotG-lip was constructed and displayed on the surface of Bacillus subtilis spores. The analysis of enzymatic properties showed that the optimal catalytic temperature and pH of the immobilized LiP were 55 °C and 4.5, respectively. Compared with free LiP (42 °C and pH4.0), the optimal reaction temperature increased by 13 °C. After incubation at 70 °C for 1 h, its activity remained above 30%, while the free LiP completely lost its activity under the same conditions. Adding Mn, DL-lactic acid, and PEG-4000 increased the CotG-LiP enzyme activity to 313%, 146%, and 265%, respectively. The recyclability of spore display made the fusion protein CotG-LiP retain more than 50% enzyme activity after four cycles. The excellent recycling rate indicated that LiP displayed on the spore surface had a good application prospect in sewage treatment and other fields, and also provided a reference for the rapid and low-cost immobilized production of enzyme preparations.
木质素过氧化物酶（LiP）在木质素降解、环境处理、秸秆饲料等行业具有良好的应用前景。然而，其应用受到酶制剂价格高、稳定性低的限制。本研究构建了大肠杆菌-枯草芽孢杆菌（E. coli-B. subtilis）穿梭表达载体 pHS-cotG-lip，并将其展示在枯草芽孢杆菌孢子表面。酶学性质分析表明，固定化 LiP 的最适催化温度和 pH 分别为 55°C 和 4.5。与游离 LiP（42°C 和 pH4.0）相比，最适反应温度提高了 13°C。在 70°C 孵育 1 小时后，其活性仍保持在 30%以上，而游离 LiP 在相同条件下完全失去活性。添加 Mn、DL-乳酸和 PEG-4000 可使 CotG-LiP 酶活分别提高 313%、146%和 265%。孢子展示的可重复性使得融合蛋白 CotG-LiP 在经过四个循环后仍保留超过 50%的酶活。良好的循环回收率表明，表面展示的 LiP 在污水处理等领域具有良好的应用前景，也为酶制剂的快速、低成本固定化生产提供了参考。
