You Shuai, Xie Chen, Ma Rui, Huang Huo-Qing, Herman Richard Ansah, Su Xiao-Yun, Ge Yan, Cai Hui-Yi, Yao Bin, Wang Jun, Luo Hui-Ying
1Key Laboratory for Feed Biotechnology of the Ministry of Agriculture, Feed Research Institute, Chinese Academy of Agricultural Sciences, Beijing, 100081 China.
2School of Biotechnology, Jiangsu University of Science and Technology, Zhenjiang, 212018 People's Republic of China.
Biotechnol Biofuels. 2019 Dec 3;12:278. doi: 10.1186/s13068-019-1620-7. eCollection 2019.
Xylanase is one of the most extensively used biocatalysts for biomass degradation. However, its low catalytic efficiency and poor thermostability limit its applications. Therefore, improving the properties of xylanases to enable synergistic degradation of lignocellulosic biomass with cellulase is of considerable significance in the field of bioenergy.
Using fragment replacement, we improved the catalytic performance and thermostability of a GH10 xylanase, XylE. Of the ten hybrid enzymes obtained, seven showed xylanase activity. Substitution of fragments, M3, M6, M9, and their combinations enhanced the catalytic efficiency (by 2.4- to fourfold) as well as the specific activity (by 1.2- to 3.3-fold) of XylE. The hybrids, XylE-M3, XylE-M3/M6, XylE-M3/M9, and XylE-M3/M6/M9, showed enhanced thermostability, as observed by the increase in the (3-4.7 °C) and (1.1-4.7 °C), and extended (by 1.8-2.3 h). In addition, the synergistic effect of the mutant xylanase and cellulase on the degradation of mulberry bark showed that treatment with both XylE-M3/M6 and cellulase exhibited the highest synergistic effect. In this case, the degree of synergy reached 1.3, and the reducing sugar production and dry matter reduction increased by 148% and 185%, respectively, compared to treatment with only cellulase.
This study provides a successful strategy to improve the catalytic properties and thermostability of enzymes. We identified several xylanase candidates for applications in bioenergy and biorefinery. Synergistic degradation experiments elucidated a possible mechanism of cellulase inhibition by xylan and xylo-oligomers.
木聚糖酶是生物质降解中应用最广泛的生物催化剂之一。然而，其低催化效率和较差的热稳定性限制了其应用。因此，改善木聚糖酶的性能以实现与纤维素酶协同降解木质纤维素生物质在生物能源领域具有重要意义。
通过片段替换，我们提高了GH10木聚糖酶XylE的催化性能和热稳定性。在获得的10种杂合酶中，7种具有木聚糖酶活性。片段M3、M6、M9及其组合的替换提高了XylE的催化效率（提高了2.4至4倍）以及比活性（提高了1.2至3.3倍）。杂合体XylE-M3、XylE-M3/M6、XylE-M3/M9和XylE-M3/M6/M9表现出增强的热稳定性，通过Tm（提高了3-4.7℃）和T1/2（提高了1.1-4.7℃）的增加以及t1/2（延长了1.8-2.3小时）可以观察到。此外，突变木聚糖酶和纤维素酶对桑树皮降解的协同作用表明，用XylE-M3/M6和纤维素酶处理表现出最高的协同效应。在这种情况下，协同度达到1.3，与仅用纤维素酶处理相比，还原糖产量和干物质减少量分别增加了148%和185%。
本研究提供了一种成功的策略来改善酶的催化性能和热稳定性。我们鉴定了几种可用于生物能源和生物炼制的木聚糖酶候选物。协同降解实验阐明了木聚糖和木寡糖抑制纤维素酶的可能机制。
