Key Laboratory of Biomass Chemical Engineering of Ministry of Education, College of Chemical and Biological Engineering, Zhejiang University, Hangzhou, 310027, China.
J Ind Microbiol Biotechnol. 2018 Sep;45(9):803-811. doi: 10.1007/s10295-018-2041-5. Epub 2018 Jun 16.
The industrial application of Trichoderma reesei has been greatly limited by insufficient β-glucosidase activity in its cellulase system. In this study, a novel β-glucosidase expression cassette was constructed and integrated at the target site in T. reesei ZU-02, which achieved the overexpression of β-glucosidase gene and in situ disruption of the cellulase transcriptional repressor ACE1. The resulting transformants showed significant increase in both β-glucosidase activity (BGA) and filter paper activity (FPA). The BGA and FPA increased to 25.13 IU/mL and 20.06 FPU/mL, respectively, 167- and 2.45-fold higher than that of the host strain. Meanwhile, the obtained cellulase system exhibited improved ratio of BGA to FPA, leading to better synergistic effect between cellulase components. Furthermore, submerged fermentation of the transformant was established in 50 m fermenter yielding 112.2 IU/mL β-glucosidase and 89.76 FPU/mL total cellulase. The newly constructed T. reesei transformant achieved improved hydrolysis yield (90.6%) with reduced enzyme loading (15 FPU/g substrate).
里氏木霉的工业应用受到其纤维素酶系统中β-葡萄糖苷酶活性不足的极大限制。在这项研究中，构建了一种新型的β-葡萄糖苷酶表达盒，并整合到里氏木霉 ZU-02 的靶位，实现了β-葡萄糖苷酶基因的过表达和纤维素酶转录阻遏物 ACE1 的原位破坏。所得转化体的β-葡萄糖苷酶活性（BGA）和滤纸活性（FPA）均显著增加。BGA 和 FPA 分别增加到 25.13 IU/mL 和 20.06 FPU/mL，比宿主菌株分别高 167 倍和 2.45 倍。同时，获得的纤维素酶系统表现出改善的 BGA 与 FPA 比值，从而使纤维素酶成分之间具有更好的协同作用。此外，在 50 m 发酵罐中进行了转化体的液体深层发酵，产β-葡萄糖苷酶 112.2 IU/mL，总纤维素酶 89.76 FPU/mL。新构建的里氏木霉转化体在降低酶用量（15 FPU/g 底物）的情况下实现了提高的水解产率（90.6%）。
