Improved Production of Majority Cellulases in Trichoderma reesei by Integration of cbh1 Gene From Chaetomium thermophilum
Improved Production of Majority Cellulases in Trichoderma reesei by Integration of cbh1 Gene From Chaetomium thermophilum
复制标题
通过整合来自嗜热毛壳菌的 cbh1 基因提高里氏木霉中大多数纤维素酶的产量
DOI：
10.3389/fmicb.2020.01633
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发表时间：
2020-07-14
影响因子：
5.2
通讯作者：
Qin, Lina
中科院分区：
文献类型：
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作者：
Jiang, Xianzhang;Du, Jiawen;Qin, Lina
木质纤维素是一种丰富的废弃资源，被认为是生产生物燃料或其他有价值生物产品的一种有前景的材料。目前，木质纤维素材料经济利用的主要瓶颈之一是将木质纤维素转化为可发酵的可溶性糖的成本效益。解决这一问题的一种方法是寻找更优的木质纤维素降解酶或进一步提高当前木质纤维素酶的产量。在本研究中，首先对一种嗜热真菌嗜热毛壳菌（Chaetomium thermophilum）的木质纤维素降解能力进行了评估，并与生物技术常用菌株里氏木霉（Trichoderma reesei）进行了比较。数据表明，与里氏木霉相比，嗜热毛壳菌在纤维素酶产量相对较低的情况下展现出显著更高的纤维素利用效率，这表明嗜热毛壳菌中可能存在更好的纤维素酶。对嗜热毛壳菌和里氏木霉之间的蛋白质分泌组的比较显示，这两个物种分泌的蛋白质类别差异很大。此外，为了证明嗜热毛壳菌中的纤维素酶具有更好的酶学特性，首先分别对嗜热毛壳菌和里氏木霉的主要纤维素酶纤维二糖水解酶I（CBH1）进行了特性分析。数据显示，在较宽的温度和pH范围内，嗜热毛壳菌CBH1的比活性比里氏木霉CBH1高约4.5倍。为了探究提高里氏木霉中CBH1的活性是否有助于提高里氏木霉的整体纤维素利用效率，通过将嗜热毛壳菌cbh1基因整合到里氏木霉cbh1基因位点，用嗜热毛壳菌cbh1基因替换了里氏木霉cbh1基因。数据令人惊讶地表明，这种基因替换不仅使纤维二糖水解酶活性提高了约4.1倍，还导致其他纤维素酶基因的诱导更强，使得滤纸酶活性、偶氮 - 羧甲基纤维素（Azo - CMC）活性和β - 葡萄糖苷酶活性分别提高了约2.2倍、1.9倍和2.3倍。本研究不仅为优质纤维素酶基因提供了新资源以及提高里氏木霉纤维素酶产量的新策略，还有助于为嗜热毛壳菌的基础研究开辟道路。
Lignocellulose is an abundant waste resource and has been considered as a promising material for production of biofuels or other valuable bio-products. Currently, one of the major bottlenecks in the economic utilization of lignocellulosic materials is the cost-efficiency of converting lignocellulose into soluble sugars for fermentation. One way to address this problem is to seek superior lignocellulose degradation enzymes or further improve current production yields of lignocellulases. In the present study, the lignocellulose degradation capacity of a thermophilic fungus Chaetomium thermophilum was firstly evaluated and compared to that of the biotechnological workhorse Trichoderma reesei. The data demonstrated that compared to T. reesei, C. thermophilum displayed substantially higher cellulose-utilizing efficiency with relatively lower production of cellulases, indicating that better cellulases might exist in C. thermophilum. Comparison of the protein secretome between C. thermophilum and T. reesei showed that the secreted protein categories were quite different in these two species. In addition, to prove that cellulases in C. thermophilum had better enzymatic properties, the major cellulase cellobiohydrolase I (CBH1) from C. thermophilum and T. reesei were firstly characterized, respectively. The data showed that the specific activity of C. thermophilum CBH1 was about 4.5-fold higher than T. reesei CBH1 in a wide range of temperatures and pH. To explore whether increasing CBH1 activity in T. reesei could contribute to improving the overall cellulose-utilizing efficiency of T. reesei, T. reesei cbh1 gene was replaced with C. thermophilum cbh1 gene by integration of C. thermophilum cbh1 gene into T. reesei cbh1 gene locus. The data surprisingly showed that this gene replacement not only increased the cellobiohydrolase activities by around 4.1-fold, but also resulted in stronger induction of other cellulases genes, which caused the filter paper activities, Azo-CMC activities and β-glucosidase activities increased by about 2.2, 1.9, and 2.3-fold, respectively. The study here not only provided new resources of superior cellulases genes and new strategy to improve the cellulase production in T. reesei, but also contribute to opening the path for fundamental research on C. thermophilum.