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首页> 外文期刊>ACS Sustainable Chemistry & Engineering >Synergism of Recombinant Podospora anserina PaAA9B with Cellulases Containing AA9s Can Boost the Enzymatic Hydrolysis of Cellulosic Substrates
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Synergism of Recombinant Podospora anserina PaAA9B with Cellulases Containing AA9s Can Boost the Enzymatic Hydrolysis of Cellulosic Substrates

机译：重组Podospora Anserina PaaA9b与含有Aa9s的纤维素酶的协同作用可以提高纤维素基材的酶水解

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To achieve a fast and efficient cellulose hydrolysis with low enzyme loading remains a challenge for an economically feasible biomass biorefinery process. The synergism between cellulase and lytic polysaccharide monooxygenases (LPMOs) has demonstrated great promise, but it appears that such synergistic effects are highly substrate-dependent. In addition, the need for an efficient method of LPMO production has also limited its application. In this study, the production of Podospora anserina AA9B (PaAA9B), one of the most active LPMOs, was optimized by using double-plasmid coexpression in Pichia pastoris, and its hydrolysis-boosting effects on cellulases were assessed on model cellulosic materials and on our in-house-optimized atmospheric glycerol organosolv (AGO)-pretreated lignocellulosic substrates. The results showed that the double-plasmid coexpression technique successfully improved the PaAA9B production, where up to three times more PaAA9B (3.34 g L-1) was expressed in the 5 L bioreactor after 4 days of induction. The addition of recombinant PaAA9B to Cellic CTec2 (CTec2) significantly boosted the cellulose hydrolysis of a filter paper and Avicel by 2.3- and 1.4-folds, respectively, while no effects were observed on carboxymethyl cellulose (CMC). When lignocellulosic substrates were assessed, the PaAA9B also successfully enhanced the cellulose hydrolysis of both add-catalyzed (ac) and alkali-catalyzed (al) AGO-pretreated sugarcane bagasses by 30 and 20%, respectively, at a 5% solid loading (w/v). At industrially relevant high-solid loading (20% w/v) hydrolysis of an al-AGO-pretreated substrate, the combination of 1.0 mg of PaAA9B and 3 FPU of cellulase per gram of substrate achieved 83% cellulose hydrolysis with 105 g L-1 of the corresponding glucose concentration after 72 h. These results indicated that the double-plasmid coexpression strategy is viable for the high-yield PaAA9B production. The mixture of PaAA9B and cellulase enzymes containing other AA9s exhibited a strong cosynergistic interaction and further boosted the enzymatic hydrolysis of both model cellulosic substrates and our optimized AGO-pretreated lignocellulosic biomass with industrially relevant enzyme loading. This study sheds light on the industrially relevant PaAA9B utilization in the enzymatic hydrolysis of lignocellulosic substrates.
机译：为了实现具有低酶负载的快速高效的纤维素水解仍然是一种经济上可行的生物量生物填充方法的挑战。纤维素酶和裂解多糖单氧基酶（LPMOS）之间的协同作用表现出很大的希望，但似乎这种协同效应依赖于高基板。此外，对LPMO生产有效方法的需求也限制了其应用。在该研究中，通过使用匹氏紫杉醇的双质粒共表达优化了Podospora Anserina Aa9b（Paaa9b）的生产，并在模型纤维素材料和我们的水解纤维素酶对纤维素酶的水解升压效应进行了优化。内部优化的大气甘油有机溶胶（前） - 普通木质纤维素底物。结果表明，双质粒共表达技术成功改善了PAAA9B的生产，其中在诱导4天后在5L生物反应器中表达了3℃的PAAA9B（3.34g L-1）的3倍。添加重组PaaA9b至Celectic CTEC2（CTEC2）显着提高了滤纸的纤维素水解，分别通过2.3和1.4倍，同时在羧甲基纤维素（CMC）上没有观察到任何效果。当评估木质纤维素基质时，PAAA9B也成功地增强了加催化（AC）和碱催化（Al）往返的纤维素水解，分别在5％固载（W. / v）。在工业相关的高固体载量（20％w / v）水解的Al往前预处理的基材上，每克底物的1.0mg PaaA9b和3fpu的纤维素酶的组合达到83％纤维素水解，105g l- 1在72小时后的相应葡萄糖浓度。这些结果表明，高产PAAA9B生产可行的双质粒共表达策略是可行的。含有其他AA9S的PaaA9b和纤维素酶的混合物表现出强烈的舒适性相互作用，并进一步提高了模型纤维素基质的酶水解，并通过工业相关的酶负载进行了预处理的木质纤维素生物量。本研究阐明了在酶促水解的工业相关的PAAA9B利用中的阐明。

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