同时增强里氏木霉纤维素酶的β-外切协同作用和外切-外切协同作用，以提高纤维素降解能力。
Simultaneous enhancement of the beta-exo synergism and exo-exo synergism in Trichoderma reesei cellulase to increase the cellulose degrading capability.
机构信息
College of Life Sciences, Northwest A&F University, 22 Xinong Road, Yangling, 712100, Shaanxi, China.
Biomass Energy Center for Arid and Semi-arid Lands, Northwest A&F University, 22 Xinong Road, Yangling, 712100, Shaanxi, China.
出版信息
Microb Cell Fact. 2019 Jan 18;18(1):9. doi: 10.1186/s12934-019-1060-x.
BACKGROUND
Cellulase is the one of the largest contributors to the high production costs of the lignocellulose-based biorefineries. As the most widely used cellulase producer, Trichoderma reesei has two weaknesses, deficiencies in β-glucosidase and cellobiohydrolase II. This work aimed at solving this problem by simultaneous enhancement of the beta-exo synergism and exo-exo synergism in T. reesei cellulase to increase the cellulose degrading capability, i.e. enhanced co-expression of the β-glucosidase gene the cellobiohydrolase II gene of T. reesei.
RESULTS
Enhanced co-expression of the β-glucosidase gene and the cellobiohydrolase II gene in T. reesei using the strong promoter Pcbh1 was found successful in overcoming the two weaknesses. Filter paper activities of T. reesei cellulase were greatly elevated, which were 7.21 ± 0.45 (E7, Aabgl1 and Trcbh2) and 7.69 ± 0.42 (F6, Anbgl1 and Trcbh2) FPIU/mL. They were much higher than that of the parental strain Rut-C30, 2.45 ± 0.36 FPIU/mL. Enzymatic hydrolysis yields were also improved, from 67.22 ± 1.61% by Rut-C30 cellulase to 87.98 ± 0.65% by E7 cellulase and 86.50 ± 1.01% by F6 cellulase. The substrate loading for 1 g glucose release from SECS were decreased, from 2.9637 g SECS using Rut-C30 cellulase to 2.0291 g SECS using E7 cellulase and 2.0573 g SECS using F6 cellulase. As a result, the efficiency of the process from SECS to glucose was substantially improved.
CONCLUSIONS
Enhanced co-expression of the β-glucosidase gene and the cellobiohydrolase II gene in T. reesei using the strong promoter Pcbh1 in T. reesei was proven triumphal in the simultaneous enhancement of the beta-exo synergism and exo-exo synergism in T. reesei cellulase. This strategy also improved the cellulase production, enzymatic hydrolysis yield and the efficiency of the process from SECS to glucose in the context of on-site cellulase production. This work is a commendable attempt in the cellulase composition optimization at the transcriptional level.
背景
纤维素酶是木质纤维素生物炼制高生产成本的最大贡献者之一。里氏木霉作为最广泛使用的纤维素酶生产菌，存在β-葡萄糖苷酶和纤维二糖水解酶 II 活性不足的缺陷。本工作旨在通过同时增强里氏木霉纤维素酶的β-外切协同作用和外切-外切协同作用来解决这个问题，即增强里氏木霉的β-葡萄糖苷酶基因和纤维二糖水解酶 II 基因的共表达。
结果
用强启动子 Pcbh1 成功地实现了里氏木霉中β-葡萄糖苷酶基因和纤维二糖水解酶 II 基因的增强共表达，克服了这两个弱点。滤纸纤维素酶活力大大提高，分别为 7.21±0.45（E7、Aabgl1 和 Trcbh2）和 7.69±0.42（F6、Anbgl1 和 Trcbh2）FPIU/mL，显著高于出发菌株 Rut-C30 的 2.45±0.36 FPIU/mL。酶解产率也得到了提高，由 Rut-C30 纤维素酶的 67.22±1.61%提高到 E7 纤维素酶的 87.98±0.65%和 F6 纤维素酶的 86.50±1.01%。从 SECS 释放 1g 葡萄糖的底物加载量也降低了，由 Rut-C30 纤维素酶的 2.9637gSECS 降低到 E7 纤维素酶的 2.0291gSECS 和 F6 纤维素酶的 2.0573gSECS。结果表明，从 SECS 到葡萄糖的过程效率得到了显著提高。
结论
用强启动子 Pcbh1 在里氏木霉中增强共表达β-葡萄糖苷酶基因和纤维二糖水解酶 II 基因，成功地同时增强了里氏木霉纤维素酶的β-外切协同作用和外切-外切协同作用。该策略还提高了纤维素酶的产量、酶解产率和现场生产纤维素酶从 SECS 到葡萄糖的过程效率。这项工作是在转录水平上对纤维素酶组成进行优化的一项值得称赞的尝试。
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