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Cellulase hyper-production by Trichoderma reesei mutant SEU-7 on lactose.
Biotechnol Biofuels. 2017; 10:228.BB

Abstract

BACKGROUND

The induction of cellulase production by insoluble carbon source cellulose was a common and efficient strategy, but has some drawbacks, such as difficult fermentation operation, substantial cellulase loss, long fermentation time, and high energy-consumption, resulting in high cost of cellulase production in industry. These drawbacks can be overcome if soluble carbon sources are utilized as the inducers for cellulase production. However, until now the induction efficiency of most soluble carbon sources, especially lactose and glucose, is still inferior to cellulose despite extensive efforts have been made by either optimizing the fermentation process or constructing the recombinant strains. Therefore, strain improvement by metabolic engineering for high induction efficiency of soluble carbon sources is of great interest.

RESULTS

Trichoderma reesei mutant SEU-7 was constructed from T. reesei RUT-C30 with the overexpression of endogenous gene β-glucosidase (BGL1) by insertional mutagenesis via Agrobacterium tumefaciens-mediated transformation (AMT). Compared to RUT-C30, SEU-7 displays substantially enhanced activities of both cellulase and hemicellulase when grown on either lactose or cellulose. The induction efficiency with lactose was found to be higher than cellulose in strain SEU-7. To the best of our knowledge, we achieved the highest FPase activity in SEU-7 in both batch culture (13.0 IU/mL) and fed-batch culture (47.0 IU/mL) on lactose. Moreover, SEU-7 displayed unrivaled pNPGase activity on lactose in both batch culture (81.0 IU/mL) and fed-batch culture (144.0 IU/mL) as compared to the other reported T. reesei strains in the literature grown in batch or fed-batch experiments on cellulose or lactose. This superiority of SEU-7 over RUT-C30 improves markedly the saccharification ability of SEU-7 on pretreated corn stover. The overexpression of gene BGL1 was found either at the mRNA or at the protein level in the mutant strains with increased cellulase production in comparison with RUT-C30, but only SEU-7 displayed much higher expression of gene BGL1 on lactose than on cellulose. Two copies of gene BGL1 were inserted into the chromosome of T. reesei SEU-7 between KI911141.1:347357 and KI911141.1:347979, replacing the original 623-bp fragment that is not within any genes' coding region. The qRT-PCR analysis revealed that the mRNA levels of both cellulase and hemicellulase were upregulated significantly in SEU-7, together with the MFS transporter CRT1 and the XYR1 nuclear importer KAP8.

CONCLUSIONS

Recombinant T. reesei SEU-7 displays hyper-production of both cellulase and hemicellulase on lactose with the highest FPase activity and pNPGase activity for T. reesei, enabling highly efficient saccharification of pretreated biomass. For the first time, the induction efficiency for cellulase production by lactose in T. reesei was reported to be higher than that by cellulose. This outperformance of T. reesei SEU-7, which is strain-specific, is attributed to both the overexpression of gene BGL and the collateral mutation. Moreover, the increased transcription levels of cellulase genes, the related transcription factors, and the MFS transporter CRT1 contribute to the outstanding cellulase production of SEU-7. Our research advances strain improvement to enhance the induction efficiency of soluble carbon sources to produce cost-effective cellulase and hemicellulase in industry.

Authors+Show Affiliations

State Key Laboratory of Bioelectronics, School of Biological Science and Medical Engineering, Southeast University, Nanjing, 210096 China.State Key Laboratory of Bioelectronics, School of Biological Science and Medical Engineering, Southeast University, Nanjing, 210096 China. 35 Jinxianghe Road, Xuanwu District, Nanjing, 210008 Jiangsu Province China.State Key Laboratory of Bioelectronics, School of Biological Science and Medical Engineering, Southeast University, Nanjing, 210096 China.Key Laboratory of Systems Bioengineering (Ministry of Education), School of Chemical Engineering and Technology, Tianjin University, Weijin Road 92, Nankai District, Tianjin, 300072 People's Republic of China.Key Laboratory of Systems Bioengineering (Ministry of Education), School of Chemical Engineering and Technology, Tianjin University, Weijin Road 92, Nankai District, Tianjin, 300072 People's Republic of China.Key Laboratory of Synthetic Biology, Institute of Plant Physiology and Ecology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai, 200032 China.School of Environmental and Biological Engineering, Nanjing University of Science and Technology, Nanjing, 210094 China.Department of Chemistry, University of Michigan, 930 North University Avenue, Ann Arbor, MI 48109 USA.

Pub Type(s)

Journal Article

Language

eng

PubMed ID

29034003

Citation

Li, Chengcheng, et al. "Cellulase Hyper-production By Trichoderma Reesei Mutant SEU-7 On Lactose." Biotechnology for Biofuels, vol. 10, 2017, p. 228.
Li C, Lin F, Zhou L, et al. Cellulase hyper-production by Trichoderma reesei mutant SEU-7 on lactose. Biotechnol Biofuels. 2017;10:228.
Li, C., Lin, F., Zhou, L., Qin, L., Li, B., Zhou, Z., Jin, M., & Chen, Z. (2017). Cellulase hyper-production by Trichoderma reesei mutant SEU-7 on lactose. Biotechnology for Biofuels, 10, 228. https://doi.org/10.1186/s13068-017-0915-9
Li C, et al. Cellulase Hyper-production By Trichoderma Reesei Mutant SEU-7 On Lactose. Biotechnol Biofuels. 2017;10:228. PubMed PMID: 29034003.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Cellulase hyper-production by Trichoderma reesei mutant SEU-7 on lactose. AU - Li,Chengcheng, AU - Lin,Fengming, AU - Zhou,Le, AU - Qin,Lei, AU - Li,Bingzhi, AU - Zhou,Zhihua, AU - Jin,Mingjie, AU - Chen,Zhan, Y1 - 2017/10/04/ PY - 2017/05/16/received PY - 2017/09/23/accepted PY - 2017/10/17/entrez PY - 2017/10/17/pubmed PY - 2017/10/17/medline KW - Cellulase KW - Hemicellulase KW - Hyper-production KW - Soluble carbon source KW - Trichoderma reesei KW - β-Glucosidase SP - 228 EP - 228 JF - Biotechnology for biofuels JO - Biotechnol Biofuels VL - 10 N2 - BACKGROUND: The induction of cellulase production by insoluble carbon source cellulose was a common and efficient strategy, but has some drawbacks, such as difficult fermentation operation, substantial cellulase loss, long fermentation time, and high energy-consumption, resulting in high cost of cellulase production in industry. These drawbacks can be overcome if soluble carbon sources are utilized as the inducers for cellulase production. However, until now the induction efficiency of most soluble carbon sources, especially lactose and glucose, is still inferior to cellulose despite extensive efforts have been made by either optimizing the fermentation process or constructing the recombinant strains. Therefore, strain improvement by metabolic engineering for high induction efficiency of soluble carbon sources is of great interest. RESULTS: Trichoderma reesei mutant SEU-7 was constructed from T. reesei RUT-C30 with the overexpression of endogenous gene β-glucosidase (BGL1) by insertional mutagenesis via Agrobacterium tumefaciens-mediated transformation (AMT). Compared to RUT-C30, SEU-7 displays substantially enhanced activities of both cellulase and hemicellulase when grown on either lactose or cellulose. The induction efficiency with lactose was found to be higher than cellulose in strain SEU-7. To the best of our knowledge, we achieved the highest FPase activity in SEU-7 in both batch culture (13.0 IU/mL) and fed-batch culture (47.0 IU/mL) on lactose. Moreover, SEU-7 displayed unrivaled pNPGase activity on lactose in both batch culture (81.0 IU/mL) and fed-batch culture (144.0 IU/mL) as compared to the other reported T. reesei strains in the literature grown in batch or fed-batch experiments on cellulose or lactose. This superiority of SEU-7 over RUT-C30 improves markedly the saccharification ability of SEU-7 on pretreated corn stover. The overexpression of gene BGL1 was found either at the mRNA or at the protein level in the mutant strains with increased cellulase production in comparison with RUT-C30, but only SEU-7 displayed much higher expression of gene BGL1 on lactose than on cellulose. Two copies of gene BGL1 were inserted into the chromosome of T. reesei SEU-7 between KI911141.1:347357 and KI911141.1:347979, replacing the original 623-bp fragment that is not within any genes' coding region. The qRT-PCR analysis revealed that the mRNA levels of both cellulase and hemicellulase were upregulated significantly in SEU-7, together with the MFS transporter CRT1 and the XYR1 nuclear importer KAP8. CONCLUSIONS: Recombinant T. reesei SEU-7 displays hyper-production of both cellulase and hemicellulase on lactose with the highest FPase activity and pNPGase activity for T. reesei, enabling highly efficient saccharification of pretreated biomass. For the first time, the induction efficiency for cellulase production by lactose in T. reesei was reported to be higher than that by cellulose. This outperformance of T. reesei SEU-7, which is strain-specific, is attributed to both the overexpression of gene BGL and the collateral mutation. Moreover, the increased transcription levels of cellulase genes, the related transcription factors, and the MFS transporter CRT1 contribute to the outstanding cellulase production of SEU-7. Our research advances strain improvement to enhance the induction efficiency of soluble carbon sources to produce cost-effective cellulase and hemicellulase in industry. SN - 1754-6834 UR - https://www.unboundmedicine.com/medline/citation/29034003/Cellulase_hyper_production_by_Trichoderma_reesei_mutant_SEU_7_on_lactose_ L2 - https://biotechnologyforbiofuels.biomedcentral.com/articles/10.1186/s13068-017-0915-9 DB - PRIME DP - Unbound Medicine ER -
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