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Cellulase hyper-production by Trichoderma reesei mutant SEU-7 on lactose
Chengcheng Li1, Fengming Lin1,2, Le Zhou1
1State Key Laboratory of Bioelectronics, School of Biological Science and Medical Engineering, Southeast University, Nanjing, 210096 China.
Biotechnology for Biofuels
|October 17, 2017
Summary
Engineered Trichoderma reesei SEU-7 overexpresses β-glucosidase (BGL1), achieving higher cellulase and hemicellulase production on lactose than cellulose. This breakthrough enables cost-effective enzyme production and biomass saccharification.
Area of Science:
- Biotechnology
- Enzyme Engineering
- Industrial Microbiology
Background:
- Cellulase production traditionally uses insoluble cellulose, facing operational challenges and high costs.
- Soluble carbon sources offer potential but exhibit lower induction efficiency compared to cellulose.
- Metabolic engineering aims to enhance soluble carbon source induction for cost-effective enzyme production.
Purpose of the Study:
- To engineer Trichoderma reesei for improved cellulase and hemicellulase production using soluble inducers.
- To investigate the potential of lactose as a superior inducer for enzyme production.
- To enhance the cost-effectiveness of cellulase and hemicellulase production for industrial applications.
Main Methods:
- Construction of Trichoderma reesei mutant SEU-7 from RUT-C30 via Agrobacterium tumefaciens-mediated transformation (AMT).
- Overexpression of the endogenous β-glucosidase (BGL1) gene through insertional mutagenesis.
- Comparative analysis of cellulase and hemicellulase activities on lactose and cellulose using batch and fed-batch cultures.
- Gene expression analysis using qRT-PCR to assess BGL1, cellulase, hemicellulase, CRT1, and KAP8 levels.
Main Results:
- Trichoderma reesei SEU-7 exhibited significantly enhanced cellulase and hemicellulase activities compared to RUT-C30 on both lactose and cellulose.
- Lactose demonstrated higher induction efficiency than cellulose in the SEU-7 strain.
- SEU-7 achieved record FPase (47.0 IU/mL) and pNPGase (144.0 IU/mL) activities in fed-batch culture on lactose, surpassing previously reported strains.
- Improved saccharification of pretreated corn stover was observed with SEU-7.
- Overexpression of BGL1 was confirmed, with SEU-7 showing higher BGL1 expression on lactose.
Conclusions:
- Recombinant Trichoderma reesei SEU-7 hyper-produces cellulase and hemicellulase on lactose, achieving industry-leading enzyme activities.
- This study reports, for the first time, higher cellulase induction efficiency by lactose than cellulose in Trichoderma reesei.
- The enhanced performance is attributed to BGL1 overexpression and collateral mutations, leading to increased transcription of cellulase genes and related factors.
- Strain improvement using soluble carbon sources like lactose offers a pathway to cost-effective industrial enzyme production.