High activity cellulase production by recombinant Trichoderma reesei ZU-02 with the enhanced cellobiohydrolase production

• Published in: Bioresource technology Vol. 144; pp. 693 - 697
• Main Authors: Fang, Hao, Xia, Liming
• Format: Journal Article
• Language: English
• Published: Kidlington Elsevier Ltd 01.09.2013; Elsevier
• Subjects: Agrobacterium - metabolism; Agronomy. Soil science and plant productions; Biological and medical sciences; Biotechnology; Cassettes; Cellobiohydrolase; Cellulase; Cellulose 1,4-beta-Cellobiosidase - biosynthesis; Enhanced expression; Enzymatic hydrolysis of corn stover; Food industries; Fundamental and applied biological sciences. Psychology; General agronomy. Plant production; Genes; Hydrolysis; Plasmids - metabolism; Recombinant; Recombinant Trichoderma reesei; Recombination, Genetic - genetics; Screening; Transformation, Genetic; Transformations; Trichoderma - enzymology; Trichoderma - genetics; Trichoderma reesei; Use and upgrading of agricultural and food by-products. Biotechnology; Use of agricultural and forest wastes. Biomass use, bioconversion; Recombinant Trichoderma reesei; Cellulase; Enhanced expression; Enzymatic hydrolysis of corn stover; Cellobiohydrolase; Monocotyledones; Recombination; Cereal crop; Straw; Fungi; Gramineae; Angiospermae; Fungi Imperfecti; Zea mays; Enzyme; Cellulose 1,4-β-cellobiosidase; Trichoderma reesei; Glycosylases; Hydrolysis; Cellulolytic enzyme; Glycosidases; Enzymatic digestion; Hydrolases; Spermatophyta; Corn by product
• ISSN: 0960-8524; 1873-2976
• DOI: 10.1016/j.biortech.2013.06.120

•The transformation rate of Trichoderma reesei was significantly increased.•Two steps of screening were designed for efficient transformant selection.•The filter paper activity of the cellulase was greatly improved.•The cellobiohydrolase production was notably enhanced.•The exo-exo-synergism played a role in enzymatic hydrolysis of lignocellulose. The cbh1 strong promoter was employed to over-express the cbh2 gene for enhancing cellobiohydrolase (CBH) production in Trichoderma reesei because CBH II component has higher specific activity than CBH I and is an important component in cellulase. The recombinant plasmid pCAMBIA1300-hph-PsCT containing strong expression cassette was constructed and transformed into T. reesei via optimized Agrobacterium-mediated transformation, producing 324 positive T. reesei transformants for the two steps of screening. Ten fast-growing T. reesei transformants were selected, amongst which C10 was found to have the highest filter paper activity 28.92±2.45IU/mL, 4.3-fold higher than that of ZU-02, 6.71±0.79IU/mL. C10 also has the highest cellobiohydrolase activity 122.44±7.42U/mL, 5.4 times higher than that of ZU-02, 22.49±2.27U/mL. The cellulase from C10 performed better (93.06±2.83%) than the one from ZU-02 in enzymatic hydrolysis because the exo–exo-synergism played a role.