Enhanced production of cellobiohydrolases in Trichoderma reesei and evaluation of the new preparations in biofinishing of cotton

• Published in: Journal of biotechnology 2005-03, Vol.116 (3), p.305-317
• Main Authors: Miettinen-Oinonen, Arja, Paloheimo, Marja, Lantto, Raija, Suominen, Pirkko
• Format: Article
• Language: English
• Subjects: Biofinishing; Biological and medical sciences; Biotechnology; Cellobiohydrolase; cellulose 1,4-beta-cellobiosidase; Cellulose 1,4-beta-Cellobiosidase - biosynthesis; Cellulose 1,4-beta-Cellobiosidase - chemistry; Cellulose 1,4-beta-Cellobiosidase - genetics; cotton; Cotton Fiber - classification; Cotton Fiber - methods; enzyme activity; Fundamental and applied biological sciences. Psychology; gene overexpression; Genetic Enhancement - methods; genetically engineered microorganisms; Hypocrea jecorina; Overproduction; processing technology; promoter regions; Protein Engineering - methods; Recombinant Proteins - biosynthesis; Recombinant Proteins - chemistry; T. reesei; Trichoderma - enzymology; Trichoderma - genetics
• ISSN: 0168-1656; 1873-4863
• DOI: 10.1016/j.jbiotec.2004.10.017

In the search for suitable cellulase combinations for industrial biofinishing of cotton, five different types of Trichoderma reesei strains were constructed for elevated cellobiohydrolase production: CBHI overproducers with and without endoglucanase I (EGI), CBHII overproducers with and without endoglucanase II (EGII) and strains overproducing both CBHI and CBHII without the major endoglucanases I and II. One additional copy of cbh1 gene increased production of CBHI protein 1.3-fold, and two copies 1.5-fold according to ELISA (enzyme-linked immunosorbent assay). The level of total secreted proteins was increased in CBHI transformants as compared to the host strain. One copy of the cbh2 expression cassette in which the cbh2 was expressed from the cbh1 promoter increased production of CBHII protein three- to four-fold when compared to the host strain. T. reesei strains producing elevated amounts of both CBHI and CBHII without EGI and EGII were constructed by replacing the egl1 locus with the coding region of the cbh1 gene and the egl2 locus with the coding region of cbh2. The cbh1 was expressed from its own promoter and the cbh2 gene using either the cbh1 or cbh2 promoter. Production of CBHI by the CBH-transformants was increased up to 1.6-fold and production of CBHII up to 3.4-fold as compared with the host strain. Approximately similar amounts of CBHII protein were produced by using cbh1 or cbh2 promoters. When the enzyme preparation with elevated CBHII content was used in biofinishing of cotton, better depilling and visual appearance were achieved than with the wild type preparation; however, the improvement was not as pronounced as with preparations with elevated levels of endoglucanases (EG).