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Direct and efficient production of ethanol from cellulosic material with a yeast strain displaying cellulolytic enzymes

Author:
Fujita, Y., Takahashi, S., Ueda, M., Tanaka, A., Okada, H., Morikawa, Y., Kawaguchi, T., Arai, M., Fukuda, H., Kondo, A.
Source:
Applied and environmental microbiology 2002 v.68 no.10 pp. 5136-5141
ISSN:
0099-2240
Subject:
Hypocrea jecorina, Saccharomyces cerevisiae, genetically modified organisms, gene transfer, Aspergillus aculeatus, beta-glucosidase, genes, gene expression, beta-glucanase, hydrolysis, enzyme activity, ethanol production, saccharification
Abstract:
For direct and efficient ethanol production from cellulosic materials, we constructed a novel cellulose-degrading yeast strain by genetically codisplaying two cellulolytic enzymes on the cell surface of Saccharomyces cerevisiae. By using a cell surface engineering system based on alpha-agglutinin, endoglucanase II (EGII) from the filamentous fungus Trichoderma reesei QM9414 was displayed on the cell surface as a fusion protein containing an RGSHis6 (Arg-Gly-Ser-His6) peptide tag in the N-terminal region. EGII activity was detected in the cell pellet fraction but not in the culture supernatant. Localization of the RGSHis6-EGII-alpha-agglutinin fusion protein on the cell surface was confirmed by immunofluorescence microscopy. The yeast strain displaying EGII showed significantly elevated hydrolytic activity toward barley beta-glucan, a linear polysaccharide composed of an average of 1,200 glucose residues. In a further step, EGII and beta-glucosidase 1 from Aspergillus aculeatus No. F-50 were codisplayed on the cell surface. The resulting yeast cells could grow in synthetic medium containing beta-glucan as the sole carbon source and could directly ferment 45 g of beta-glucan per liter to produce 16.5 g of ethanol per liter within about 50 h. The yield in terms of grams of ethanol produced per gram of carbohydrate utilized was 0.48 g/g, which corresponds to 93.3% of the theoretical yield. This result indicates that efficient simultaneous saccharification and fermentation of cellulose to ethanol are carried out by a recombinant yeast cells displaying cellulolytic enzymes.
Agid:
1409745