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Microbial β-glucosidases from cow rumen metagenome enhance the saccharification of lignocellulose in combination with commercial cellulase cocktail

Del Pozo, Mercedes V, Fernández-Arrojo, Lucía, Gil-Martínez, Jorge, Montesinos, Alejandro, Chernikova, Tatyana N, Nechitaylo, Taras Y, Waliszek, Agnes, Tortajada, Marta, Rojas, Antonia, Huws, Sharon A, Golyshina, Olga V, Newbold, Charles J, Polaina, Julio, Ferrer, Manuel, Golyshin, Peter N
Biotechnology for biofuels 2012 v.5 no.1 pp. 187
Trichoderma, bacteria, beta-glucosidase, biomass, corn stover, cows, dry matter content, endo-1,4-beta-glucanase, ethanol production, hydrolysis, lignocellulose, metagenomics, microbial communities, pH, polymers, rumen, saccharification, temperature
BACKGROUND: A complete saccharification of plant polymers is the critical step in the efficient production of bio-alcohols. Beta-glucosidases acting in the degradation of intermediate gluco-oligosaccharides produced by cellulases limit the yield of the final product. RESULTS: In the present work, we have identified and then successfully cloned, expressed, purified and characterised 4 highly active beta-glucosidases from fibre-adherent microbial community from the cow rumen. The enzymes were most active at temperatures 45–55°C and pH 4.0-7.0 and exhibited high affinity and activity towards synthetic substrates such as p-nitrophenyl-beta-D-glucopyranoside (pNPbetaG) and pNP-beta-cellobiose, as well as to natural cello-oligosaccharides ranging from cellobiose to cellopentaose. The apparent capability of the most active beta-glucosidase, herein named LAB25g2, was tested for its ability to improve, at low dosage (31.25 units g⁻¹ dry biomass, using pNPbetaG as substrate), the hydrolysis of pre-treated corn stover (dry matter content of 20%; 350 g glucan kg⁻¹ dry biomass) in combination with a beta-glucosidase-deficient commercial Trichoderma reseei cellulase cocktail (5 units g⁻¹ dry biomass in the basis of pNPbetaG). LAB25g2 increased the final hydrolysis yield by a factor of 20% (44.5 ± 1.7% vs. 34.5 ± 1.5% in control conditions) after 96–120 h as compared to control reactions in its absence or in the presence of other commercial beta-glucosidase preparations. The high stability (half-life higher than 5 days at 50°C and pH 5.2) and 2–38000 fold higher (as compared with reported beta-glucosidases) activity towards cello-oligosaccharides may account for its performance in supplementation assays. CONCLUSIONS: The results suggest that beta-glucosidases from yet uncultured bacteria from animal digestomes may be of a potential interest for biotechnological processes related to the effective bio-ethanol production in combination with low dosage of commercial cellulases.