BackgroundA 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.ResultsIn 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.ConclusionsThe 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.
Raw glycerol is an industrial byproduct from biodiesel production and is one of the most promising substrates for 2,3-butanediol (2,3-BD) production; however, 2,3-BD is not yet produced by fermentation from glycerol on a commercial scale due to poor process economics. Class 1 microorganism collections were screened and Raoultella planticola strain CECT 843 proved to be the best 2,3-BD producer, achieving (23.3 ± 1.4) g 2,3-BD per L and a yield of 36% (g 2,3-BD per g glycerol). To further increase product concentration and yield, R. planticola CEC T843 was subjected to random mutagenesis using ultra-violet (UV) light and ethyl methane sulfonate (EMS). Two mutant strains were found to produce at least 30% more 2,3-BD than the wild type: R. planticola IA1 [(30.8 ± 3.9) g 2,3-BD per L and 49% yield] and R. planticola IIIA3 [(30.5 ± 0.4) g 2,3-BD per L and 49% yield].
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