Cloning and characterization of a new β-Glucosidase from a metagenomic library of Rumen of cattle feeding with Miscanthus sinensis

Abstract

BackgroundThe study on the second generation bio-fuel is a hot area of current research of renewable energy. Among series of key points in this area, the role of β-glucosidase in the degradation of intermediate gluco-oligosaccharides limits the rate of the complete saccharification of lignocellulose.ResultsIn this study, a new β-glucosidase gene, unglu135B12, which was isolated from a metagenomic library of rumen of cattle feeding with Miscanthus sinensis by the function-based screening, encodes a 779 amino acid polypeptide that contains a catalytic domain belonging to glycoside hydrolase family 3 (GH3). It was recombinantly expressed, purified and biochemically characterized. The recombinant β-glucosidase, unglu135B12, displayed optimum enzymatic activity at pH 5.0 at 38°C, and showed the highest specific activity of 2.5 × 103 U/mg under this optimal condition to p-nitrophenyl-β-D-glucopyranoside (pNPG), and its Km and Vmax values were 0.309 mmol/L and 7.292 μmol/min, respectively. In addition, the presence of Ca2+, K+, Na+ slightly improved β-glucosidase activity of unglu135B12 by about 5%, while about 10 ~ 85% loss of β-glucosidase activity was induced by addition of Mn2+, Fe3+, Zn2+, Cu2+. Interestingly, unglu135B12 was activated by glucose at the concentration lower than 40 mM.ConclusionsOur findings indicate that unglu135B12 is a new β-glucosidase derived from rumen of cattle, and it might be a potent candidate for saccharification of lignocellulose in industrial application.Electronic supplementary materialThe online version of this article (doi:10.1186/1472-6750-14-85) contains supplementary material, which is available to authorized users.

Identification and characterization of a novel β-glucosidase via metagenomic analysis of Bursaphelenchus xylophilus and its microbial flora

“…The unique properties of Cen502 are summarized as follows: Cen502 is characterized by high catalytic efficiency at high pH, as indicated by its optimum enzymatic activity at pH 8.0. The optimum pH of purified Cen502 was higher than most known β-glucosidases, which typically exhibit optimal pHs below 7.0 13 , 15 , 22 – 25 . Of note, Cen502 is active over a wide range of pHs, retaining 56% of its activity at pH 9.0, and is stable over the pH range 5.0–9.0.…”

“…The β-glucosidase from Fusarium oxysporum has optimal activity at pH 5.0, and the enzyme is stable over the pH range 4.0–7.0 24 . Likewise, the optimal pH of the β-glucosidase Unglu135B12 from cattle rumen is 5.0, and the enzyme is stable only in the pH range 5.0–6.0 25 . The β-glucosidase Ho BGLA from Halothermothrix orenii has maximal activity at pH 6.0 and is stable in the pH range 4.5–7.5 26 .…”

Catalytic properties, functional attributes and industrial applications of β-glucosidases

“…Most bioconversion is focused on the production of a sugar platform of simple sugars which can then biologically or chemically be converted into fuels (e.g., ethanol, butanol and hydrocarbons) (Cherubini 2010 ). Recently, Li et al ( 2014 ) cloned and characterized a β-glucosidase from rumen microbes of cattle feeding with Miscanthus sinensis and this plant is used as an ideal source of biofuel. Coating of β-glucosidase on polymer nanofibers was also found more efficient for cellulosic ethanol production (Lee et al 2010 ).…”

Gene cloning and expression of fungal lignocellulolytic enzymes from the rumen of gayal (<i>Bos frontalis</i>)

“…Hong et al (2006) have found that an enzyme with both b-glucosidase and bxylosidase activity showed a better cellulolytic efficiency. Li et al (2014) have found in ruminal microorganisms from B. taurus a novel b-glucosidase gene, which encodes for a protein, unglu135B12, with a molecular weight of 84.1 kDa. This protein has an optimal reaction temperature of 38∞C and an optimal pH of 5.0 and shows the highest similarity with proteins from Bacteroidetes.…”