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.
Highlights
The study on the second generation bio-fuel is a hot area of current research of renewable energy
The conversion of the most abundant part of fiber, cellulose, into bio-fuel will depend on the synergistical hydrolysis of enzymes at least including three main cellulases: endo-β-1,4-glucanase (EC3.2.1.4), cellobiohydrolase (EC3.2.1.91) and β-glucosidase (EC3.2.1.21) [5]. β-glucosidase is widely distributed in nature [6,7], which catalyses the cleavage of the glycosidic bonds existing in disaccharides, oligosaccharides and alkyl- or aryl- β-glucosides [8]
Β-glucosidase does not act on cellulose directly, it is of great importance in cellulose deconstruction by eliminating cellobiose inhibition on endoglucanases and exoglucanases, allowing the cellulolytic enzymes to function more efficiently [5,9]
Summary
The study on the second generation bio-fuel is a hot area of current research of renewable energy. The study on the second generation bio-fuel, which can be constantly derived from the waste of agriculture and forestry and the widespread distribution of non-food plants, is a hot area of current research of renewable energy [1,2]. Among non-food plants, as the raw material of bio-fuel, Miscanthus sinensis has been focused on in recent years as an ideal bio-source plant. This fiber-rich plant is widely distributed around South East Asia, and it’s a common specie in China [3]. The activity of β-glucosidase is considered as the rate-limiting factor in cellulose degradation
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