Cloning, expression and characterization of an β-1,3(4)-glucanase from Microbulbifer arenaceous

Abstract:

[Background] β-glucan is a non-starch polysaccharide, widely existed in nature, especially in cell wall of cereal plants. β-glucanase can catalyze β-glucan into β-glucooligosaccharides, it plays an important role in food, feed and papermaking industry. [Objective] gene of β-1,3(4)-glucanase from marine bacteria Microbulbifer arenaceous was cloned and expressed in Escherichia coli, and its enzymatic characteristics and hydrolysis properties were studied. [Methods] The β-1,3(4)-glucanase gene (MaGlu16A) was cloned from the genomic DNA of Microbulbifer arenaceous and the recombinant plasmid (pET-28a-MaGlu16A) was constructed. The recombination strain was successfully expressed in E. coli BL21(DE3), and purified by Ni-NTA affinity chromatography, then enzymatic characteristics was studied. [Results] The optimal pH of MaGlu16A was 6.0 and optimal temperature was 40 °C. It was stable between pH 5.0 and 10.5, and below 35 °C. The enzyme showed obvious resistance to EDTA, and it could maintain 99.3% and 82.5% activity at the concentration of 1 mmol/L and 10 mmol/L. In addition, MaGlu16A showed broad substrate specificity. It could hydrolyze not only curdlan and laminarin but also barley β-glucan, lichenan, oat β-glucan, and yeast β-glucan, whose hydrolytic products were mainly glucose, disaccharide, trisaccharide and tetragasaccharide. [Conclusion] Cloning, expression and characterization of β-1,3(4)-glucanase from marine bacteria Microbulbifer arenaceous could provide a basis for the exploration of β-glucanase and preparation of β-glucooligosaccharides.