Expression and characterization of β-1,3-1,4-glucanase of Aspergillus usamii in Escherichia coli and its application in sourdough bread making.

Abstract

A β-1,3-1,4-glucanase gene (Auglu12A) from Aspergillus usamii was successfully expressed in Escherichia coli BL21(DE3). The recombinant enzyme, reAuglu12A was efficiently purified using the one-step nickel-nitrilotriacetic acid affinity chromatography. The specific activity of reAuglu12A was 694.8U/mg, with an optimal temperature of 55°C and pH of 5.0. The reAuglu12A exhibited stability at temperatures up to 60°C and within the pH range of 4.0-5.5. The reAuglu12A hydrolytic activity was increased in the presence of metal ions, especially K+ and Na+ , whereas it exhibited a Km and Vmax of 8.35mg/mL and 1254.02µmol/min/mg, respectively, toward barley β-glucan at pH 5.0 and 55°C. The addition of reAuglu12A significantly increased the specific volume (p<0.05) and reduced crumb firmness and chewiness (p<0.05) of wheat-barley sourdough bread during a 7-day storage period compared to the control. Overall, the quality of wheat-barley sourdough bread was improved after incorporation of reAuglu12A (especially at 3000U/300g). These changes were attributed to the synergistic effect of acidification by sourdough and its metabolites which provided a conducive environment for the optimal action of reAuglu12A in the degradation of β-glucans of barley flour in sourdough. This stabilized the dough structure, thereby enhancing the quality, texture, and shelf life of the bread. These findings suggest that reAuglu12A holds promise as a candidate for β-glucanase application in the baking industry.