Highly Efficient Expression of Tannase in Aspergillus niger and Its Enzymatic Properties

Abstract

The recombinant strains of Aspergillus niger with high expression of tannase were constructed by means of genetic engineering. The activity and enzymatic properties of the recombinant enzyme were studied to improve the expression of tannase and better realize the important role of tannase in tea beverage, feed and other industries. GlaA multi-copy strong promoter PglaA6R and signal peptide SglaA were integrated to construct tannase gene AnTan Aspergillus niger recombinant expression vector pSZHG6R-AnTan with the endogenous highly expressed glaA gene site as the integration target site, and the Aspergillus niger pure recombinant strain A1 of tannase was obtained by agrobacterium-mediated transformation of Aspergillus niger. The results of polyacrylamide gel electrophoresis (SDS-PAGE) detection results showed that the size of the target protein was about 76 kDa. The maximum enzyme activity reached 134.36 U·mL−1 when 10% dextrin was used as carbon source for shaking flask fermentationat to 11 days, which was about 192 times of the host strain. The study of enzymatic properties showed that the optimal temperature was 50 ℃ and the thermal stability was poor at high temperature, the optimal pH was 7.0. K+ and Ca2+ had no effect on recombinant enzyme, Mg2+ had weak promoting effect on recombinant enzyme, Zn2+ had obvious promoting effect on recombinant enzyme, Cu2+ and Fe2+ had weak inhibition on recombinant enzyme, Mn2+and EDTA had obvious inhibitory effect on recombinant enzyme. The results provide a valuable reference for further optimizing the secretion and expression of tannase.

Keywords

• tannase
• aspergillus niger
• high expression
• enzymology properties
• recombinant strains
• genetic engineering