High-level secretory expression of Aspergillus exo-inulinase and its use in the preparation of fructose syrup from inulin

• Published in: Journal of molecular catalysis. B, Enzymatic Enzymatic, 2016-11, Vol.133, p.S543-S551
• Main Authors: Chen, Guang-Jun, Yang, Jiang-Ke, Peng, Xiao-Bo, He, Jing-Ren
• Format: Article
• Language: English
• Subjects: Codon optimization; Endoplasmic reticulum protein; Fructose; Inulin; Inulinase
• ISSN: 1381-1177; 1873-3158
• DOI: 10.1016/j.molcatb.2017.09.001

[Display omitted] •De novo codon design improved the expression of exo-inulinase gene.•Co-expression with HAC1 improved the secretory expression of exo-inulinase gene.•Inulinase gene was high-level expressed in a pilot-scale bioreactor.•Reaction parameters optimization facilitated the hydrolysis of inulin by inulinase.•Exo-inulinase could hydrolyze inulin into syrup with 95% fructose. Inulin is a type of fructose polymer that is commonly present in plants as a storage carbohydrate. Enzymatic hydrolysis of inulin via exo-inulinase to produce fructose is an efficient, green and state-of-the-art technique. To achieve the high-level secretory expression of inulinase and to realize enzymatic preparation of fructose syrup from inulin, an Aspergillus exo-inulinase gene inu was codon-optimized and co-expressed with the endoplasmic reticulum secretion protein in Pichia cells. After inducible expression in a 500-L pilot scale bioreactor, the inulinase activity of the recombinant strains reached 10,480U/mL of cultivation broth. Next, according to the determined enzymatic characteristics of inulinase INU, we optimized the parameters for inulinase to hydrolyse inulin. Under the optimal condition of the enzyme/inulin ratio of 5000U/g, 15% substrate and an incubation temperature of 50°C for 4h, the hydrolysis ratio of inulin reached 100%. The hydrolysis products of inulin contain two components, 95% fructose, and 5% glucose. This study has fulfilled the scaled-up production of inulinase and facilitated its industrial application for enzymatic preparation of fructose from inulin.