Inulin hydrolysis and citric acid production from inulin using the surface-engineered Yarrowia lipolytica displaying inulinase
The INU1 gene encoding exo-inulinase cloned from Kluyveromyces marxianus CBS 6556 was ligated into the surface display plasmid and expressed in the cells of the marine-derived yeast Yarrowia lipolytica which can produce citric acid. The expressed inulinase was immobilized on the yeast cells. The activity of the immobilized inulinase with 6 × His tag was found to be 22.6 U mg−1 of cell dry weight after cell growth for 96 h. The optimal pH and temperature of the displayed inulinase were 4.5 and 50 °C, respectively and the inulinase was stable in the pH range of 3–8 and in the temperature range of 0–50 °C. During the inulin hydrolysis, the optimal inulin concentration was 12.0% and the optimal amount of added inulinase was 181.6 U g−1 of inulin. Under such conditions, over 77.9% of inulin was hydrolyzed within 10 h and the hydrolysate contained main monosaccharides and disaccharides, and minor trisaccharides. During the citric acid production in the flask level, the recombinant yeast could produce 77.9 g L−1 citric acid and 5.3 g L−1 iso-citric acid from inulin while 68.9 g L−1 of citric acid and 4.1 g L−1 iso-citric acid in the fermented medium were attained within 312 h of the 2-L fermentation, respectively.
Journal: Metabolic Engineering - Volume 12, Issue 5, September 2010, Pages 469–476