Development of a Chemically Defined Fermentation Medium for the Production of a New Recombinant Fructosyltransferase

The industrial-scale production of fructooligosaccharides from sucrose requires large quantities of the enzyme fructosyltransferase. An Aspergillus terreus fructosyltransferase was therefore expressed in Kluyveromyces lactis GG799 and secreted into the medium. K. lactis was cultivated in shaking flasks at 30°C and 250 rpm using either rich or chemically defined fermentation media. In order to limit the accumulation of unwanted side products that tend to form in rich media such as yeast extract peptone dextrose, a chemically defined FM22 medium was optimized in a two stages, focusing on biomass accumulation and enzyme production, respectively. A design-of-experiments strategy was used to screen for essential vitamins. A two-level fractional factorial design revealed that only biotin, nicotinic acid, pyridoxine and D-pantothenic acid were necessary for biomass accumulation, and an additional D-optimal design was used to optimize the concentration of inorganic salts (MgSO4, (NH4)2SO4, CaSO4, FeSO4 and KH2PO4) and the fermentation temperature. For enzyme production, the integrated LAC4 promoter was induced with galactose, which was provided in addition to glucose as the carbon source in the adapted FM22 medium. 