Enzymatic production of prebiotic fructo‐oligosteviol glycosides

Two-Step Production of Neofructo-Oligosaccharides Using Immobilized Heterologous Aspergillus terreus 1F-Fructosyltransferase Expressed in Kluyveromyces lactis and Native Xanthophyllomyces dendrorhous G6-Fructosyltransferase

“…Because we used a two-step consecutive process, the slowest partial reaction determined the overall speed. The results indicated that neoFOS production could be increased by optimizing enzyme activity, which could be achieved by enzyme immobilization and/or expression under the control of a strong inducible promoter [11,35]. The use of 1-FFT for the first catalytic step did not achieve the clear initiation of the neoseries using already-elongated FOS in any of the consecutive second catalytic steps, probably reflecting the higher affinity of the enzymes for sucrose rather than inulin-type oligosaccharides.…”

“…Because 1-FFT was expressed in K. lactis, the resulting mixture of FOS represented the combined activity of native 6-SFT and recombinant 1-FFT. However, inulin-type FOS remained dominant because the strong LAC4 promoter was used for 1-FFT expression [11].…”

“…Heterologous A. terreus 1-FFT expressed in K. lactis was tested in the pH range 4-8 and the temperature range 40-80 °C ( Figure 4a) based on previous findings [11]. Similarly, an equivalent design was used to study the effect of sucrose concentration (0.77-2.10 M) and reaction times ranging from 14 to 107 min ( Figure 4b) [11,13]. The significant reduced quadratic model predicted a local maximum FOS yield at pH 5.8 and 70 °C.…”

“…Kluyveromyces lactis strain GG799 was obtained from New England Biolabs (Frankfurt, Germany) and was used to express native 6-SFT and the integrated fructosyltransferase (1-FFT) gene from Aspergillus terreus NIH2624, which was described previously [11,12]. Xanthophyllomyces dendrorhous (DSM 5626) was obtained from the Leibniz Institute DSMZ German Collection of Microorganisms and Cell Cultures (Germany) and was used to express native G 6 -FFT.…”

“…FOS are short-chain fructans, mainly consisting of fructose units, with a low calorific value [4][5][6][7][8]. Their prebiotic effects reflect their ability to stimulate the growth of beneficial bifidobacteria and lactobacilli while inhibiting undesirable bacteria such as Clostridium perfringens [9][10][11][12]. The sensorial qualities of FOS are similar to sucrose, but they do not promote dental caries.…”