Biosynthesis of lactosucrose by a new source of β-fructofuranosidase from Bacillus methanolicus LB-1

Lactosucrose (LS) is a prebiotic trisaccharide enzymatically synthesized by transglycosylation from lactose and sucrose with beneficial health effect. The β-fructofuranosidase used for synthesis of LS was produced from Bacillus methanolicus LB-1, which was isolated from traditional rice wine. A maximal yield of 8.63 U/mL of the enzyme was obtained by fermentation with B. methanolicus LB-1 under the optimized conditions: 10 g/L of glucose, 5 g/L of yeast extract, initial medium pH at 7.0, 37 °C, 24 h. The enzyme was purified and identified by ammonium sulfate fractional precipitation, Sephadex G-75 gel filtration chromatography and LC-MS, and SDS-PAGE of the purified enzyme showed a major protein band at 45 kDa. Biosynthesis of LS was performed using the purified β-fructofuranosidase, and production of LS reached 110 g/L under the optimized reaction conditions: pH at 7.0, 37 °C, 6.0 U/g sucrose of enzyme, 15% of sucrose, 15% of lactose, 28 h. HPLC analysis of the reaction products showed a distinct peak for LS at about 30 min of elution, confirming that B. methanolicus LB-1 β-fructofuranosidase had effective transfructosylation activity. Therefore, this new microbial source of β-fructofuranosidase may be a candidate with potential application prospect in biosynthesis of prebiotic LS.