Synthesis of β-Galactooligosaccharide Using Bifidobacterial β-Galactosidase Purified from Recombinant Escherichia coli .

Galactooligosaccharides (GOSs) are known to be selectively utilized by Bifidobacterium , which can bring about healthy changes of the composition of intestinal microflora. In this study, β-GOS were synthesized using bifidobacterial β-galactosidase (G1) purified from recombinant E. coli with a high GOS yield and with high productivity and enhanced bifidogenic activity. The purified recombinant G1 showed maximum production of β-GOSs at pH 8.5 and 45°C. A matrix-assisted laser desorption ionization time-of-flight mass spectrometry analysis of the major peaks of the produced β-GOSs showed MW of 527 and 689, indicating the synthesis of β-GOSs at degrees of polymerization (DP) of 3 and DP4, respectively. The trisaccharides were identified as β-D-galactopyranosyl-(1→4)- O -β-D -galactopyranosyl-(1→4)- O -β-D -glucopyranose, and the tetrasaccharides were identified as β-D-galactopyranosyl-(1→4)- O -β-D -galactopyranosyl- (1→4)- O -β-D -galactopyranosyl-(1→4)- O -β-D -glucopyranose. The maximal production yield of GOSs was as high as 25.3% (w/v) using purified recombinant β-galactosidase and 36% (w/v) of lactose as a substrate at pH 8.5 and 45°C. After 140 min of the reaction under this condition, 268.3 g/l of GOSs was obtained. With regard to the prebiotic effect, all of the tested Bifidobacterium except for B. breve grew well in BHI medium containing β-GOS as a sole carbon source, whereas lactobacilli and Streptococcus thermophilus scarcely grew in the same medium. Only Bacteroides fragilis , Clostridium ramosum , and Enterobacter cloacae among the 17 pathogens tested grew in BHI medium containing β-GOS as a sole carbon source; the remaining pathogens did not grow in the same medium. Consequently, the β-GOS are expected to contribute to the beneficial change of intestinal microbial flora.