Synthesis and characterization of lactose and lactulose derived oligosaccharides by glucansucrase and trans-sialidase enzymes

Human milk oligosaccharides (hMOS) have an essential role in infants’ health by stimulating growth of health-beneficial gut bacteria for the human host. These hMOS also directly reduce pathogenic microbial infections by serving as antiadhesives against pathogenic microbes, and stimulating immune responses. Many babies have limited access to human milk, and alternative sources for hMOS are not available in nature. Infant formula with the commercial prebiotics lack pathogen exclusion and immune and barrier modulating effects exerted by hMOS. Development of efficient routes for synthesis of hMOS or structurally/functionally effective mimics is currently focus of attention. At present, whole cell biosynthetic routes and single/multiple enzyme biocatalytic systems for synthesis of hMOS (mimics) are studied. In this project, the glucansucrases Gtf180-ΔN and GtfA-ΔN from Lactobacillus reuteri were used to glucosylate lactose or GOS. The mixture of glucosylated lactose derivatives obtained showed selective effects on growth of various gut bacteria, including lactobacilli, bifidobacteria and commensal bacteria. Mutational analysis of Gtf180-ΔN revealed that three amino acid residues (N1029, W1065 and Q1140) play important roles in determining linkage specificity in lactose trans-glycosylation. Finally, trans-sialidase from Trypanosoma cruzi was used to transfer sialic acid to mixtures of glucosylated-lactose, galactosylated-lactulose and galacto-oligosaccharide (Vivinal GOS) molecules. The newly synthesized galactose-containing oligosaccharides and sialylated oligosaccharides hold strong potential for further applications as hMOS mimics. Glucansucrase and trans-sialidase enzymes are promising tools as biocatalysts for efficient synthesis of new functional oligosaccharides