Red onion is a popular ingredient in many Thai dishes and has recently been promoted for commercial cultivation. In this study, inulin-fructooligosaccharides (inulin-FOSs) were extracted from red onions in a simplified extraction method. The extract contained 24.00 ± 0.38 g/L free glucose, fructose and sucrose, while the level of FOSs was recorded at 74.0 ± 2.80 g/L with a degree of polymerization of 4.1. The extract was resistant to simulated gastrointestinal conditions, while selectively promoting probiotic lactobacilli. These outcomes resulted in inhibitory effects against various pathogenic bacteria. The in vitro batch culture fermentation of the extract by natural mixed culture indicated that an unknown sugar identified as neokestose was more rapidly fermented than 1-kestose and other longer-chain inulin-FOSs. Notably, neokestose selectively encouraged a bifidogenic effect, specifically in terms of the growth of Bifidobacteirum breve, which is an infant-type probiotic bacterium. This is the first report to state that neokestose could selectively enhance the bifidogenic effect. In summary, inulin-FOSs extract should be recognized as a multifunctional ingredient that can offer benefits in food and pharmaceutical applications.

Glycoconjugates are the most diverse biomolecules of life. Mostly located at the cell surface, they translate into cell-specific “barcodes” and offer a vast repertoire of functions, including support of cellular physiology, lifestyle, and pathogenicity. Functions can be fine-tuned by non-carbohydrate modifications on the constituting monosaccharides. Among these modifications is pyruvylation, which is present either in enol or ketal form. The most commonly best-understood example of pyruvylation is enol-pyruvylation of N-acetylglucosamine, which occurs at an early stage in the biosynthesis of the bacterial cell wall component peptidoglycan. Ketal-pyruvylation, in contrast, is present in diverse classes of glycoconjugates, from bacteria to algae to yeast—but not in humans. Mild purification strategies preventing the loss of the acid-labile ketal-pyruvyl group have led to a collection of elucidated pyruvylated glycan structures. However, knowledge of involved pyruvyltransferases creating a ring structure on various monosaccharides is scarce, mainly due to the lack of knowledge of fingerprint motifs of these enzymes and the unavailability of genome sequences of the organisms undergoing pyruvylation. This review compiles the current information on the widespread but under-investigated ketal-pyruvylation of monosaccharides, starting with different classes of pyruvylated glycoconjugates and associated functions, leading to pyruvyltransferases, their specificity and sequence space, and insight into pyruvate analytics.

Diverse members of the Bacteroidetes phylum have general protein O-glycosylation systems that are essential for processes such as host colonization and pathogenesis. Here, we analyzed the function of a putative fucosyltransferase (FucT) family that is widely encoded in Bacteroidetes protein O-glycosylation genetic loci. We studied the FucT orthologs of three Bacteroidetes species—Tannerella forsythia, Bacteroides fragilis, and Pedobacter heparinus. To identify the linkage created by the FucT of B. fragilis, we elucidated the full structure of its nine-sugar O-glycan and found that l-fucose is linked β1,4 to glucose. Of the two fucose residues in the T. forsythia O-glycan, the fucose linked to the reducing-end galactose was shown by mutational analysis to be l-fucose. Despite the transfer of l-fucose to distinct hexose sugars in the B. fragilis and T. forsythia O-glycans, the FucT orthologs from B. fragilis, T. forsythia, and P. heparinus each cross-complement the B. fragilis ΔBF4306 and T. forsythia ΔTanf_01305 FucT mutants. In vitro enzymatic analyses showed relaxed acceptor specificity of the three enzymes, transferring l-fucose to various pNP-α-hexoses. Further, glycan structural analysis together with fucosidase assays indicated that the T. forsythia FucT links l-fucose α1,6 to galactose. Given the biological importance of fucosylated carbohydrates, these FucTs are promising candidates for synthetic glycobiology.

By using molecular dynamics simulations with an efficient enhanced sampling technique and in combination with nuclear magnetic resonance (NMR) spectroscopy quantitative structural information on $\alpha$ -2,8-linked sialic acids is presented. We used a bottom-up approach to obtain a set of larger ensembles for tetra- and deca-sialic acid from model dimer and trimer systems that are in agreement with the available J-coupling constants and nuclear Overhauser effects. The molecular dynamic (MD) simulations with enhanced sampling are used to validate the force field used in this study for its further use. This empowered us to couple NMR observables in the MD framework via J-coupling and distance restraining simulations to obtain conformations that are supported by experimental data. We used these conformations in thermodynamic integration and one-step perturbation simulations to calculate the free-energy of suggested helical conformations. This study brings most of the available NMR experiments together and supplies information to resolve the conflict on the structures of poly- $\alpha$ -2,8-linked sialic acid.

UDP-Gal: glycoprotein-N-acetylgalactosamine β-1,3-galactosyltransferase (T-synthase, EC 2.4.1.122) catalyses the transfer of the monosaccharide galactose from UDP-Gal to GalNAc-Ser/Thr, synthesizing the core 1 mucin type O-glycan. Such glycans play important biological roles in a number of recognition processes. The crucial role of these glycans is acknowledged for mammals, but a lot remains unknown regarding invertebrate and especially mollusc O-glycosylation. Although core O-glycans have been found in snails, no core 1 β-1,3-galactosyltransferase has been described so far. Here, the sequence of the enzyme was identified by a BlastP search of the NCBI Biomphalaria glabrata database using the human T-synthase sequence (NP_064541.1) as a template. The obtained gene codes for a 388 amino acids long transmembrane protein with two putative N-glycosylation sites. The coding sequence was synthesised and expressed in Sf9 cells. The expression product of the putative enzyme displayed core 1 β-1,3-galactosyltransferase activity using pNP-α-GalNAc as the substrate. The enzyme showed some sequence homology (49.40% with Homo sapiens, 53.69% with Drosophila melanogaster and 49.14% with Caenorhabditis elegans) and similar biochemical parameters with previously characterized T-synthases from other phyla. In this study we present the identification, expression and characterisation of the UDP-Gal: glycoprotein-N-acetylgalactosamine β-1,3-galactosyltransferase from the fresh-water snail Biomphalaria glabrata, which is the first cloned T-synthase from mollusc origin.