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Obesity has become a global epidemic. Sargassum fusiforme fucoidan (Fuc) is a group of water-soluble heteropolysaccharides that exhibits a wide range of medicinal functions. It consists of l-fucose and sulfate groups, with l-fucose as the main monosaccharide. This study investigated the therapeutic effects of Fuc on diet-induced obesity (DIO) in C57BL/6J female mice. Fuc significantly alleviated obesity in mice induced by high-fat high-fructose (HFHF) feeding, inhibiting body weight gain, reducing fat accumulation, and improving hepatic steatosis. In addition, Fuc significantly improved glucose tolerance and insulin sensitivity by enhancing the phosphorylation level of AKT (at Ser473) in the adipose tissues. Mechanistically, although Fuc did not decrease the energy intake in DIO mice, it significantly increased the energy expenditure by up-regulating the expression of uncoupling protein 1 (UCP1) in the adipose tissues. Notably, Fuc also improved the obesity-driven dysbiosis of gut microbiota and decreased the relative abundance of the obesity-related intestinal bacteria. However, Fuc was unable to alleviate DIO-induced metabolic disorders in pseudo-sterile mice. Our findings suggested that Fuc might remodel gut microbiota and exert its weight loss and hypolipidemic effects by increasing the energy expenditure, thus providing a novel perspective for treating obesity and related complications.

In this work, we reported an in situ exopolysaccharide (in situ-EPS1) containing rare fucose produced by Lactobacillus helveticus MB2-1 in Sayram ketteki yoghurt, which made it unique. Its fine structure was characterized by GPC, HPLC, FT-IR, GC-MS

Early weaning stress (EWS) in piglets is associated with intestinal dysfunction. Here, utilizing a pig EWS model to mimic early-life stress (ELS) in humans, we investigated the mechanism of ELS-induced intestinal diseases through integrated multi-omics analyses of proteome, glycome, and microbiome. Our results demonstrated that EWS resulted in disrupted the ileal barrier integrity by reducing tight junction-related gene expression and interfering with cell-cell adhesion paralleled the increased proportion of pathogens such as Escherichia_Shigella and Helicobacter. Furthermore, Proteome data revealed that the accumulation of unfolded proteins and insufficient unfolded protein response (UPR) process caused by EWS led to ER stress. Data from proteome and glycome found that EWS induced aberrant mucin O-glycans, including truncated glycans, reduction in acidic glycans, and increased in fucosylated glycans. In addition, correlation test by taking fucose and inflammatory response into account suggested that enhancement of fucose expression might be a compensatory host response. Taken together, these results extend the comprehensive knowledge of the detrimental impacts and pathogenesis of EWS and help to provide intervention targets for ELS-induced intestinal diseases in the future.

Polysaccharides coded as CP were extracted from Athyrium Multidentatum (Doll.) Ching and then fractionated into five fractions (FP-1, FP-2, FP-3, FP-4 and FP-5). A purified polysaccharide designated as FP-3-4 was prepared from FP-3 by Sephadex G-100 column chromatography. Chemical analysis disclosed that CP and these fractions were heteropolysaccharides and mainly composed of glucose, galactose, arabinose, mannose, rhamnose, xylose, fucose, ribose and uronic acid with different molar ratios. They presented different images of SEM. FP-3-4 was highly branched polymers with sixteen types of linkages. The in vitro immunomodulatory results stated that CP and these fractions could promote macrophage proliferation, enhance macrophage phagocytosis and increase the production of NO, TNF-α, IFN-γ, IL-1β, IL-6, IL-10 and IL-2, indicating remarkable immune enhancement activities. RNA sequencing analysis revealed that CP and FP-3 induced macrophage activation mainly through MAPK and alternative NF-κΒ signaling pathways via CD14/TLR4 and Dectin-2 receptors, which were verified by RT-qPCR and western blot.

B cell superantigens crosslink conserved domains of B cell receptors (BCRs) and cause dysregulated, polyclonal B cell activation irrespective of normal BCR-antigen complementarity. The cells typically succumb to activation-induced cell death, which can impede the adaptive immune response and favor infection. In the present study, we demonstrate that the fucose-binding lectin of Burkholderia ambifaria, BambL, bears functional resemblance to B cell superantigens. By engaging surface glycans, the bacterial lectin activated human peripheral blood B cells, which manifested in the surface expression of CD69, CD54 and CD86 but became increasingly cytotoxic at higher concentrations. The effects were sensitive to BCR pathway inhibitors and excess fucose, which corroborates a glycan-driven mode of action. Interactome analyses in a model cell line suggest BambL binds directly to glycans of the BCR and regulatory coreceptors. In vitro, BambL triggered BCR signaling and induced CD19 internalization and degradation. Owing to the lectin’s six binding sites, we propose a BCR activation model in which BambL functions as a clustering hub for receptor glycans, modulates normal BCR regulation, and induces cell death through exhaustive activation. Supplementary Information The online version contains supplementary material available at 10.1007/s00018-021-04009-z.

Background In endothelial cells, phospholipase C (PLC) β1-activated Ca2+ is a crucial second messenger for the signaling pathways governing angiogenesis. PLCβ1 is inactivated by complexing with an intracellular protein called translin-associated factor X (TRAX). This study demonstrates specific interactions between Globo H ceramide (GHCer) and TRAX, which highlight a new angiogenic control through PLCβ1 activation. Methods Globo-series glycosphingolipids (GSLs), including GHCer and stage-specific embryonic antigen-3 ceramide (SSEA3Cer), were analyzed using enzyme-linked immunosorbent assay (ELISA) and Biacore for their binding with TRAX. Angiogenic activities of GSLs in human umbilical vein endothelial cells (HUVECs) were evaluated. Molecular dynamics (MD) simulation was used to study conformations of GSLs and their molecular interactions with TRAX. Fluorescence resonance energy transfer (FRET) analysis of HUVECs by confocal microscopy was used to validate the release of PLCβ1 from TRAX. Furthermore, the in vivo angiogenic activity of extracellular vesicles (EVs) containing GHCer was confirmed using subcutaneous Matrigel plug assay in mice. Results The results of ELISA and Biacore analysis showed a stable complex between recombinant TRAX and synthetic GHCer with Kd of 40.9 nM. In contrast, SSEA3Cer lacking a fucose residue of GHCer at the terminal showed ~ 1000-fold decrease in the binding affinity. These results were consistent with their angiogenic activities in HUVECs. The MD simulation indicated that TRAX interacted with the glycan moiety of GHCer at amino acid Q223, Q219, L142, S141, and E216. At equilibrium the stable complex maintained 4.6 ± 1.3 H-bonds. TRAX containing double mutations with Q223A and Q219A lost its ability to interact with GHCer in both MD simulation and Biacore assays. Removal of the terminal fucose from GHCer to become SSEA3Cer resulted in decreased H-bonding to 1.2 ± 1.0 by the MD simulation. Such specific H-bonding was due to the conformational alteration in the whole glycan which was affected by the presence or absence of the fucose moiety. In addition, ELISA, Biacore, and in-cell FRET assays confirmed the competition between GHCer and PLCβ1 for binding to TRAX. Furthermore, the Matrigel plug assay showed robust vessel formation in the plug containing tumor-secreted EVs or synthetic GHCer, but not in the plug with SSEA3Cer. The FRET analysis also indicated the disruption of colocalization of TRAX and PLCβ1 in cells by GHCer derived from EVs. Conclusions Overall, the fucose residue in GHCer dictated the glycan conformation for its complexing with TRAX to release TRAX-sequestered PLCβ1, leading to Ca2+ mobilization in endothelial cells and enhancing angiogenesis in tumor microenvironments.

Human milk oligosaccharides (HMOs) and its most abundant component, 2’-Fucosyllactose (2’-FL), are known to be immunomodulatory. Previously, it was shown that HMOs and 2’-FL bind to the C-type lectin receptor DC-SIGN. Here we show, using a ligand-receptor competition assay, that a whole mixture of HMOs from pooled human milk (HMOS) and 2’-FL inhibit the binding of the carbohydrate-binding receptor DC-SIGN to its prototypical ligands, fucose and the oligosaccharide Lewis-B, (Leb) in a dose-dependent way. Interestingly, such inhibition by HMOS and 2’-FL was not detected for another C-type lectin, Langerin, evolutionary similar to DC-SIGN. The cell-ligand competition assay using DC-SIGN expressing cells confirmed that 2’-FL inhibits the binding of DC-SIGN to Leb. Molecular dynamics (MD) simulations show that 2’-FL exists in a preorganized bioactive conformation before binding to DC-SIGN and this conformation is retained after binding to DC-SIGN. Leb has more flexible conformations and utilizes two binding modes, which operate one at a time via its two fucoses to bind to DC-SIGN. 2’-FL may have a reduced entropic penalty due to its preorganized state compared to Leb, and it has lower binding enthalpy, suggesting better binding to DC-SIGN. Thus, due to the better binding to DC-SIGN, 2’-FL may replace Leb from its binding pocket in DC-SIGN. MD simulations also showed that 2’-FL does not bind to Langerin. Our studies confirm 2’-FL as a specific ligand for DC-SIGN and suggest that 2’-FL can replace other DC-SIGN ligands from its binding pocket during ligand-receptor interactions in possible immunomodulatory processes.

The conformational properties of carbohydrates can contribute to protein structure directly through covalent conjugation in the cases of glycoproteins and proteoglycans and indirectly in the case of transmembrane proteins embedded in glycolipid-containing bilayers. However, there continue to be significant challenges associated with experimental structural biology of such carbohydrate-containing systems. All-atom explicit-solvent molecular dynamics simulations provide a direct atomic resolution view of biomolecular dynamics and thermodynamics, but the accuracy of the results depends on the quality of the force field parametrization used in the simulations. A key determinant of the conformational properties of carbohydrates is ring puckering. Here, we applied extended system adaptive biasing force (eABF) all-atom explicit-solvent molecular dynamics simulations to characterize the ring puckering thermodynamics of the ten common pyranose monosaccharides found in vertebrate biology (as represented by the CHARMM carbohydrate force field). The results, along with those for idose, demonstrate that the CHARMM force field reliably models ring puckering across this diverse set of molecules, including accurately capturing the subtle balance between 4C1 and 1C4 chair conformations in the cases of iduronate and of idose. This suggests the broad applicability of the force field for accurate modeling of carbohydrate-containing vertebrate biomolecules such as glycoproteins, proteoglycans, and glycolipids.

Enzymatic hydrolysis of α-L-fucose from fucosylated glycoconjugates is consequential in bacterial infections and the neurodegenerative lysosomal storage disorder fucosidosis. Understanding human α-L-fucosidase catalysis, in an effort toward drug design, has been hindered by the absence of three-dimensional structural data for any animal fucosidase. Here, we have used cryoelectron microscopy (cryo-EM) to determine the structure of human lysosomal α-L-fucosidase (FucA1) in both an unliganded state and in complex with the inhibitor deoxyfuconojirimycin. These structures, determined at 2.49 Å resolution, reveal the homotetrameric structure of FucA1, the architecture of the catalytic center, and the location of both natural population variations and disease-causing mutations. Furthermore, this work has conclusively identified the hitherto contentious identity of the catalytic acid/base as aspartate-276, representing a shift from both the canonical glutamate acid/base residue and a previously proposed glutamate residue. These findings have furthered our understanding of how FucA1 functions in both health and disease.

The effects of water activity (aw), pH, and temperature on transglycosylation activity of α-L-fucosidase from Thermotoga maritima in the synthesis of fucosylated oligosaccharides were evaluated using different water-organic cosolvent reaction systems. The optimum conditions of transglycosylation reaction were the pH range between 7 and 10 and temperature 90–95 °C. The addition of organic cosolvent decreased α-L-fucosidase transglycosylation activity in the following order: acetone > dimethyl sulfoxide (DMSO) > acetonitrile (0.51 > 0.42 > 0.18 mM/h). However, the presence of DMSO and acetone enhanced enzyme-catalyzed transglycosylation over hydrolysis as demonstrated by the obtained transglycosylation/hydrolysis rate (rT/H) values of 1.21 and 1.43, respectively. The lowest rT/H was calculated for acetonitrile (0.59), though all cosolvents tested improved the transglycosylation rate in comparison to a control assay (0.39). Overall, the study allowed the production of fucosylated oligosaccharides in water-organic cosolvent reaction media using α-L-fucosidase from T. maritima as biocatalyst.

The testis of bamboo shark is characterized by diametric development leading to zonation architecture. Here, we investigated the staining pattern of 12 lectins in 6 groups of differential binding specificities within the germ, somatic, and interstitial cells of each zone. The neutral mucopolysaccharides appeared in the interstitial tissue in all the zones and became more significant in the spermatozoal–Sertoli cell junction. The cellular localization of the lectins varies in testicular zones and cell types. There was a gradual increase in glycosylation toward the degenerative zone. The increased intensity of most lectins in the interstitial cells indicates the association of glycoconjugates in their androgen-secreting activity. Statistical analyses showed a significant correlation between the groups of lectins and each lectin used, stronger response to lectins in the interstitial cells (ICs) than other cell types. Moreover, the response to glucosamine (GlcNAc), galactosamine (GalNAc), and fucose tended to be higher than glucose and galactose. Furthermore, the intensity of response was increased toward the degenerative zone. In addition, we can use peanut agglutinin (PNA) as an acrosomal marker in combination with other marker proteins for studying shark spermatogenesis. These findings refer to the crucial role of glycoconjugates in spermatogenesis in the bamboo shark testis.

Angelica sinensis polysaccharide (ASP) is one of the main active components of Angelica sinensis (AS) that is widely used in traditional Chinese medicine. ASP is water-soluble polysaccharides, and it is mainly composed of glucose (Glc), galactose (Gal), arabinose (Ara), rhamnose (Rha), fucose (Fuc), xylose (Xyl) and galacturonic acid (GalUA). The extraction methods of ASP include hot water extraction and ultrasonic wave extraction, and different extraction methods can affect the yield of ASP. ASP has a variety of pharmacological activities, including hematopoietic activity, promoting immunity, antitumor, anti-inflammatory, antioxidant, anti-aging, anti-virus, liver protection, and so on. As a kind of natural polysaccharide, ASP has potential application as drug carriers. This review provides a comprehensive summary of the latest extraction and purification methods of ASP, the strategies used for monosaccharide compositional analysis plus polysaccharide structural characterization, pharmacological activities and drug carrier applications, and it can provide a basis for further study on ASP.

Consumption of marine alga-based polysaccharides as additional functional foods can endow with health benefits by diminishing the risk of chronic diseases. A polygalacto-fucopyranose characterized as [→1)-2, 4-SO3-α-Fucp-(3 → 1)-{2-SO3-α-Fucp-(3→}] with [(4 → 1)-6-OAc-β-Galp-(4→] side chain isolated from marine alga Sargassum wightii exhibited potential antihypertensive activity. Upon treatment with studied polygalactofucan (50 mg/kg BW), serum hypertension biomarkers troponin-T (1.3 pg/mL), troponin-I (1.2 μg/dL) and angiotensin-II converting enzyme (0.18 pg/mL) were significantly recovered in hypertensive rats compared to disease control. Serum cardiovascular risk indices of diseased rats were significantly decreased ( 17%), and were comparable with standard antihypertensive agent telmisartan (8.3-10.2% at 2 mg/kg BW). The studied compound was safe for consumption as obvious from the high LD50 value (>5 g/kg), and could be developed as a prospective functional food ingredient attenuating the pathophysiological attributes causing hypertension-related conditions.

The aim of this study was to establish the fine structure of fucoidan from Sargassum oligocystum and to study the radiosensitizing effect of fucoidans from three algae of genus Sargassum (S. oligocystum, S. duplicatum, and S. feldmannii) with different structures. The fucoidan SoF2 from S. oligocystum was sulfated (32%) galactofucan (Fuc:Gal = 2:1), with a Mw of 183 kDa (Mw/Mn = 2.0). Its supposed structure was found to be predominantly 1,3-linked fucose as the main chain, with branching points at C2 and C4. The branches could be single galactose and/or fucose short chains with terminal galactose residues. Sulfate groups were found at positions C3, C2, and/or C4 of fucose residues and at C2 and/or C4 of galactose residues. The radiosensitizing effect of galactofucans from S. oligocystum, S. duplicatum, and S. feldmannii against human melanoma SK-MEL-28, colon HT-29, and breast MDA-MB-231 cancer cells was investigated. The influence of all investigated polysaccharides treatments with/without X-ray radiation on colony formation of human melanoma cells SK-MEL-28 was weak. Fucoidan from S. feldmannii has been shown to be the most promising radiosensitizing compound against human colon HT-29 and breast MDA-MB-231 cancer cells.

Cellulosimicrobium strain SE3T was isolated from the San Elijo coastal lagoon near San Diego. A whole genome-based phylogenetic comparison shows great heterogeneity within the Cellulosimicrobium genus. Based on average nucleotide identity, whole genome-based comparison, and the presence of a unique l-fucose metabolic pathway, strain SE3T was shown to belong to a novel species within the genus, together with five other strains. The name Cellulosimicrobium fucosivorans sp. nov. is proposed, with strain SE3T as the type strain. The strain encodes a unique alpha-l-fucosidase and the l-fucose metabolic pathway is homologous to the one recently described in Campylobacter jejuni. C. fucosivorans is able to grow on l-fucose, and interestingly, the biosynthesis of the yellow carotenoid is dependent on the presence of l-fucose in the media. The ability to metabolize fucose and the linked production of carotenoids are expected to provide C. fucosivorans with a competitive advantage in the sunny coastal lagoon area.

O-linked modification of nuclear and cytosolic proteins with monosaccharides is essential in all eukaryotes. While many aspects of this post-translational modification are highly conserved, there are striking differences between plants and the animal kingdom. In animals, dynamic cycling of O-GlcNAc is established by two essential single copy enzymes, the O-GlcNAc transferase OGT and O-GlcNAc hydrolase OGA. In contrast, plants balance O-GlcNAc with O-fucose modifications, catalyzed by the OGT SECRET AGENT (SEC) and the protein O-fucosyltransferase (POFUT) SPINDLY (SPY). However, specific glycoside hydrolases for either of the two modifications have not yet been identified. Nucleocytoplasmic O-glycosylation is still not very well understood in plants, even though a high number of proteins were found to be affected. One important open question is how specificity is established in a system where only two enzymes modify hundreds of proteins. Here, we discuss the possibility that O-GlcNAc- and O-fucose-binding proteins could introduce an additional flexible layer of regulation in O-glycosylation-mediated signaling pathways, with the potential of integrating internal or external signals.

The anti-glycation effects of polysaccharides from Ecklonia cava were examined according to extraction method-hot buffer (HP), ultrasound (UP), enzyme (EP), or a combination of ultrasound and enzyme (UEP). The physicochemical properties, monosaccharide compositions, and structural characteristics of the polysaccharides were determined. UP, EP, and UEP had higher fucose and galactose compositions than HP. The FT-IR spectra of samples showed the presence of sulfate esters and 4-sulfate galactose. 1H NMR indicated that alginate was removed by purification. UP, EP, and UEP possessed higher sulfate contents than HP. UEP presented with the highest extraction yield and lowest protein and uronic acid contents. The levels of AGE formation, as well as fructosamine, α-dicarbonyl, and protein carbonyl contents were determined during a 3-week incubation in a BSA/fructose system. UEP and UP effectively inhibited AGE, although the inhibition effect was lower than that of aminoguanidine. However, UP and UEP showed higher inhibition of fructosamine, α-dicarbonyl, and protein carbonyl than aminoguanidine. AGE formation was negatively correlated with sulfate content and some monosaccharide compositions (fucose, galactose, and glucose), but positively correlated with molecular weight. Overall, the present study suggests that UEP is a suitable extraction method for obtaining anti-glycation agents from E. cava.

The α1,6-fucosyltransferase, FUT8, is the sole enzyme catalyzing the core-fucosylation of N-glycoproteins in mammalian systems. Previous studies using free N-glycans as acceptor substrates indicated that a terminal β1,2-GlcNAc moiety on the Man-α1,3-Man arm of N-glycan substrates is required for efficient FUT8-catalyzed core-fucosylation. In contrast, we recently demonstrated that, in a proper protein context, FUT8 could also fucosylate Man5GlcNAc2 without a GlcNAc at the non-reducing end. We describe here a further study of the substrate specificity of FUT8 using a range of N-glycans containing different aglycones. We found that FUT8 could fucosylate most of high-mannose and complex-type N-glycans, including highly branched N-glycans from chicken ovalbumin, when the aglycone moiety is modified with a 9-fluorenylmethyloxycarbonyl (Fmoc) moiety or in a suitable peptide/protein context, even if they lack the terminal GlcNAc moiety on the Man-α1,3-Man arm. FUT8 could also fucosylate paucimannose structures when they are on glycoprotein substrates. Such core-fucosylated paucimannosylation is a prominent feature of lysosomal proteins of human neutrophils and several types of cancers. We also found that sialylation of N-glycans significantly reduced their activity as a substrate of FUT8. Kinetic analysis demonstrated that Fmoc aglycone modification could either improve the turnover rate or decrease the KM value depending on the nature of the substrates, thus significantly enhancing the overall efficiency of FUT8 catalyzed fucosylation. Our results indicate that an appropriate aglycone context of N-glycans could significantly broaden the acceptor substrate specificity of FUT8 beyond what has previously been thought.

The absence of fucose on asparagine-297 of the human immunoglobulin G (IgG) heavy chain has been shown to enhance antibody-dependent cellular cytotoxicity (ADCC) activity by 10- to 100-fold compared to fucosylated antibody. Our lab is studying the use of adeno-associated virus (AAV) as a vector for the delivery of HIV-specific antibodies for therapeutic purposes. Since the antibody is produced by vector-transduced cells in vivo, current techniques of glycoengineering cannot be utilized. In order to achieve similar enhancement of ADCC with AAV-delivered antibodies, short hairpin RNAs (shRNAs) that target fucosyltransferase-8 (FUT8), were designed, tested, and cloned into AAV vectors used to deliver HIV-specific broadly neutralizing antibodies (bNAbs). Antibodies produced by our glycoengineered-AAV (GE-AAV) vectors were analyzed for fucose content and ADCC. GE-AAV constructs were able to achieve over 80% knockdown of FUT8. Results were confirmed by lectin western blot for α1-6 fucose, which revealed almost a complete absence of fucose on GE-AAV-produced antibodies. GE-AAV-produced antibodies revealed >10-fold enhancement of ADCC, while showing identical neutralization and gp140 trimer binding compared to their fucosylated counterparts. ADCC was enhanced 40- to 60-fold when combined with key Fc mutations known to enhance binding to FcγRIIIA. Our findings define a powerful approach for supercharging AAV-delivered anti-HIV antibodies., Graphical Abstract, Termini et al. describe the creation and validation of a novel glycoengineered-AAV vector for the delivery of anti-HIV antibodies. These AAV are capable of knocking down fucosyltransferase-8 following transduction and, when combined with antibody-Fc mutations, produce broadly neutralizing antibodies with 40- to 60-fold higher ADCC.

The preparation, chemical properties and bio-activities of polysaccharides derived from halophytes have gained an increasing interest in the past few years. Phytochemical and pharmacological reports have shown that carbohydrates are important biologically active compounds of halophytes with numerous biological potentials. It is believed that the mechanisms involved in these bio-activities are due to the modulation of immune system. The main objective of this summary is to appraise available literature of a comparative study on the extraction, structural characterizations and biological potentials, particularly immunomodulatory effects, of carbohydrates isolated from halophytes (10 families). This review also attempts to discuss on bioactivities of polysaccharides related with their structure-activity relationship. Data indicated that the highest polysaccharides yield of around 35% was obtained under microwave irradiation. Structurally, results revealed that the most of extracted carbohydrates are pectic polysaccharides which mainly composed of arabinose (from 0.9 to 72%), accompanied by other monosaccharides (galactose, glucose, rhamnose, mannose and xylose), significant amounts of uronic acids (from 18.9 to 90.1%) and some proportions of fucose (from 0.2 to 8.3%). The molecular mass of these pectic polysaccharides was varied from 10 to 2650 kDa. Hence, the evaluation of these polysaccharides offers a great opportunity to discover novel therapeutic agents that presented especially beneficial immunomodulatory properties. Moreover, reports indicated that uronic acids, molecular weights, as well as the presence of sulfate and unmethylated acidic groups may play a significant role in biological activities of carbohydrates from halophyte species.

The deoxyhexose sugar L-fucose is important for many biological processes within the human body and the associated microbiota. This carbohydrate is abundant in host gut mucosal surfaces, numerous microbial cell surface structures, and some dietary carbohydrates. Fucosylated oligosaccharides facilitate the establishment of a healthy microbiota and provide protection from infection. However, there are instances where pathogens can also exploit these fucosylated structures to cause infection. Furthermore, deficiencies in host fucosylation are associated with specific disease outcomes. This review focuses on our current understanding of the impact of fucosylation within the mucosal environment of the gastrointestinal tract with a specific emphasis on the mediatory effects in host-microbe interactions.

The L-fucose-specific lectin from Aspergillus niger (ANL), isolated from the corneal smears of a keratitis patient was reported earlier. Here, we examined the interaction of ANL with human hepatocellular and colon cancer cells, evaluated its anti-cancer activity and diagnostic potential to detect aberrantly glycosylated tumour-associated serum glycoproteins such as alpha-fetoprotein (AFP). We observed that ANL strongly bound to both HepG2 and HT-29 cell-lines and this interaction was effectively blocked with L-fucose and mucin in a dose and time-dependent manner with an IC50 of 1.25 and 5 μg/mL for HepG2 and HT-29 cells respectively at 48 hours. ANL treatment increased hypodiploidy and decreased the number of HepG2 cell in G0 -G1 phase at both 24 and 48 hours. Furthermore, ANL increased the level of apoptosis in both HepG2 and HT-29 cells in a time-dependent manner via enhanced production of reactive oxygen species and altered mitochondrial membrane potential, indicative of intrinsic apoptotis pathway activation. Immunoblot analysis confirmed the time-dependent elevation of levels of cytochrome c, initiator caspase-9 and activation of caspase-3. ANL immunohistochemistry on colon cancer tissue and quantification of AFP in HCC patient serum samples by developing an ANL-anti-AFP antibody sandwich enzyme-linked immunosorbent assay confirmed the diagnostic potential of ANL. Here, interaction of ANL with AFP could be effectively blocked in the presence of competing fucose-bearing glycans. We found ANL to be more sensitive than Lens culinaris lectin, a well-known fucose-specific lectin and currently used diagnostic agent. ANL can be further explored as a diagnostic and anti-cancer agent.

Fucose (Fuc), a monosaccharide present at the core or the termini of glycans, critically regulates various biological phenomena and is associated with various diseases. Specifically detecting Fuc residues or inhibiting the fucosylation pathway is pivotal in understanding the mechanisms of how fucosylated glycans are related to biological processes and diseases and in developing novel therapeutic agents.This review focuses on chemical biology approaches using Fuc analogs developed for metabolically labeling fucosylated glycans or inhibiting the biosynthesis of fucosylated glycans.Developed Fuc analogs have different potency, specificity and effects on protein and cellular functions. Developing highly enzyme-specific probes and inhibitors is desirable for future investigations.Chemical glycobiology approaches using sugar analogs are useful for revealing novel mechanisms of inter-relationships among sugar metabolism pathways and manipulating glycan expression to develop new glycan-targeted therapies.

Mucins are large gel-forming polymers inside the mucus barrier that inhibit the yeast-to-hyphal transition of Candida albicans, a key virulence trait of this important human fungal pathogen. However, the molecular motifs in mucins that inhibit filamentation remain unclear despite their potential for therapeutic interventions. Here, we determined that mucins display an abundance of virulence-attenuating molecules in the form of mucin O-glycans. We isolated and cataloged >100 mucin O-glycans from three major mucosal surfaces and established that they suppress filamentation and related phenotypes relevant to infection, including surface adhesion, biofilm formation and cross-kingdom competition between C. albicans and the bacterium Pseudomonas aeruginosa. Using synthetic O-glycans, we identified three structures (core 1, core 1 + fucose and core 2 + galactose) that are sufficient to inhibit filamentation with potency comparable to the complex O-glycan pool. Overall, this work identifies mucin O-glycans as host molecules with untapped therapeutic potential to manage fungal pathogens.

Deletion of α-1,3/4-fucosidase activity in Arabidopsis thaliana resulted in the accumulation of GN1-type free N-glycans with the Lewis a epitope (GN1-FNG). This suggests that the release of α-fucose residue(s) may trigger rapid degradation of the plant complex-type (PCT) GN1-FNG. The fact that PCT-GN1-FNG has rarely been detected to date is probably due to its easier degradation compared with PCT-GN2-FNG.

Galactosamine (GalN) is a well-known agent for inducing viral hepatitis models in rodents, but it can cause toxicity on different organs. Vitamin U (Vit U) has been proved as a powerful antioxidant on many toxicity models. The present study was designed to investigate the protective effects of Vit U on GalN-induced stomach injury. Rats were divided into four groups as follows: control (group I), Vit U given animals (50 mg/kg per day; group II), GalN administered animals (500 mg/kg at a single dose; group III), GalN + Vit U given animals (at the same dose and time, group IV). At the end of the 3rd day, animals were killed, and stomach tissues were taken. They were homogenized and centrifuged. In comparison to the control group, glutathione, total antioxidant capacity levels, catalase, superoxide dismutase, glutathione peroxidase, glutathione reductase, glutathione-S-transferase, and Na

Sulfated galactofucans, as the active compositions of fucoidan, were reported to exhibit antitumor activity. In the current study, a sulfated galactofucan (SGF) from Sargassum thunbergii and its three derivatives (SGF-H, SGF-L, and SGF-S) were prepared for structural analysis. Structural analysis showed that SGF-H was a high molecular weight sulfated galactofucan (51.5/17.8 kDa) with a high molar ratio of galactose (Gal) to fucose (Fuc) (0.66 : 1), SGF-L was a low molecular weight sulfated galactofucan (17.7 kDa) with a low molar ratio of Gal to Fuc (0.20 : 1), and SGF-S was a mixture (1.7 kDa) of sulfated galacto-fuco-oligomers or fuco-oligomers. It was noteworthy that the linkage of Gal residues in SGF-H was a β-linkage while SGF-L was an α-linkage. A comparative study on the anti-lung cancer activity in vitro and in vivo, antimetastatic effects, the metastasis-associated protein expression, and binding abilities to fibroblast growth factors (FGFs) of SGF, SGF-H, and SGF-L was performed to understand the structure–activity relationship. To some extent, SGF-L showed the strongest activity in the inhibition of human lung cancer cells A549 cell proliferation, while SGF-H exhibited the strongest activity in the inhibition of human bronchial epithelial cells BEAS-2B cell proliferation. SGF-L showed the strongest antimetastatic activity, followed by SGF-H and SGF. The expression of metastasis-associated proteins showed only a small difference. The in vivo tumor inhibition of SGF, SGF-H, and SGF-L was 45%, 41%, and 31%, respectively. SPR analysis showed SGF-H binds preferentially to FGF1 and FGF2, while SGF-L preferentially binds to FGF7 and FGF10, suggesting that the anti-lung cancer activity from sulfated galactofucan could involve the FGF-FAK/mTOR pathway.

The effects of galactofucan from Laminaria japonica on the digestion and intestinal microbiota of human were investigated in the present study. Crude fraction of the sulfated polysaccharide from L. japonica (CF) and its molecular-weight homogeneous fraction (CGF-3) were prepared and characterized. In the simulated digestion model for the human saliva and gastrointestinal tract, no obvious changes in the molecular weight or the reducing sugar content of CGF-3 were observed, indicating CGF-3 is resistant to the human digestive system. Then CGF-3 did not affect the α-amylase activity while it dose-dependently inhibited the activity of pancreatic lipase partly depending on its sulfate groups. In the in vitro fermentation with the human fecal microbiota, CF did not change the total carbohydrate, reducing sugar and short chain fatty acids contents, which indicated CF was not utilized by the microbiota. However, the microbiota composition was modulated greatly by CF intervention. These findings shed a light on the better understanding of the impacts of dietary galactofucan on the digestion and intestinal microbiota.

Pancreatic ductal adenocarcinoma (PDAC) is an intractable malignancy for which novel therapeutic targets are in high demand. To uncover glycans expressed within PDAC, we previously performed glycome profiling of PDAC cell lines using lectin microarray and found that the lectin rBC2LCN with specificity to a Fucα1-2Galβ1-3 motif exhibited strong binding to a PDAC cell line (Capan-1) and to all tumor tissues derived from 69 pancreatic cancer patients. Nevertheless, no information was available as to whether glycans containing the Fucα1-2Galβ1-3 motif are expressed within PDAC. Here we used HPLC combined with MALDI-TOFMS to perform a structural and quantitative glycome analysis targeting both N- and O-glycans derived from two types of patient-derived PDAC xenograft mouse models, PC3 (well-differentiated) and PC42 (poorly-differentiated). A higher percentage of highly branched and sialylated complex-type N-glycans was detected in PC42 relative to PC3. The percentage of core 1 O-glycans was higher in PC42 relative to PC3, whereas that of core 3 O-glycans was higher in PC3. Cancer-related glycan epitopes such as Lewis A and Lewis Y were detected in core 3 O-glycans of both PC3 and PC42. H-type3 containing the Fucα1-2Galβ1-3 motif was detected in Core 2 O-glycans in both models, explaining the molecular mechanism of the binding of rBC2LCN to PDAC.

Summary Carbohydrate-binding proteins from pathogenic bacteria and fungi have been shown to be implicated in various pathological processes, where they interact with glycans present on the surface of the host cells. These interactions are part of the initial processes of infection of the host and are very important to study at the atomic level. Here, we report the room temperature neutron structures of PLL lectin from Photorhabdus laumondii in its apo form and in complex with deuterated L-fucose, which is, to our knowledge, the first neutron structure of a carbohydrate-binding protein in complex with a fully deuterated carbohydrate ligand. A detailed structural analysis of the lectin-carbohydrate interactions provides information on the hydrogen bond network, the role of water molecules, and the extent of the CH-π stacking interactions between fucose and the aromatic amino acids in the binding site.

α-Linked galactose is a common carbohydrate motif in nature that is processed by a variety of glycoside hydrolases from different families. Terminal Galα1–3Gal motifs are found as a defining feature of different blood group and tissue antigens, as well as the building block of the marine algal galactan λ-carrageenan. The blood group B antigen and linear α-Gal epitope can be processed by glycoside hydrolases in family GH110, whereas the presence of genes encoding GH110 enzymes in polysaccharide utilization loci from marine bacteria suggests a role in processing λ-carrageenan. However, the structure–function relationships underpinning the α-1,3-galactosidase activity within family GH110 remain unknown. Here we focus on a GH110 enzyme (PdGH110B) from the carrageenolytic marine bacterium Pseudoalteromonas distincta U2A. We showed that the enzyme was active on Galα1–3Gal but not the blood group B antigen. X-ray crystal structures in complex with galactose and unhydrolyzed Galα1–3Gal revealed the parallel β-helix fold of the enzyme and the structural basis of its inverting catalytic mechanism. Moreover, an examination of the active site reveals likely adaptations that allow accommodation of fucose in blood group B active GH110 enzymes or, in the case of PdGH110, accommodation of the sulfate groups found on λ-carrageenan. Overall, this work provides insight into the first member of a predominantly marine clade of GH110 enzymes while also illuminating the structural basis of α-1,3-galactoside processing by the family as a whole.

Genomic analysis of the marine bacterium Wenyingzhuangia fucanilytica CZ1127T revealed the presence of four fucoidanase genes fwf1, fwf2, fwf3, fwf4 that belonged to the glycoside hydrolase family 107 (GH107, CAZy), which is located in one gene cluster putatively involved in fucoidan catabolism. Genes encoding two fucoidanases fwf1 and fwf2 were cloned, and the proteins FWf1 and FWf2 were produced in Escherichia coli cells. The recombinant fucoidanases were purified and the biochemical properties of these enzymes were studied. The amino acid sequences of FWf1 and FWf2 showed 41 and 51% identity respectively with a fucoidanase FcnA from the marine bacterium Mariniflexile fucanivorans, with the established 3D structure. Structures of the oligosaccharides produced during enzymatic hydrolysis of fucoidan by FWf1 and FWf2 have been determined by NMR spectroscopy. Detailed substrate specificities of FWf1 and FWf2 were studied using fucoidans and sulfated fucooligosaccharides with different structures. Both fucoidanases catalyzed hydrolysis of 1→4-glycosidic bonds between sulfated α-l-fucose residues but had different specificities regarding sulfation patterns of the fucose residues in fucoidan molecules. Specific cleavage sites recognizable by the fucoidanases in fucoidan molecules were determined. The obtained results provide new knowledge about differences between specificities of the fucoidanases belonging to the GH107 family.

An L-fucose lectin, ANL from the corneal smears of a mycotic keratitis patient was reported earlier. Interaction of ANL with immortalized Human Corneal Epithelial Cells (HCECs) was studied in order to assign the role of ANL in pathogenesis. ANL showed strong binding to HCECs which could be blocked by L-fucose and mucin. At concentrations below 0.6 μg/mL ANL showed proliferative effect and highest at 0.07 μg/mL leading to expression of proinflammatory cytokines IL-6 and IL-8. ANL induced proinflammatory response is mediated by TLR-2,-4, MyD88, NFkB and C-Jun dependent signaling. In contrast, ANL at concentrations above 0.6 μg/mL showed growth inhibitory effect at 48 h with an IC50 of 2.75 μg/mL. Western blot analysis revealed that HCECs treated with ANL at lower concentration induced the expression of proinflammatory signaling proteins TLR-2, 4, MyD88, NFkB and C-Jun which maintain high cell proliferating state. At higher concentration ANL induced apoptotic effect in HCECs with an increase in early apoptotic population as demonstrated by Annexin V-PI assay. ANL induced the expression of apoptotic proteins FADD, Caspase 8 and -3 mediated by MyD88. These findings demonstrate implication of ANL in pathogenesis and the findings are of clinical significance in developing strategy for controlling the infection leading to mycotic keratitis.

Damage from ultraviolet (UV)B exposure is cumulative and proceeds via augmenting cellular oxidative stress. The present study investigates UVB-protective effects of Sargassum confusum derived fucoidans in human keratinocytes. Algae was extracted using Celluclast, and fucoidan fractions were recovered by step gradient ethanol precipitation. Refined fucoidan fractions were treated to human HaCaT keratinocytes and exposed to UVB (50 mJ cm−2). Among fucoidan fractions, SCFC4, the lowest molecular weight (MW) fraction indicated better UVB-protective effects with dose-dependent reduction of intracellular ROS levels, while recovering the cell viability. SCFC4 suppressed UVB-induced apoptotic body formation, sub-G1 cell content, and DNA damage by hindering the mitochondria-mediated apoptotic pathway. Also, SCFC4 repressed upstream mediators of UVB-induced inflammatory responses, which would impair stratum corneum hydration. The above therapeutic effects of SCFC4 could be attributed to suppression of nuclear factor (NF)-κB and mitogen-activated protein kinase (MAPK) pathways while activating nuclear factor erythroid 2-related factor 2 (Nrf2) mediated production of antioxidant enzymes. SCFC4 was identified as a fucoidan (MW ≈ 20 kDa) with a 64.64% fucose and 23.62% sulfate contents. Further analysis of shelf life, safety, and efficacy could promote SCFC4's use as a cosmetic ingredient.

The aim of the current study was to compare crude polysaccharides extracted from Schinus terebinthifolius Raddi (PSTF) and S. molle L. (PSMF) fruits based on their structures, physicochemical characteristics, monosaccharide composition, as well as in vitro and in vivo assays. The extraction yield of PSTF (4.26%) was higher than that of PSMF (3.56%). Remarkable variability was detected in the content of carbohydrates (80.64 ± 0.98%), protein (1.80 ± 0.28%), fat (0.04 ± 0.005%) and ash (6.32 ± 0.26%). FT-IR assay and 1H and 13C NMR spectroscopy revealed that fruits extract showed similar structural characteristics. Thin layer chromatography together with HPLC-RID analysis showed that the monosaccharide composition varied significantly between species. Both contained arabinose (40.55–42.03%) galacturonic acid (31.21–41.15%), and fucose (10.90–17.63%), but PSTF had glucose (9.13%) whereas PSMF had galactose (7.40%). Functional analyses demonstrated that samples exhibited favorable water- and oil-retention capacity, emulsifying properties, and foaming qualities. PSTF exhibited the highest antioxidant effects. Both of them showed a remarkable in vitro antidiabetic effect. PSMF highly mitigated H2O2-induced hemolysis and exhibited ~80% antihemolytic activity. The extracted polysaccharides showed potent inhibitory activity against AAPH-induced plasmid DNA damage. PSTF and PSMF revealed interesting in vivo antinociceptive and anti-inflammatory capacities.

Fungal lectin can bind specific carbohydrate structures of the host and work in recognition and adhesion or as a toxic factor. AofleA, as a fucose-specific lectin from widely studied nematode predatory fungus Arthrobotrys oligospora, possibly plays a key role in the event of capturing nematodes, but the mechanism remains unknown. Here we report the crystal structure of AofleA, which exists as a homodimer with each subunit folds as a six-bladed β-propeller. Our structural and biological results revealed that three of the six putative binding sites of AofleA had fucose-binding abilities. In addition, we found that AofleA could bind to the pharynx and intestine of the nematode in a fucose-binding-dependent manner. Our results facilitate the understanding of the mechanism that fucose-specific lectin mediates fungi-nematodes interaction, and provide structural information for the development of potential applications of AofleA.

A polysaccharide (ESPS) purified from Eupolyphaga sinensis Walker by ion exchange chromatography and gel chromatography was investigated, including its structure characterization and antitumor activity. The results showed that ESPS was composed of rhamnose, fucose, arabinose, xylose, glucose, and galactose in a molar ratio of 7.4: 3.1: 13.9: 9.3: 39.7: 26.5, with the mean weight (Mw) of 2.14 × 104 Da; the main chain of ESPS was mainly composed of → 4) - α - D - Glcp - (1 → and → 3) - β - D - Galp - (1 →, and the side chains were connected to the main chain through the O-6 atom of glucose and O-4 and O-6 atom of galactose. In addition, ESPS promoted the lymphocyte proliferation and inhibited liver cancer cells growth through enhancing lymphocyte activity in vitro, mainly NK cells. Moreover, ESPS markedly stimulated immunity in H22-bearing mice by increasing the spleen and thymus indices and effectively inhibited H22 cell growth in vivo. These data indicated that ESPS was a polysaccharide component possessing high anti-hepatocellular carcinoma activity, representing a potential immunotherapy candidate for the treatment of liver cancer.

Sanghuangporus is a traditional medicine that has been used for nearly 2000 years in Asia. It has been found to enhance immunity and have anticancer properties. Some studies determined that Sanghuangporus polysaccharide can inhibit tumors in vitro, although the underlying mechanisms remain unknown. Therefore, the present study investigated the antitumor effects of polysaccharides from the fruiting body of S. vaninii and their associated mechanisms. Water-soluble S. vaninii polysaccharide (SVP) with a molecular weight of 3.156 × 104 Da was isolated and found to be mainly composed of mannose, rhamnose, glucuronic acid, galacturonic acid, glucosamine, glucose, galactosamine, galactose, xylose, arabinose, and fucose in molar ratios of 1.63:0.04:0.36:0.03:0.13:8.39:0.08:1.08:0.25:1.07:0.40. SVP showed modest anti-proliferative activity against HeLa, SHG-44, SMMC-7721, and MCF-7 cells. Furthermore, it effectively regulated cell cycle, promoted apoptosis, and reduced the migratory and invasive capacities of MCF-7 cells. Mechanistically, SVP enhanced activation of p53-related genes and down-regulated MMP expression. These findings describe a potential natural polysaccharide for tumor therapy and a basis for the development of fungal polysaccharides as a type of health food.

The escalation of fine particulate matter (PM) air pollution has recently become a global concern. Evidence is fast accumulating on PM exposure-related skin damage. The present study explored the therapeutic potentials of fucoidan purified from Sargassum natans against damaging effects of PM exposure on human HaCaT keratinocytes. Fucoidan (SNF7) was purified from S. natans by an enzyme-assisted extraction and purified by anion exchange chromatography. SNF7 (≈50 kDa) was identified as a fucoidan containing 70.97% fucose and 36.41 ± 0.59% of sulfate. Treatment of fine dust from Beijing, China (CFD) increased intracellular ROS levels in HaCaT cells triggering DNA damage and apoptosis. Treatment of SNF7 dose-dependently attenuated CFD-induced surge of intracellular ROS levels in keratinocytes by increasing antioxidant defense enzymes. Moreover, SNF7 chelated metal ions Pb2+, Ba2+, Sr2+, Cu2+, Fe2+, and Ca2+ coming from CFD. The results substantiated the potential therapeutic effects of SNF7 against CFD-induced oxidative stress. Further studies could promote SNF7's use as an active ingredient in cosmetics.

A fucoidan (LJSF4) purified from Saccharina japonica was found to show a strong anti-inflammatory effect via activity assay in lipopolysaccharide (LPS) induced RAW 264.7 macrophage cells and zebrafish. Chemical and structural analysis indicated that LJSF4 with a sulfate content of 30.72% was composed of fucose, galactose, rhamnose, xylose and mannose with molar ratio percentages of 79.49%, 16.76%, 0.82%, 1.08% and 1.84%. NMR spectroscopy showed that LJSF4 is a polysaccharide with a backbone of alternating 1 → 3 linked α- l -fucopyranosyl and →4-α- l -fucopyranosyl with sulfate groups mainly at C–4 and partially at C–2 positions. Moreover, it also contained branches in the form of β- d -Galp-(1 → 4) units. The results of anti-inflammatory effect in vitro demonstrated that LJSF4 decreased the production of nitric oxide (NO) and cytokines, including TNF-α, IL-1β and IL-6. The mechanism revealed to be associated with the down-regulated expression of signal pathways including MAPK and NF-κB. By in vivo assay, LJSF4 showed a significantly protective effect by reducing the cell death rate, and the production of NO and ROS on LPS exposed zebrafish. Our results indicated that LJSF4 has the potential to be developed as an anti-inflammatory agent applied in functional food and cosmetic industries.

Background: Fucose is a deoxyhexose sugar. While the biological roles of L-fucose remain unclear, the sugar is known to accelerate the malignant potential of cancer cells. Therefore, this study aimed to evaluate the viability pattern of human cancer and normal cell lines treated with fucose. Methods: The human gingival fibroblast (HGF-1), colorectal adenocarcinoma (HT-29) and skin malignant melanoma (A375) cell lines were cultured and treated with fucose at three concentrations of 1, 5, and 10 mg/ml. Cell viability was then measured using (3-(4, 5-dimethylthiazolyl-2)-2, 5- diphenyltetrazolium bromide (MTT) assay. The data were analyzed using Statistical Package for the Social Sciences software. Results: The percentage of HGF-1 cell viability showed a rapid decline after day 1 of treatment. HT-29 and A375 were capable of surviving treatment with high fucose concentrations. The data were highly significant at p < 0.001. Conclusion: Whereas a high concentration of fucose is toxic to the HGF-1 cell line, the HT-29 and A375 cell lines could potentially adapt to this condition. Down- or upregulation of certain molecules that could induce or inhibit cell death may explain such adaptation. Further testing of up- and downregulated molecules should be conducted in future work.

Three polysaccharides (SH-1, SH-2 and SH-3) were purified from a brown macroalgea, Sargassum hemiphyllum. The autohydrolysis products from each polysaccharide were separated to three fractions (S fractions as oligomers, L fractions as low molecular weight polysaccharides and H fractions as high molecular weight polysaccharides). Mass spectroscopy of S fractions (SH-1-S, SH-2-S and SH-3-S) showed that these three polymers all contained short stretches of sulfated fucose. The structures of L fractions (SH-1-L, SH-2-L and SH-3-L) were determined by nuclear magnetic resonance (NMR). SH-1-L was composed of two units, unit A (sulfated galactofucan) and unit B (sulfated xylo-glucuronomannan). Unit A contained a backbone of (1, 6-linked β-D-Gal) n1, (1, 3-linked 4-sulfated α-L-Fuc) n2, (1, 3-linked 2, 4-di-sulfated α-L-Fuc) n3, (1, 4-linked α-L-Fuc) n4 and (1, 3-linked β-D-Gal) n5, accompanied by some branches, such as sulfated fuco-oligomers, sulfated galacto-oligomers or sulfated galacto-fuco-oligomers. And unit B consisted of alternating 1, 4-linked β-D-glucuronic acid (GlcA) and 1, 2-linked α-D-mannose (Man) with the Man residues randomly sulfated at C6 or branched with xylose (Xyl) at C3. Both SH-2-L and SH-3-L were composed of unit A and their difference was attributed to the ratio of n1: n2: n3: n4: n5. Based on monosaccharide analysis, we hypothesize that both SH-1-H and SH-2-H contained unit A and unit B while SH-3-H had a structure similar to SH-3-L. An assessment of anti-complement activities showed that the sulfated galactofucan had higher activities than sulfated galacto-fuco-xylo-glucuronomannan. These results suggest that the sulfated galactofucans might be a good candidate for anti-complement drugs.

To clarify the property of a novel guava seed polysaccharide (GSPS), GSPS was subjected to purify using Sepharose 6B gel filtration chromatography and further characterize the property of each individual isolated fraction. GSPS further resolved into three purified fractions, guava seed polysaccharide fraction 1 (GSF1), GSF2 and GSF3. Isolated GSF1, GSF2 and GSF3 were respectively subjected to high performance size exclusion chromatography; molecular weights of three polysaccharide fractions were determined. GSPS, GSF1, GSF2 and GSF3 were suggested to be proteopolysaccharides or glycoproteins. GSPS, GSF1, GSF2 and GSF3, particularly GSF3, were found to have a Th2-inclination property and anti-inflammatory potential. Heated GSF3 did not significantly (P > .05) decreased its immunomodulatory activity, suggesting that GSF3 is a proteopolysaccharide. The deproteinated GSF3 markedly lost its immunomodulatory activity, suggesting that both protein and carbohydrate moiety in GSF3 are essential to its immunomodulatory function. Analyses of monosaccharides composition in GSF3 using a pre-column derivatization high performance liquid chromatography exhibited that GSF3 was composed of glucuronic acid (3.28%), galacturonic acid (28.13%), galactose (14.88%), mannose (3.96%), glucose (22.99%), arabinose (7.31%), ribose (1.55%), xylose (14.81%), fucose (1.68%) and rhamnose (1.43%). Overall, we evidence that GSF3 is a low molecular weight proteopolysaccharide with potent anti-inflammatory and immunomodulatory effects.

Mutarotases catalyze the α-β anomeric conversion of monosaccharide, and play a key role in utilizing sugar as enzymes involved in sugar metabolism have specificity for the α- or β-anomer. In spite of the sequential similarity to l -rhamnose mutarotase protein superfamily (COG3254: RhaM), the ACAV_RS08160 gene in Acidovorax avenae ATCC 19860 (AaFucM) is located in a gene cluster related to non-phosphorylative l -fucose and l -galactose metabolism, and transcriptionally induced by these carbon sources; therefore, the physiological role remains unclear. Here, we report that AaFucM possesses mutarotation activity only toward l -fucose by saturation difference (SD) NMR experiments. Moreover, we determined the crystal structures of AaFucM in the apo form and in the l -fucose-bound form at resolutions of 2.21 and 1.75 A, respectively. The overall structural folding was clearly similar to the RhaM members, differed from the known l -fucose mutarotase (COG4154: FucU), strongly indicating their convergent evolution. The structure-based mutational analyses suggest that Tyr18 is important for catalytic action, and that Gln87 and Trp99 are involved in the l -fucose-specific recognition.

The Notch signaling pathway is highly conserved and essential in animal development and tissue homeostasis. Regulation of Notch signaling is a crucial process for human health. Ligands initiate a signal cascade by binding to Notch receptors expressed on the neighboring cell. Notch receptors interact with ligands through their epidermal growth factor-like repeats (EGF repeats). Most EGF repeats are modified by O-glycosylation with residues, such as O-linked N-acetylglucosamine (O-GlcNAc), O-fucose, and O-glucose. A recent study revealed the distinct roles of these O-glycans in ligand binding, processing, and trafficking of Notch receptors. In particular, O-GlcNAc glycans are essential for Delta-like (DLL) ligand-mediated Notch signaling. In this study, we showed that O-GlcNAc promotes Notch1 trafficking to the cell surfaces under the condition that O-fucose and O-glucose are removed from consecutive EGF repeats of Notch1. Through in vitro experiments, we showed that O-GlcNAc mediates the stability of EGF domains in the same manner as O-fucose and O-glucose. Thus, O-GlcNAc on EGF domains possesses a shared function in the stability of EGF domains and Notch1 trafficking.

A water soluble heteroglycan (THPS) of an average molecular weight ~1.98 × 105 Da was isolated from the aqueous extract of the fruit bodies of an edible mushroom Termitomyces heimii. Structural characterization of THPS was carried out using acid hydrolysis, methylation analysis, periodate oxidation, Smith degradation and 1D/2D NMR studies. Sugar analysis indicated the presence of glucose, mannose, galactose, and fucose in a molar ratio of nearly 6:2:2:1. The repeating unit of the THPS had a backbone consisting of four (1 → 3)-β- d -glucopyranosyl, one (1 → 6)-β- d -glucopyranosyl, two (1 → 3)-α-D-manopyranosyl, and two (1 → 6)-α-D-galactopyranosyl residues, out of which one (1 → 3)-β- d -glucopyranosyl residue was branched at O-6 position with terminal β- d -glucopyranosyl residue and one (1 → 6)-α-D-galactopyranosyl residue was branched at O-2 position with terminal α-L-fucopyranosyl residue.

Lectins are the primary recognition macromolecules for various types of fucosylation, a common eukaryotic post-translational modification. In this study, we report the heterologous expression and molecular binding properties of a fucose-specific lectin, AofleA, isolated from Arthrobotrys oligospora. This is the first reported fucose-specific lectin found in nematophagous fungi. The recombinant AofleA (r-AofleA) was expressed in Escherichia coli with high efficiency, yielding at least 500 mg of soluble and functional r-AofleA per liter of broth. Using hemagglutination inhibition assay and glycan microarray analysis, r-AofleA was found to be broadly specific for fucosylated glycans or oligosaccharides including Fucα(1–2), Fucα(1–3), Fucα(1–4) and Fucα(1–6) linkages, similar to Aleuria aurantia lectin (AAL). Frontal affinity chromatography showed that r-AofleA has high affinity towards PA-L-fucose with an average Kd value of 15 nM. These findings provide a basis for improved understanding of the structure and functions of AofleA during recognition and capture of prey nematodes by nematophagous fungus A. oligospora.

We obtained a new acidic soy hull polysaccharide (SHP-1) with a molecular weight (Mw) of 4.81 × 105 g/mol through ammonium oxalate and microwave assisted extraction. SHP-1 was mainly composed of galacturonic acid, galactose, rhamnose and arabinose (molar ratio = 46.59%:17.95%:14.77%:13.97%) with small amounts of fucose, glucose, mannose and xylose. The chemical structure was presumed to be of pectin-I type, consisting of 2/3 HGA and 1/3 RG-I. Furthermore, the rheological information and the chain morphology of SHP-1 were different in five solvents. Surfactant, salt and alkali solutions enhanced the solubility and flexibility of the polysaccharide, but the polysaccharide showed decreased fluidity under acidic conditions. The addition of ions and alkali increased the consistency coefficient of the solution, but the effect was far less than that of the cross-linking morphology. The structural and morphological information of purified SHP should aid in further study of its structure-function relationships and applications.

Fucosyltransferase 8 (FUT8) and β-galactoside α-2,6-sialyltransferase 1 (ST6GAL1) are glycosyltransferases that catalyze α1,6-fucosylation and α2,6-sialylation, respectively, in the mammalian N-glycosylation pathway. They are aberrantly expressed in various human diseases. FUT8 is non-glycosylated but is responsible for the fucosylation of ST6GAL1. However, the mechanism for the interaction between these two enzymes is unknown. In this study, we show that serum levels of α2,6-sialylated N-glycans are increased in Fut8−/− mice, whereas the mRNA and protein levels of ST6GAL1 are unchanged in mouse live tissues. The level of α2,6-sialylation on IgG was also enhanced in Fut8−/− mice along with ST6GAL1 catalytic activity increase in both serum and liver. Moreover, it was observed that ST6GAL1 prefers non-fucosylated substrates. Interestingly, increased core fucosylation accompanied by a reduction in α2,6-sialylation, was detected in rheumatoid arthritis patient serum. These findings provide new insight into the interactions between FUT8 and ST6GAL1.

High-fat diet (HFD) alters the glycosylation patterns of intestinal mucins leading to several health problems. We studied by histochemical and lectin-binding methods mucin alterations in the duodenum of mice fed a HFD for 25 weeks. Histochemical methods included periodic acid–Schiff, alcian blue pH 2.5, and high-iron diamine. Lectin-binding experiments were performed with SBA, PNA, WGA, MAA-II, SNA, ConA, UEA-I, LTA, and AAA. SBA, PNA, WGA, MAA-II, and SNA were tested also after desulfation and ConA after periodate-sodium borohydrate treatments (paradoxical ConA). Duodenal mucins are secreted by Brunner’s glands and goblet cells in the villi. Brunner’s glands of HFD mice showed increased secreting activity and a general reduction of glycosylated residuals, such as fucose and terminal α1,4-linked GlcNAc. Moreover, a general reduction of glycosylated residuals in the goblet cells of villi such as the fucosylated and sulfated ones was observed. Since the cited residuals are involved in cytoprotective and cytostatic functions, as well as in interactions with the intestinal microbiota and protection against parasites and inflammatory disorders, we conclude that HFD can predispose duodenum to several possible health disorders.