L-fucose Residues Research Articles

Application of fucoidan for the encapsulation of yeast K2 toxin

Fucoidan is an anionic polysaccharide derived from brown algae and is composed of l-fucose residues linked by alternating α-(1→3) with α-(1→4) linkages or by either α-(1→3) or α-(1→4) linkages alone. Fucoidan is known for its pharmacological, antibacterial, and antifungal properties and is used in the food industry. The aim of the study was to apply fucoidan to the formulation of the yeast killer toxin K2 produced by Saccharomyces cerevisiae. K2 toxin-loaded fucoidan particles were formed by a low-cost complexation method. They were characterised by dynamic light scattering, FT-IR spectroscopy, and scanning electron microscopy methods. The particles ranged in size from 180 to 280 nm and were stable for at least one month at 4°C. The encapsulated K2 toxin retained its antimicrobial activity. In addition, complexation with fucoidan increased the stability of the K2 toxin at room temperature. Fucoidan could serve as a carrier for K2 toxin and enhance the biotechnological potential of antimicrobial agents produced by S. cerevisiae. Encapsulation of K2 toxin opens up opportunities for its use in the food and beverage industry for biopreservation.

Glycosidic linkages of fungus polysaccharides influence the anti-inflammatory activity in mice

Over decades, the source-function relationships of bioactive polysaccharides have been progressively investigated, however, it is still unclear how a defined structure may conduce to the bioactivities of polysaccharides. To explore the structure-function relationship of fungus polysaccharides, we employed a dextran sulfate sodium (DSS)-induced colitis mouse model to compare the anti-inflammatory activity of two fungus polysaccharides from Dictyophora indusiata (DIP) and Tremella fuciformis (TFP), which exhibit distinct glycosidic linkages. The structures of DIP and TFP were characterized through molecular weight detection, molecular morphology analysis, methylation analysis, and NMR analysis. Subsequently, we employed a DSS-induced colitis model to assess the anti-inflammatory efficacy of DIP and TFP. The colitis symptoms, histological morphology, intestinal inflammatory cytokines, and the composition and function of gut microbiota before and after polysaccharides treatment in colitis mice were also investigated. DIP, l,3-β-D-glucan with 1,4-β and 1,6-β-D-Glcp as branched chains, exhibited superior therapeutic effect than that of TFP consisted of a linear 1,3-α-D-mannose backbone with D-xylose and L-fucose in the side chains. Both DIP and TFP relieved DSS-induced colitis in a gut microbiota dependent manner. Furthermore, metagenomics showed that DIP and TFP could partially reverse the bacterial function in colitis mice. Glycoside Hydrolase 1 (GH1) and GH3 were shown to cleave the glucose linkages in DIP, while GH92 and GH29 were likely active on the α-1,3-linked mannose linkages and the glycosidic bonds of L-fucose residues in TFP. Our findings highlight the pivotal role of glycosidic linkages in anti-inflammatory activities of fungus polysaccharides and would promote the design and discovery of polysaccharides with designated activity to be used as functional foods and/or therapeutics.

The contribution of fucosyltransferases to cancer biology.

Fucosyltransferases are enzymes that transfer L-fucose residues from a donor substrate to target molecules. These enzymes are encoded by genes known as FUTs (FUT1 to FUT-11), along with POFUT1 and 2. Changes in FUT expression have a significant role in cancer development and malignancy. This review delves into the biochemistry and biological functions of FUTs and their contributions to cancer. Broadly, FUTs play roles in cancer tumorigenesis, survival, and metastasis. Interactions between fucosylated glycans and various molecules associated with cancer, such as E-selectins and the epidermal growth factor receptor (EGFR), offer alternative pathways for cancer development. The review also highlights FUTs as potential biomarkers for cancer prognosis and diagnosis, along with their application as targets for therapy.

Production of high- and low-molecular weight fucoidan fragments with defined sulfation patterns and heightened in vitro anticancer activity against TNBC cells using novel endo-fucanases of the GH107 family

Fucoidans are complex fucose-containing sulfated polysaccharides with pronounced anticancer effects. Their structure-anticancer activity relationships are difficult to determine due to fucoidans' complex, often irregularities-including structures. Fucoidan-active enzymes can be used for this propose. We have investigated two new recombinant endo-fucanases FWf3 and FWf4 from the marine bacterium Wenyingzhuangia fucanilytica CZ1127T that belong to the 107 family of glycoside hydrolases (GH). Both enzymes cleaved α-(1→4)-glycosidic bonds but in fucoidan fragments with different sulfation patterns. FWf3 is the first characterized endo-fucanase that cleaves glycosidic bonds between 2O- and 2,4diO-sulfated L-fucose residues. The obtained endo-fucanases were used to produce low- and high-molecular weight fucoidan derivatives with different sulfate group locations. Low- and high-molecular weight fucoidan derivatives rich with 2,4diO-sulfation were shown to inhibit MDA-MB-231 cell colony formation more efficiently than the native fucoidan and the derivatives sulfated otherwise. Such derivatives effectively suppressed the mitochondrial membrane potential of MDA-MB-231 cells and reduced the expression of the glucose transporter 1 (GLUT1). Co-treatment of MDA-MB-231 cells with the fucoidan derivatives and oligomycin (an OXPHOS inhibitor) resulted in a synergistic anticancer effect. The data obtained demonstrate, that fucoidan and its 2,4diO-sulfated derivatives can be an effective adjunct in TNBC therapy targeting cell metabolism.

Dual role of fucosidase in cancers and its clinical potential.

Glycosidases and glycosyltransferases greatly impact malignant phenotype of tumors though genetics and epigenetics mechanisms. As the member of glycoside hydrolase (GH) families 29A, α-L-fucosidases (AFUs) are involved in the hydrolysis of terminal L-fucose residues linked via α-1,2, α-1,3, α-1,4 or α-1,6 to the reducing end of N-acetyl glucosamine (GlcNAc) of oligosaccharide chains. The defucosylation process mediated by AFUs contributes to the development of various diseases, such as chronic inflammatory diseases, immune disorders, and autoimmune diseases by reducing the interaction between fucosylated adhesion molecules supporting leukocyte extravasation. AFUs also impair crucial cell-extracellular matrix (ECM) interactions and presumably subsequent cell signaling pathways, which lead to changes in tumor function and behavior. There are two isoforms of AFUs in human, namely α-L-fucosidase 1 (FUCA1) and α-L-fucosidase 2 (FUCA2), respectively. FUCA1 is a p53 target gene and can hydrolyze different fucosylation sites on epidermal growth factor receptor (EGFR), thereby determining the activation of EGFR. FUCA2 mediates the adhesion between Helicobacter pylori and gastric mucosa and is upregulated in 24 tumor types. Besides, based on the participation of AFU in signaling pathways and tumor progression, we discuss the prospect of AFU as a therapeutic target.

Discovery of a fucoidan endo-4O-sulfatase: Regioselective 4O-desulfation of fucoidans and its effect on anticancer activity in vitro

Fucoidans are a class of sulfated fucose-containing bioactive polysaccharides produced by brown algae. The biological effects exhibited by fucoidans are thought to be related to their sulfation. However, the lack of methods for sulfation control does not allow for a reliable conclusion about the influence of the position of certain sulfate groups on the observed biological effects. We identified the gene encoding the endo-acting fucoidan sulfatase swf5 in the marine bacterium Wenyingzhuangia fucanilytica CZ1127T. This is the first report on the sequence of fucoidan endo-sulfatase. Sulfatase SWF5 belongs to the subfamily S1_22 of the family S1. SWF5 was shown to remove 4O-sulfation in fucoidans composed from the alternating α-(1→3)- and α-(1→4)-linked residues of sulfated L-fucose but not from fucoidans with the α-(1→3)-linked backbone. The endo-sulfatase was used to selectively prepare 4O-desulfated fucoidan derivatives. It was shown that the 4O-desulfated fucoidans inhibit colony formation of DLD-1 and MCF-7 cells less effectively than unmodified fucoidans. Presumably, 4O-sulfation makes a significant contribution to the anticancer activity of fucoidans.

Structure and Biological Activity Analysis of Fucoidan Isolated from Sargassum siliquosum.

Fucoidans are heterologous polysaccharides commonly seen in brown macroalgae and are known for their biological activity including anticancer, antiangiogenic, immunomodulation, and antiviral properties. The brown macroalga Sargassum siliquosum was used for the extraction and analysis of fucoidan in this study. The S. siliquosum fucoidan was indicated as a galactofucoidan composed of sugars, uronate, and sulfate at a ratio of 12:1:4 and its purity was 85% based on the abovementioned three major components. Structural analysis by electrospray ionization collision-induced dissociation tandem mass spectroscopy revealed that the purified fucoidan consisted of a carbohydrate chain composed of (1→3)-linked or (1→4)-linked l-fucose residues, with sulfate groups at C-2 and C-4 positions. Galactose residues with (1→4)-linkages function as the branch points and they are located at the C-3 or C-4 position of fucose residues. Galactose residues are sulfated mainly at C-4 and C-6, while some sulfation can also be seen at C-2. The fucoidan purified from S. siliquosum demonstrated antioxidant, anti-inflammatory, and antilipogenic activities.

α-l-Fucosidases from Bursaphelenchus xylophilus Secretome—Molecular Characterization and Their Possible Role in Breaking Down Plant Cell Walls

The pinewood nematode (PWN) Bursaphelenchus xylophilus, the causal agent of the pine wilt disease (PWD), enters above-ground parts of the tree, migrates through the resin canals and feeds on plant cells causing extensive damage. In order to penetrate the cell wall and establish a parasitic relationship with host trees, the PWN needs to secretea mixture of active cell wall degrading enzymes. In maritime pine, Pinus pinaster, which is high susceptible to PWN, xyloglucan is the major hemicellulosic polysaccharide in primary cells. The xyloglucan backbone is susceptible to hydrolysis by numerous endoglucanases, some of them specific to xyloglucan. However, to completely degrade xyloglucan, all substitutions on the glucan backbones must be released, and l-fucose residues in xyloglucan branches are released by α-l-fucosidases. In the present study, the molecular characterization of two α-l-fucosidases found in PWN secretome was performed. Moreover, a novel α-l-fucosidase was identified and its cDNA and gene sequence were determined. The three-dimensional structures of these α-l-fucosidases were predicted and the transcript levels were analyzed, thus providing new insights into fundamental PWN biology and the possible role of these proteins as cell wall degrading enzymes.

Arabidopsis thaliana α1,2-l-fucosyltransferase catalyzes the transfer of l-galactose to xyloglucan oligosaccharides.

l-Galactose (l-Gal) is one of the components of plant cell wall polysaccharides. In the GDP-l-fucose-deficient Arabidopsis thaliana mutant mur1, l-fucose (l-Fuc) residues in xyloglucan are substituted by l-Gal residues. l-Gal only differs from l-Fuc by the presence of an oxygen at C-6. Thus, we hypothesized that the A. thaliana xyloglucan α1,2-l-fucosyltransferase (AtFUT1) is also responsible for the l-galactosyl transfer to d-galactose residues in xyloglucan. In this study, we heterologously produced AtFUT1 in fission yeast and carried out an in vitro assay for the activities of AtFUT1 on GDP-l-Gal and xyloglucan oligosaccharide. We show that the recombinant AtFUT1 catalyzes l-Gal transfer to xyloglucan oligosaccharides although the initial velocity of l-Gal transfer is 3.1 times lower than that of l-Fuc transfer.

Application of a Janus aglycon with dual function in benzyl-free synthesis of spacer-armed oligosaccharide fragments of polysaccharides from rhizobacterium Azospirillum brasilense sp7

Both protective and pre-spacer features of 4-(2-chloroethoxy)phenyl (CEP) aglycon, which belong to the class of Janus aglycons, were engaged in a benzyl-free synthesis of oligosaccharide fragments of polysaccharides from rhizobacterium Azospirillum brasilense sp7. Introduction of α-1,4-linked L-fucose residue was performed using 3,4-di-O-benzoyl-2-O-triisopropylsilyl-α-L-fucopyranosyl N-phenyltrifluoroacetimidate in excellent stereoselectivity and high yields. The obtained deprotected di-, tri- and tetrasaccharides contain 4-(2-azidoethoxy)phenyl (AEP) spacer aglycon, which allows straightforward preparation of neoglycoconjugates that will be used for the study of the role of lipopolysaccharide of rhizobacterium A. brasilense sp7 in plant–microbe symbiosis. The intermediate protected oligosaccharide building blocks with cleavable CEP/AEP aglycons have a strong potential for further application in the synthesis of more complex oligosaccharides.

Induction of apoptosis by fucoidan isolated from a traditional food, Saccharina longissima, in U937 human leukemia cells

Background: A brown seaweed, Saccharina longissima (Laminaria angustata var. longissima) has been consumed in Okinawa for 300 years. It is native to the southernmost part of Japan, in Okinawa, but grows on the Pacific coast (Kushiro and Nemuro) in Hokkaido in the northernmost part of Japan. Previously, we reported the structural characteristics from S. longissima. The fucoidan consisted of L-fucose, D-galactose, D-glucose, D-xylose, D-glucuronic acid, and sulfuric acid. This fucoidan activated the RAW 264.7 murine macrophage cell line. The aim of this study was to investigate the effects of a fucoidan isolated from Saccharina longissima on cell anti-proliferation and apoptosis in the U937 human leukemia monocyte lymphoma cell line. Methods: U937 cells were incubated with four fucoidan concentrations. The degree of apoptosis was determined using the APOPercentage APOPtosis assay, caspaer-3 activity assay and Western blotting analysis assay. Results: The treatment of cell with a fucoidan highly substituted with sulfate, in addition to L-fucose residues, inhibited cell growth. Apoptosis increased in U937 cells in a dose-dependent manner following the addition of fucoidan. However, the fucoidan did not induce apoptosis in the presence of caspase inhibitor (z-VAD-FMK).Conclusions: The fucoidan from Saccharina longissima induced apoptosis in U937 cells. Accordingly, the kelp containing this fucoidan may contribute to the observed longevity of the Okinawan population.Keywords: fucoidan, Saccharina longissimi, brown seaweed, traditional food, human leukemia cells, apoptosis

Studies on the O-polysaccharide of Escherichia albertii O2 characterized by non-stoichiometric O-acetylation and non-stoichiometric side-chain l-fucosylation

An O-polysaccharide was isolated from the lipopolysaccharide of Escherichia albertii O2 and studied by chemical methods and 1D and 2D 1H and 13C NMR spectroscopy. The following structure of the O-polysaccharide was established: ▪. The O-polysaccharide is characterized by masked regularity owing to a non-stoichiometric O-acetylation of an l-fucose residue in the main chain and a non-stoichiometric side-chain l-fucosylation of a β-GlcNAc residue. A regular linear polysaccharide was obtained by sequential Smith degradation and alkaline O-deacetylation of the O-polysaccharide. The content of the O-antigen gene cluster of E. albertii O2 was found to be essentially consistent with the O-polysaccharide structure established.

New branched amino acids for high affinity dendrimeric DC-SIGN ligands

A branched amino acid was synthesized from methyl glucopyranoside; this amino acid presents three amino groups protected by Fmoc and one acid group and can be used in classic peptide synthesis. In parallel, similar azido terminated blocks were synthesized.Successive coupling reaction and deprotection afforded dendrimers with up to 27 azido functional groups. As an example of application, d-mannose and l-fucose residues were linked through CuAAC coupling and resulting glycodendrimers were evaluated in their interaction with DC-SIGN using SPR competition assay.

A nuclear fucosyltransferase-like protein, PtFUT, from marine red alga Pyropia tenera (Rhodophyta) confers osmotic stress tolerance

Pyropia tenera is a marine red alga that grows in the intertidal zone and may lose more than 90% of its water during twice-daily low tides. Based on the differentially expressed genes (DEGs) identified from the desiccation transcriptome of P. tenera gametophyte, a complementary DNA was isolated that upregulated strongly under desiccation stress and encoded a polypeptide of 591-amino acid residues showing a sequence homology with α-1,2-fucosyltransferase, named PtFUT. Fucosyltransferases transfer an l-fucose residue to various acceptor molecules, but information of their activity in the red algae is limited. PtFUT showed upregulation in transcription in gametophytes of P. tenera under desiccation and osmotic stress conditions induced by mannitol. Transcription of the PtFUT gene was also increased by abscisic acid as well as H2O2 treatment. The presence of a signal peptide for nucleus localization and fluorescence of the PtFUT-GFP fusion proteins in tobacco protoplasts indicated that PtFUT is located in the nucleus. When PtFUT was overexpressed in Chlamydomonas, the PtFUT improved the growth of transgenic cells under osmotic stress. These results suggest that PtFUT may play a role in tolerance to desiccation stress and provide insights into the molecular functions of the fucosyltransferases and understanding the desiccation tolerance mechanisms in marine red algae growing in intertidal zones.

Morphological and functional aspects of the epidermis of the sea lamprey Petromyzon marinus throughout development.

The development of the epidermis of sea lamprey Petromyzon marinus along the whole life cycle was studied using conventional staining techniques and lectin histochemistry. The epidermis undergoes variations in morphology and thickness throughout development. The simple cuboidal epithelium found in the epidermis of prolarvae becomes stratified cubic in the adult by increasing the number of cell layers. The cuticle thickness undergoes a steady increase during the larval period. There are changes in the glycoconjugate composition of the three main cell types of the P. marinus epidermis, mucous, granular and skein cells, which are more pronounced after metamorphosis. The Alcian blue-periodic acid Schiff (AB-PAS) histochemical method shows the presence of both acidic and neutral glycoconjugates in the mucous cells, indicating their secretory function. Moreover, lectin analysis reveals a mucous secretion containing glycoconjugates such as sulphated glycosaminoglycans (N-acetylglucosamine and N-acetylgalactosamine) and N-glycoproteins rich in mannose. Although granular cells are AB-PAS negative, they exhibit a similar glycoconjugate composition to the mucous cells. Moreover, granular cells show sialic acid positivity in larvae but this monosaccharide residue is not detected after metamorphosis. The skein cells, a unique cell of lampreys, are negative to AB-PAS staining but they mostly contain l-fucose and sialic acid residues, which also disappear after metamorphosis. The function of the granular and skein cells is still unknown but the role of their glycoconjugate composition is discussed. In addition, a different cellular origin is suggested for these two types of cells.

Application of lectinhistochemistry to study the formation and reactivity of the marginal transitional zone of the joint capsule

Aim. To analyze the formation and reactivity of the intermediate part (marginal tranzitional zone) of joint capsule using lectinhistochemistry. Methods and results. Hip joint of laboratory rats was as an object for present investigation. Method of intrafoetal antigen injection has been taken as а pattern in order to study the reactivity of above-mentioned zone. Fragments of hip joints were fixed in picroformol, decalcinated and dehydrated. We used a standard set of lectins, conjugated with horseradish peroxidase for lectinhistochemistry. In the intermediate part, we distinguished regio superficialis and regio profunda . The intensity of expression for lectin receptors in regio superficialis was always higher than in the parietal part of the joint capsule. The intermediate part is distinguishable from the surrounding tissues as early as on the 1 st day, meanwhile regio superficialis and regio profunda are discernible from each other only on the 7 th day. The intermediate part formation is associated with a decrease in the expression of lectin receptors in regio superficialis and an increase in the content of galactose-, mannose- and glycoconjugates in regio profunda occurring from the 14 th day. Changes are more pronounced in regio profunda in comparison with regio superficialis and consist in their premature development, which is accompanied by early expression of a large number of galactoseconjugates in regio profunda on the 7 th day, with a subsequent decrease in galactose – and mannose saturation in it and appearance of significant L-fucose residues amount. Conclusions. Antigenic effect leads to tension in immuno-biological relationship between articular cartilage and joint capsule and following reactive processes affecting lectin receptors glycosylation of intermediate part. Premature glycosylation of regio profunda is aimed on the early formation of the intermediate part of joint capsule having the lectin-mediated barrier, preventing excessive invasion of the joint capsule into the articular cartilage.

Primary structure, conformation in aqueous solution, and intestinal immunomodulating activity of fucoidan from two brown seaweed species Sargassum crassifolium and Padina australis

We studied the structure of fucoidans extracted from two brown seaweed species, Sargassum crassifolium and Padina australis, and their intestinal immunomodulating activity via Peyer’s patch cells of C3H/HeJ mice. ESI–MS analysis indicated that the dominant structure of both fucoidans has a backbone of α-(1→4)-linked and α-(1→3)-linked l-fucose residues and sulfate groups are attached at the C-2 and C-4 positions; branches of fucoidan from S. crassifolium are galactose residues with (1→4)- linkage and branching points are at C-4 of fucose, while fucoidan from P. australis, branches are sulfated galactose-fucose disaccharides and sulfated galactose monosaccharides attached to the main chain through (1→3)- or (1→4)- linkages. According to small angle X-ray scattering (SAXS) measurements, the two fucoidans have a branched structure. We simulated them with molecular models based on our proposed primary structure. These fucoidan samples have the ability to stimulate intestinal immunological activity via Peyer’s patch cells.

Lectin histochemistry shows the comparative biosynthesis and cellular biodistribution of alpha l-fucose residues in some tissues of tetrapoda representatives

Fucose is a monosaccharide that plays several immunological roles. This study investigated the comparative biosynthesis and cellular biodistribution of fucose residues in some tissues of tetrapoda representatives using lectin histochemistry. In this study, the mouse was used as a representative for mammalian, pigeon for avian, lizard for reptilian, and toad for amphibians. The localization of the fucose residues was seen in several cell types of mice ileum, such as villi microfold (M) cells, goblet cells, some of intestinal crypts cells, and lamina propria cells. In other tetrapoda representatives, fucose was only seen in M cells of lizard ileum and some cells of villi lamina propria of pigeon, lizard, and toad. It was also observed in the pancreatic acinar cells of the mouse and some cell aggregations of pancreatic parenchyma of the lizard. Contrarily, it was not seen either in pigeon or in toad pancreases parenchyma. Spleen of all animals showed the fucose residues in some splenic cells in the red pulp only, barring the white pulp. The liver parenchyma of all tetrapoda representatives hadn’t fucose residues. The fucose cellular biodistribution in some cells of tetrapoda representatives differed based on the cell type. In the mouse, it was highly seen in the apical cytoplasm of the villi M cells as well as in the cup-like part of goblet cells. In addition, it was seen as “rings” in the granule membranes of the Ulex europeaus agglutinin I (UEAI+) cells in the intestinal crypts cells. Furthermore, the UEAI+ cells in the lamina propria showed fucose granules in their cytoplasm. There is no clear evidence about the relation between the cellular biosynthesis of fucose residues and mucosal immune cells. The role of fucose residues in the pancreatic acinar cells are not well understood and need further investigations. In this study, fucose residues were synthesized in several types of cells in the mouse ileum, spleen and pancreas as compared with other tetrapoda. The data obtained from this study can help us to get more information about the cellular biodistribution and synthesis of fucose residues in several animal species rather than mammalians.

H-ficolin binds Aspergillus fumigatus leading to activation of the lectin complement pathway and modulation of lung epithelial immune responses.

Aspergillus fumigatus is an opportunistic fungal pathogen that typically infects the lungs of immunocompromised patients leading to a high mortality. H-Ficolin, an innate immune opsonin, is produced by type II alveolar epithelial cells and could participate in lung defences against infections. Here, we used the human type II alveolar epithelial cell line, A549, to determine the involvement of H-ficolin in fungal defence. Additionally, we investigated the presence of H-ficolin in bronchoalveolar lavage fluid from transplant patients during pneumonia. H-Ficolin exhibited demonstrable binding to A.fumigatus conidia via l-fucose, d-mannose and N-acetylglucosamine residues in a calcium- and pH-dependent manner. Moreover, recognition led to lectin complement pathway activation and enhanced fungal association with A549 cells. Following recognition, H-ficolin opsonization manifested an increase in interleukin-8 production from A549 cells, which involved activation of the intracellular signalling pathways mitogen-activated protein kinase MAPK kinase 1/2, p38 MAPK and c-Jun N-terminal kinase. Finally, H-ficolin concentrations were significantly higher in bronchoalveolar lavage fluid of patients with lung infections compared with control subjects (n=16; P=0·00726). Receiver operating characteristics curve analysis further highlighted the potential of H-ficolin as a diagnostic marker for lung infection (area under the curve=0·77; P<0·0001). Hence, H-ficolin participates in A.fumigatus defence through the activation of the lectin complement pathway, enhanced fungus-host interactions and modulated immune responses.

Are multifunctional marine polysaccharides a myth or reality?

Are multifunctional marine polysaccharides a myth or reality?