New and more efficient multivalent glyco-ligands for asialoglycoprotein receptor of mammalian hepatocytes.

New multi-valent, carbohydrate ligands that contain terminal N-acetylgalactosamine (GalNAc) or lactose (Lac) were prepared using a nitrilotriacetic acid (NTA) derivative of L-lysine as scaffold. Tri-valent structures were prepared by attaching an ω-amino glycoside of GalNAc or Lac to each of the three carboxyl groups of N(ε)-protected N(α)-dicarboxymethyl-L-lysine. In addition, a hexa-valent lactoside was synthesized by attaching N(ε)-deprotected trivalent lactoside to each of the carboxyl group of N(α)-(trifluoroacetamido)hexanoyl L-aspartic acid.

Survey of immune-related, mannose/fucose-binding C-type lectin receptors reveals widely divergent sugar-binding specificities.

C-type lectins (CTLs) are proteins that contain one or more carbohydrate-recognition domains (CRDs) that require calcium for sugar binding and share high degree of sequence homology and tertiary structure. CTLs whose CRD contain EPN (Glu-Pro-Asn) tripeptide motifs have potential to bind mannose (Man), N-acetylglucosamine (GlcNAc), glucose (Glc) and l-fucose (Fuc), whereas those with QPD (Glu-Pro-Asp) tripeptide motifs bind galactose (Gal) and N-acetylgalactosamine (GalNAc). We report here for the first time a direct comparison of monosaccharide (and some di- and trisaccharides)-binding characteristics of 11 EPX-containing (X = N, S or D) immune-related CTLs using a competition assay and an enzyme-linked immunosorbent assay, and neoglycoproteins as ligand.

Oral tolerance to food-induced systemic anaphylaxis mediated by the C-type lectin SIGNR1.

We propose that a C-type lectin receptor, SIGNR-1 (also called Cd209b), helps to condition dendritic cells (DCs) in the gastrointestinal lamina propria (LPDCs) for the induction of oral tolerance in a model of food-induced anaphylaxis. Oral delivery of BSA bearing 51 molecules of mannoside (Man(51)-BSA) substantially reduced the BSA-induced anaphylactic response. Man(51)-BSA selectively targeted LPDCs that expressed SIGNR1 and induced the expression of interleukin-10 (IL-10), but not IL-6 or IL-12 p70.

Functional interaction of common allergens and a C-type lectin receptor, dendritic cell-specific ICAM3-grabbing non-integrin (DC-SIGN), on human dendritic cells.

Fucosylated glycans on pathogens are known to shape the immune response through their interaction with pattern recognition receptors, such as C-type lectin receptors (CLRs), on dendritic cells (DCs). Similar fucosylated structures are also commonly found in a variety of allergens, but their functional significance remains unclear. To test a hypothesis that allergen-associated glycans serve as the molecular patterns in functional interaction with CLRs, an enzyme-linked immunosorbent assay-based binding assay was performed to determine the binding activity of purified allergens and allergen extracts.

Matrix-assisted laser desorption/ionization mass spectrometry of polysaccharides with 2',4',6'-trihydroxyacetophenone as matrix.

So far, there have been only a few matrices reported for detection of polysaccharides with molecular weight higher than 3000 Daltons by matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS). In this work, we found that 2',4',6'-trihydroxyacetophenone (THAP) is a good matrix for MALDI time-of-flight MS analysis of polysaccharides with broad mass range. Large polysaccharides, dextrans, glycoproteins and polysialic acids have been successfully detected by MALDI-MS with THAP as matrix.

Antigen coupled with Lewis-x trisaccharides elicits potent immune responses in mice.

Background: Glycoproteins containing Lewis-x (Le(x)) trisaccharides are often associated with the host's adaptive T(H)2-type immunity, but the mechanisms underlying the T(H)2-biased response are at present unclear.

Objective: The modulatory effect of Le(x) or its glycoconjugates on IgE/T(H)2 responses was investigated.

Methods: The levels of serum antibodies and cytokines were analyzed by means of ELISA, RT-PCR, or both.

Carbohydrate ligands of human C-reactive protein: binding of neoglycoproteins containing galactose-6-phosphate and galactose-terminated disaccharide.

Binding of carbohydrate ligand by human C-reactive protein (CRP), in both native form and structurally deviated form (neoCRP or mCRP), was investigated using galactose-6-phosphate (Gal6P)- and Galbeta3GalNAc-containing bovine serum albumin (BSA) derivatives. To this end, we synthesized glycosides of Gal6P and Galbeta3GalNAc that can potentially generate a terminal aldehyde group. omega-Aldehydo glycosides were then conjugated to BSA via reductive amination.

Glycoproteomics: protein modifications for versatile functions. Meeting on glycoproteomics.

Meeting on Glycoproteomics

Carbohydrate-binding properties of human neo-CRP and its relationship to phosphorylcholine-binding site.

Binding characteristics of two types of ligands for human neo-C-reactive protein (neo-CRP), which is a conformationally altered but physiologically relevant form of CRP, were studied fluorometrically by probing CRP immobilized on a polystyrene surface with europium-labeled ligands. Two Eu-ligands used were bovine serum albumin derivatives that contain on average 40 residues of ligand structures, one derivative containing phosphorylcholine (PC) and the other lactosyl residues. The PC-containing ligands required the presence of calcium for binding, whereas galactose-containing derivatives bound in the absence of calcium.

Inhibition of adhesion of type 1 fimbriated Escherichia coli to highly mannosylated ligands.

The inhibitory potencies of a number of mannosides, di- and trivalent mannosides, a set of mannose-terminating dendrimers, and five types of mannose-bearing neoglycoproteins were determined by using a binding assay that measures the binding of (125)I-labeled, highly mannosylated neoglycoprotein to a type 1 fimbriated Escherichia coli (K12) strain in suspension. The IC(50) values (the concentration of inhibitor that causes 50 % reduction in the bound (125)I-ligand to E. coli) obtained by this method were much lower than the equivalent values obtained by hemagglutination or in assays that involve microplate immobilization.

Mapping the binding areas of human C-reactive protein for phosphorylcholine and polycationic compounds. Relationship between the two types of binding sites.

We developed a fluorescence-based assay method for determining ligand binding activities of C-reactive protein (CRP) in solution. Using this method, we compared the phosphorylcholine (PC)- and polycation-based binding activities of human CRP. The PC-based binding required calcium, whereas a polycation (e.