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Cell aggregation in the marine sponge Microciona prolifera is mediated by a multimillion molecular-mass aggregation factor, termed MAF. Earlier investigations revealed that the cell aggregation activity of MAF depends on two functional domains: (i) a Ca(2+)-independent cell-binding domain and (ii) a Ca(2+)-dependent proteoglycan self-interaction domain. Structural analysis of involved carbohydrate fragments of the proteoglycan in the self-association established a sulfated disaccharide beta-D: -GlcpNAc3S-(1-->3)-alpha-L: -Fucp and a pyruvated trisaccharide beta-D: -Galp4,6(R)Pyr-(1-->4)-beta-D: -GlcpNAc-(1-->3)-alpha-L: -Fucp. Recent UV, SPR, and TEM studies, using BSA conjugates and gold nanoparticles of the synthetic sulfated disaccharide, clearly demonstrated self-recognition on the disaccharide level in the presence of Ca(2+)-ions. To determine binding forces of the carbohydrate-carbohydrate interactions for both synthetic MAF oligosaccharides, atomic force microscopy (AFM) studies were carried out. It turned out that, in the presence of Ca(2+)-ions, the force required to separate the tip and sample coated with a self-assembling monolayer of thiol-spacer-containing beta-D: -GlcpNAc-(1-->3)-alpha-L: -Fucp-(1-->O)(CH(2))(3)S(CH(2))(6)S- was found to be quantized in integer multiples of 30 +/- 6 pN. No binding was observed between the two monolayers in the absence of Ca(2+)-ions. Cd(2+)-ions could partially induce the self-interaction. In contrast, similar AFM experiments with thiol-spacer-containing beta-D: -Galp4,6(R)Pyr-(1-->4)-beta-D: -GlcpNAc-(1-->3)-alpha-L: -Fucp-(1-->O)(CH(2))(3)S(CH(2))(6)S- did not show a binding in the presence of Ca(2+)-ions. Also TEM experiments of gold nanoparticles coated with the pyruvated trisaccharide could not make visible aggregation in the presence of Ca(2+)-ions. It is suggested that the self-interaction between the sulfated disaccharide fragments is stronger than that between the pyruvated trisaccharide.

The acceptor specificities of ST3Gal III, ST3Gal IV, ST6Gal I and ST6Gal II were investigated using a panel of beta-D-Galp-(1-->4)-beta-D-GlcpNAc-(1-->2)-alpha-D-Manp-(1-->O)(CH(2))(7)CH(3) analogues. Modifications introduced at either C2, C3, C4, C5, or C6 of terminal D-Gal, as well as N-propionylation instead of N-acetylation of subterminal D-GlcN were tested for their influence on the alpha-2,3- and alpha-2,6-sialyltransferase acceptor activities. Both ST3Gal enzymes displayed the same narrow acceptor specificity, and only accept reduction of the Gal C2 hydroxyl function. The ST6Gal enzymes, however, do not have the same acceptor specificity. ST6Gal II seems less tolerant towards modifications at Gal C3 and C4 than ST6Gal I, and prefers beta-D-GalpNAc-(1-->4)-beta-D-GlcpNAc (LacdiNAc) as an acceptor substrate, as shown by replacing the Gal C2 hydroxyl group with an N-acetyl function. Finally, a particularly striking feature of all tested sialyltransferases is the activating effect of replacing the N-acetyl function of subterminal GlcNAc by an N-propionyl function.

The immunogenic O-glycan of circulating anodic antigen (CAA) is a high-molecular-mass polysaccharide with the unique -->6)-[beta-D-GlcpA-(1-->3)]-beta-D-GalpNAc-(1--> repeating unit. To obtain information at the molecular level about the specificity of monoclonal antibodies against CAA, the immunoreactivity of two series of bovine serum albumin-coupled synthetic oligosaccharides related to the CAA O-glycan was monitored using ELISA and surface plasmon resonance spectroscopy. The importance of the axial hydroxyl group of beta-D-GalpNAc for antibody binding was investigated using the following series of analogues: beta-D-GlcpA-(1-->3)-beta-D-GlcpNAc-(1-->O); beta-D-GlcpNAc-(1-->6)-[beta-D-GlcpA-(1-->3)]-beta-D-GlcpNAc-(1-->O); and beta-D-GlcpA-(1-->3)-beta-D-GlcpNAc-(1-->6)-[beta-D-GlcpA-(1-->3)]-beta-D-GlcpNAc-(1-->O). In the second series of analogues, beta-D-Glcp6S-(1-->3)-beta-D-GalpNAc-(1-->O), beta-D-GalpNAc-(1-->6)-[beta-D-Glcp6S-(1-->3)]-beta-D-GalpNAc-(1-->O), and beta-D-Glcp6S-(1-->3)-beta-D-Gal-pNAc-(1-->6)-[beta-D-Glcp6S-(1-->3)]-beta-D-GalpNAc-(1-->O), the native beta-D-GlcpA moiety was replaced by beta-D-Glcp6S to evaluate the influence of the nature of the charge on antibody recognition. Comparison of the immunoreactivity of these series with that measured for conjugates containing corresponding synthetic CAA fragments showed that the antibody binding levels can be correlated to the antibody specificity to CAA fragments. For the most reactive antibodies, the structural changes chosen (beta-D-GalpNAc replaced by beta-D-GlcpNAc, and beta-D-GlcpA replaced by beta-D-Glcp6S) completely eradicated the binding.

The gut-associated circulating anodic antigen (CAA) is one of the major excretory antigens produced by the parasite Schistosoma mansoni. The immunoreactive part of CAA is a threonine-linked polysaccharide composed of long stretches of the unique repeating disaccharide-->6)-[beta-D-GlcpA-(1-->3)]-beta-D-GalpNAc-(1-->. Previously, using surface plasmon resonance and ELISA techniques, it has been shown that some anti-CAA IgM monoclonal antibodies (MAbs) also recognize members of a series of bovine serum albumin (BSA)-coupled synthetic di- to penta-saccharide fragments of the CAA glycan. To generate information on the molecular level about the glycan specificity of the relevant IgM MAbs, two series of oligosaccharides related to the CAA disaccharide epitope were synthesized, and coupled to BSA. The first three analogues, beta-D-GlcpA-(1-->3)-[small beta]-D-GlcpNAc-(1-->O), beta-D-GlcpNAc-(1-->6)-[beta-D-GlcpA-(1-->3)]-beta-D-GlcpNAc-(1-->O), and beta-D-GlcpA-(1-->3)-beta-D-GlcpNAc-(1-->6)-[beta-D-GlcpA-(1-->3)]-beta-D-GlcpNAc-(1-->O), wherein the native beta-D-GalpNAc moiety was replaced by beta-D-GlcpNAc, were synthesized to investigate the specificity of the selected MAbs to the carbohydrate backbone of CAA. The second series of analogues, beta-D-Glcp6S-(1-->3)-beta-D-GalpNAc-(1-->O), beta-D-GalpNAc-(1-->6)-[beta-D-Glcp6S-(1-->3)]-beta-D-GalpNAc-(1-->O), and beta-D-Glcp6S-(1-->3)-beta-D-GalpNAc-(1-->6)-[beta-D-Glcp6S-(1-->3)]-beta-D-GalpNAc-(1-->O), wherein the native beta-D-GlcpA moiety was replaced by beta-D-Glcp6S, was synthesized to evaluate the importance of the type/nature of the charge of CAA for the MAb recognition.

A new, powerful method is presented for screening the binding in real time and taking place under dynamic conditions of oligosaccharides to lectins. The approach combines an SPR biosensor and HPLC profiling with fluorescence detection, and is applicable to complex mixtures of oligosaccharides in terms of ligand-fishing. Labeling the oligosaccharides with 2-aminobenzamide ensures a detection level in the fmol range. In an explorative study the binding of RNase B-derived oligomannose-type N-glycans to biosensor-immobilized concanavalin A (Con A) was examined, and an affinity ranking could be established for Man(5)GlcNAc(2) to Man(9)GlcNAc(2), as monitored by HPLC. In subsequent experiments and using well-defined labeled as well as nonlabeled oligosaccharides, it was found that the fluorescent tag does not interfere with the binding and that the optimum epitope for the interaction with Con A comprises the tetramannoside unit Manalpha2Manalpha6(Manalpha3)Man[D(3)B(A)4'], rather than the generally accepted trimannoside Manalpha6 (Manalpha3)Man [B(A)4' or 4(4')3]. In a similar experimental setup, the interaction of various fucosylated human milk oligosaccharides with the fucose-binding lectin from Lotus tetragonolobus purpureaus was studied, and it appeared that oligosaccharides containing blood group H could selectively be retained and eluted from the lectin-coated surface. Finally, using the same lectin and a mixture of O-glycans derived from bovine submaxillary gland mucin, minor constituents but containing fucose could selectively be picked from the analyte solution as demonstrated by HPLC profiling.

Starting from a tumor-associated synthetic MUC1-derived peptide MUC1a' and using a completely enzymatic approach for the synthesis of the core-2 sialyl Lewis X glycopart, the following glycopeptide was synthesized: AHGV[Neu5Ac(alpha2-3)Gal(beta1-4)[Fuc(alpha1-3)]GlcNAc(beta1-6)[Gal(beta1-3)]GalNAc(alpha1-O)]TSAPDTR. First, polypeptide N-acetylgalactosaminyltransferase 3 was used to site-specifically glycosylate MUC1a' to give MUC1a'-GalNAc. Then, in a one-pot reaction employing beta-galactosidase and core-2 beta6-N-acetylglucosaminyltransferase the core-2 O-glycan structure was prepared. The core-2 structure was then sequentially galactosylated, sialylated, and fucosylated by making use of beta4-galactosyltransferase 1, alpha3-sialyltransferase 3, and alpha3-fucosyltransferase 3, respectively, resulting in the sialyl Lewis X glycopeptide. The overall yield of the final compound was 23% (3.2 mg, 1.4 micromol). During the synthesis three intermediate glycopeptides containing O-linked GalNAc, Gal(beta1-4)GlcNAc(beta1-6)[Gal(beta1-3)]GalNAc, and Neu5Ac(alpha2-3)Gal(beta1-4)GlcNAc(beta1-6)[Gal(beta1-3)]GalNAc, respectively, were isolated in mg quantities. All products were characterized by mass spectrometry and NMR spectroscopy.

A series of well-defined oligosaccharide fragments of the capsular polysaccharide of Streptococcus pneumoniae type 3 has been generated. Partial-acid hydrolysis of the capsular polysaccharide, followed by fractionation of the oligosaccharide mixture by Sepharose Q ion-exchange chromatography yielded fragments containing one to seven [-->3)-beta-D-GlcpA-(1-->4)-beta-D-Glcp-(1-->] repeating units. The isolated fragments were analysed for purity by high-pH anion-exchange chromatography with pulsed amperometric detection (HPAEC-PAD) using an IonPac AS11 column, and their structures were verified by 1H NMR spectroscopy and nano-electrospray mass spectrometry. The oligosaccharides can be used to produce neoglycoprotein vaccines with a defined carbohydrate part.

The chemical synthesis of alpha-L-Fucp-(1 --> 3)-beta-D-GalpNAc-(1 --> 4)-beta-D-GlcpNAc-(1 --> 3)-alpha-D-GalpO(CH2)5NH2, beta-D-GalpNAc-(1 --> 4)-[alpha-L-Fucp-(1 --> 3)-]beta-D-GlcpNAc-(1 --> 3)-alpha-D-GalpO(CH2)5NH2, and alpha-L-Fucp-(1 --> 3)-beta-D-GalpNAc-(1 --> 4)-[alpha-L-Fucp-(1 --> 3)-]beta-D-GlcpNAc-(1 --> 3)-alpha-D-GalpO(CH2)5NH2 is described. These structures represent fucosylated oligosaccharide fragments of the glycocalyx glycan of the cercarial stage of the parasite Schistosoma mansoni, and in protein-conjugated form they are potential diagnostics in the search for antibodies raised against the glycan in the serum of infected humans.

The primary structures of 21 novel monoantennary and diantennary N-glycans of the glycoprotein αD -hemocyanin (α D-Hc) of Helix pomatia have been determined. Outer oligosaccharide fragments (antennae) were released from the glycoprotein by Smith degradation of an α D-Hc pronase digest. The major antenna, obtained following HPLC fractionation on Lichrosorb-NH2, was characterized using ¹H-NMR spectroscopy, fast-atom-bombardment mass spectrometr3,, and linkage analysis, and corresponds to a pentasaccharide fragment. The intact carbohydrate chains of αD-Hc were released with peptide-N4-(N-acetyl-β-glucosaminyl)asparagine amidase-F digestion, separated from the protein on Bio-Gel P-100, and subfractionated on Bio-Gel P-4. A portion of subfractions was reduced with sodium borodeuteride, and the non-reduced and reduced .samples were further fractionated on CarboPac PA-1. Lichrosorb-NH2/Lichrosphere-NH2, and/or Lichrosphere-C18. Purified oligosaccharides and oligosaccharide-alditols were analyzed using 500⁄600-MHz ¹H-NMR spectroscopy. In total, four novel types of antenna were identified, namely. [This equation cannot be represented into ASCII Text] which are all attached to O-2 of αMan residues of the trimannosyl-N,N′-diacetylchitobiose core element. which is generally β-l,2-xylosylated and α-1,6-fucosylated. Man(α1–6)[Man(α1–3)l[±Xyl(β1–2)]Man(β1–4)GlcNAc(β1–4)[±Fuc(α1–6)]GlcNAc. [ABSTRACT FROM AUTHOR]

Proteoglycans of bovine nasal septal cartilage bear predominantly chondroitin 4-sulfate. After exhaustive chondroitinase ABC digestion of a chondromucoprotein preparation rich in proteoglycans and subsequent reductive β-elimination, five hexasaccharide alditols were isolated from the glycosaminoglycan-protein linkage region. They were analyzed by enzymatic digestion in conjunction with HPLC and by one-dimensional and two-dimensional ¹H-NMR spectroscopy. They share the conventional core saccharide structure Δ4,5HexAα1-3GalNAcβ1-4GlcAβ1-3Galβ1-3Galβ1-4Xyl-ol (where Δ4,5HexA is 4,5-unsaturated hexuronic acid), but have different sulfation profiles. One compound (I) does not contain sulfate. Two of the three monosulfated compounds (II and III) have an O-sulfate group at either C6 or at C4 of the GalNAc residue. The other monosulfated compound (IV) is hitherto unreported and has a O-sulfate at C4 of the Gal residue preceding the GlcA residue, whereas the GalNAc is not sulfated. The disulfated compound (V) has sulfate groups at C4 of both the Gal residue preceding GlcA and the GalNAc residue. The molar ratio of compounds I–V is 38.3:5.9:43.0:1.6:11.2. The structural heterogeneity of these hexasaccharide alditols reflects the polydispersity in the linkage region of the chondroitin sulfate chains. In addition, two trisaccharide and two tetrasaccharide alditols derived from the repeating disaccharide region of the chondroitin sulfate chains were also isolated. Their structures were unambiguously determined by enzymatic analysis and by ¹H-NMR spectroscopy as δΔ4,5HexAα1-3GalNAc(4-O- or 6-O-sulfate)β1-4GlcA-ol and δΔ4,5HexAα1-3GalNAc(4-O- or 6-O-sulfate)β1-4GlcAβ1-3GalNAc(4-O-sulfate)-ol, respectively. [ABSTRACT FROM AUTHOR]

The N-linked carbohydrate chains of porcine zona pellucida glycoproteins were released by digestion with peptide-N4-(N-acetyl-β- glucosaminyl)asparagine amidase F and subsequently separated from the O- glycoprotein by gel-permeation chromatography on Bio-Gel P-100. The O- linked carbohydrate chains were released from the O-glycoprotein by alkaline borohydride treatment. Fractionation of the extremely heterogeneous mixture of O-linked oligosaccharide alditols was achieved by a combination of chromatographic techniques comprising gel- permeation chromatography on Bio-Gel P-4 and P-6, anion-exchange FPLC on Mono Q, and high-pH anion-exchange chromatography on CarboPac PA-1. The primary structures of 32 O-glycans were determined by one- and two- dimensional ¹H-NMR spectroscopy. The major part of the analyzed compounds contain a combination of the structural elements Gal β 1- 4GlcNAc β 1-3Gal β 1-3GalNAc-ol, Gal β 1-4(6SO4 )GlcNAc, and α 2-3-linked Neu5Gc or Neu5Ac. This series of compounds has the following structure, where n = 0 to > 6: [Neu5Gc/Ac α2-3]0-1[Gal β 1-4(6SO4-)GlcNAc β1-3]nGal β 1-4GlcNAc β 1-3Gal β 1- 3GalNAc-ol. In addition, smaller compounds were identified in which the Gal β 1-3GalNAc-ol core is substituted by Neu5Gc/Ac α 2-6-linked to GalNAc-ol and/or Neu5Gc/Ac α 2-3-linked to Gal. Furthermore, oligosaccharides were obtained in which the distribution of 6-O- sulfated GlcNAc residues differs from that in the above-mentioned general structure, and a small portion of the oligosaccharides has the GlcNAc β 1-3GalNAc-ol core structure. Analysis of the endo-β- galactosidase digests of pools of N- and O-glycans indicated that the two types of oligosaccharides contain qualitatively similar poly(N- acetyllactosamine) chains. [ABSTRACT FROM AUTHOR]

The carbohydrate chains of the respiratory-mucus glycoproteins of a patient (blood group O) suffering from bronchiectasis due to Kartagenerߣs syndrome, were released by alkaline borohydride treatment of a pronase digest. The structures of 82 neutral and low-molecular-mass sialylated oligosaccharides have been described previously [van Kuik A., de Waard P., Vliegenthart J. F. G., Klein A., Carnoy C., Lamblin G. Roussel P. (1991) Eur. J. Biochem. 198, 169 — 182]. In the present work, medium-size sialylated oligosaccharides were obtained after ion-exchange chromatography and were subsequently separated into 36 fractions utilizing gel filtration, HPLC on normal-phase alkylamine-bonded silica and reverse-phase HPLC. From these fraction, the following six sialylated hepta- and octa-saccharide-alditols have been characterized by employing 500-MHz [sup 1]H-NMR spectroscopy, in conjunction with fast-atom-bombardment mass spectroscopy and methylation analysis. [ABSTRACT FROM AUTHOR]

From the carbohydrate-protein linkage region of whale cartilage proteoglycans, which bear predominantly chondroitin 4-sulfate, one nonsulfated, two monosulfated and one disulfated hexasaccharide alditols were isolated after exhaustive digestions with Actinase E and chondroitinase ABC, and subsequent β-elimination. Their structures were analyzed by chondroitinase ACII digestion in conjunction with HPLC and by 500-MHz ¹H-NMR spectroscopy. The nonsulfated compound (A) had the following conventional structure: ΔGlcA(β1–3)-GalNAc(β1–4)GlcA(β1–3)Gal(β1–3)Gal(β1–4)Xylol, where GlcA, ΔGlcA and GalNAc are glucuronic acid; 4,5-unsaturated glucuronic acid and 2-deoxy-2-N-acetylamino-D-galactose, respectively. The other compounds were sulfated derivatives of compound A. Two monosulfated compounds (B and C) had an ester sulfate on C4 or C6 of the GalNAc residue, respectively and the disulfated compound (D) had two ester sulfate groups, namely, one on C4 of the GalNAc and the other on C4 of the Gal residue substituted by GlcA. The molar ratio of A/B/C/D was 0.21:0.16:0.36:0.27. The compound containing Gal-4-O-sulfate was previously isolated by us in the form of a sulfated glycoserine [ΔGlcA(β1–3)GalNAc(4-O-sulfate)(β1–4)GlcA(β1–3)Gal(4-O-sulfate)(β1–3)Gal(β1–4)Xyl/β1-O-Ser] from the carbohydrate-protein linkage region of rat chondrosarcoma chondroitin-4-sulfate proteoglycans [Sugahara K., Yamashina, I., DeWaard, P., Van Halbeek, H. & Vliegenthart, J. F. G. (1988) J. Biol. Chem. 263, 10168–10174]. The discovery of this structure in the carbohydrate-protein linkage region of chondroitin 4-sulfate proteoglycans from nontumorous cartilage indicates that it is not a tumor-associated product but rather a physiological biosynthetic product since it represents a significant proportion. The biological significance of this structure is discussed in relation to glycosaminoglycan biosynthesis. [ABSTRACT FROM AUTHOR]

Oligosaccharide-alditols derived from the respiratory mucins of a patient suffering from bronchiectasis were separated by HPLC into 46 fractions. The structures of 11 oligosaccharides with GlcNAcβ(1 → 3)Galβ(1 → 4)GlcNAcβ(1 → 6)GalNAc-ol in common have been established (preceding paper in this journal). In this second part the structures of 12 oligosaccharides were established 8 of which have not been described before. [ABSTRACT FROM AUTHOR]

New neutral oligosaccharides from cow colostrums κ-casein were identified and characterized by 500-MHz ¹H-NMR spectroscopy. Their structures are Galβ(1→3)GalNAc-ol, Galβ(1→3)[GlcNAcβ(1→6)]GalNAc-ol, Galβ(1→3)[Galβ(1→4)GlcNAcβ(1→6)]GalNAc-ol, Galβ(1→3){Fucα(1→3){Galβ(1→4)]GlcNAcβ(1→60}Gal-Nac-ol. The tetrasaccharide and the cow colostrums κ-caseinoglycopeptide which contains this oligosaccharide inhibit the hemagglutination of blood group I human erythrocytes. In cow mature milk only the disaccharide is characterized. The variability of these neutral oligosaccharides in cow κ-casein as a function of time after calving is studied. [ABSTRACT FROM AUTHOR]

Hemocyanin from the freshwater snail Lymnaea stagnalis is a high-molecular-mass copper-containing glycoprotein which functions as oxygen carrier in the hemolymph. To release the carbohydrate chains, the protein was digested by pronase followed by hydrazinolysis and reduction. The oligosaccharide-alditols were purified by gel permeation chromatography on Bio-Gel P-4, followed by HPLC on a Lichrosorb-NH2 column. Using 500-MHz ¹H-NMR spectroscopy, in conjunction with sugar, methylation and deamination analysis, the following seven novel primary oligosaccharide structures could be unravelled. [ABSTRACT FROM AUTHOR]

Glycoproteins were isolated from the ovary of the starfish Asterias rubens (L.). After delipidation, sugar analysis revealed the presence of mannose, glucose and N-acetylglucosamine in a molar ratio of 9.0:1.3:2.3. Subsequently, hydrazinolysis, re-N-acetylation, reduction and high-voltage paper electrophoresis were carried out, resulting in a mixture of neutral oligosaccharide alditols which was fractionated on Bio-Gel P-4. The alditols, investigated by 500-MHz ¹H-NMR spectroscopy, turned out to be of the oligomannose type or of the glucooligomannose type containing 9 mannose and 1-3 glucose residues. The most abundant compounds were established to be: Manα1 → 2Manα1 → 6 Manα1 →6 6 Manα1 → 2Manα1 → 3 Manβ1→ 4GlcNAcβ1 → 4GlcNAc-ol Manα1 → 2Manα1 → 2Manα1 → 3 and Manα1 → 2Manα1 → 6Man&alpha :1→ 6 Man&alpha:1 &rarr:2M anα1→3 Manβ1→ β1 → 4GlcNAc·ol Glcα1 → 3Manα1 → 2Manα1 → 2Manα1 → 3. [ABSTRACT FROM AUTHOR]

The carbohydrate chains of the bronchial-mucus glycoproteins of six cystic fibrosis patients with blood group O were released by alkaline borohydride treatment. Low-molecular-mass, monosialyl oligosaccharide-alditols were isolated by anion-exchange chromatography and fractionated by high-performance liquid chromatography. Structural characterization was performed by 500-MHz ¹H-NMR spectroscopy in combination with quantitative sugar analysis. The established structures range in size from tetra- up to heptasaccharides. They are all sialyl analogs of neutral oligosaccharides that were characterized previously [Lamblin G., Boersma A., Lhermitte M., Roussel P., Mutsaers J. H. G. M., Van Halbeek H. & Vliegenthart J. F. G. (1984) Eur. J. Biochem. 143, 227–236]. The NeuAc residue was found to occur either in a(2→3)-linkage to Gal, or in α(2→6)-linkage to GalNAcol or Gal. [ABSTRACT FROM AUTHOR]

Hemocyanin from the freshwater snail Lymnaea stagnalis is a high-molecular-mass copper-containing oxygen-transport protein, which occurs freely dissolved in the hemolymph. It is a glycoprotein containing fucose, xylose, 3-O-methylmannose, 3-O-methylgalactose, mannose, galactose, N-acetylgalactosamine and N-acetylglucosamine residues as sugar constituents. The N-glycosidic carbohydrate chains of this glycoprotein were released by hydrazinolysis of a pronase digest and subsequently fractionated as oligosaccharide-alditols on Bio-Gel P-4 followed by Lichrosorb-NH2. Investigation with 500-MHz ¹H-NMR spectroscopy, in conjunction with sugar and methylation analysis revealed the lowest-molecular-mass glycan chain to have the structure: ... [ABSTRACT FROM AUTHOR]

The article discusses the primary structure of a new tetraantennary glycan of the N- acetyllactosaminic type isolated from human factor VIll/von Willebrand factor. In the present study, N-glycosylpeptides and oligosaccharide-alditols released by alkaline reductive treatment have been fractionated using serial affinity chromatography on ConA-Sepharose and LCA-Sepharose. By this way, a minor glycopeptide not bound to ConA but retained by and eluted from LCA lectin has been purified. By associating data obtained by permethylation analysis and 500-MHz H-NMR spectroscopy, it proposes a monosialylated, monofucosylated tetraantennary structure of the N-acetyllactosaminic type which has not previously been described in human glycoproteins.

Alkaline borohydride reductive cleavage (β-elimination) of glycophorin A isolated from one individual of the rare blood group Cad, resulted in the release of six acidic oligosaccharide-alditols which were separated by high-performance liquid chromatography (HPLC) on an alkyl amine silicagel column. The structure of four of them has been determined by the application of methanolysis, methylation analysis and ¹H-NMR spectroscopy at 500 MHz. The structures and relative amounts were as follows: oligosaccharide 1: NeuAc(α2-3)Gal(β1- 3)GalNAc-ol (3.5%); oligosaccharide 3: GalNAc(β1- 4)[NeuAc(α2 - 3)]Gal(β1- 3)GalNAc-ol (10.5%); oligosaccharide 5: NeuAc(α2 - 3)Gal(β1- 3)[NeuAc(α2 - 6)]GalNAc-ol (10.4%); oligosaccharide 6: GalNAc(β1- 4)[NeuAc(α2-3)]Gal(β1- 3)[NeuAc(α2-6)]GalNAc-ol (71.2%). The two other oligosaccharides (2 and 4) were obtained in very low amount. The major pentasaccharide (oligosaccharide 6) carries the blood group Cad determinant and is a potent inhibitor of human anti-Sda antibody. [ABSTRACT FROM AUTHOR]

From bronchial mucins of cystic fibrosis patients with blood group O, the carbohydrate chains were released in the form of oligosaccharide-alditols by alkaline borohydride treatment. Application of high-performance liquid chromatography directly on the pool of neutral oligosaccharides afforded 23 fractions. Twenty oligosaccharide structures were characterized by employing 500-MHz 1H-NMR spectroscopy in conjunction with sugar analysis. Thirteen among these had been revealed before to occur in human bronchial nucins [Van Halbeek, H., Dorland, L., Vliegenthart, J. F. G., Hull, W. E., Lamblin, G., Lhermitte, M., Boersma, A., and Roussel, P. (1982) Eur. J. Biochem. 127, 7–20], when paper-chromatographic fractionation of this pool of neutral oligosaccharides was employed. High-performance liquid chromatography enabled to obtain another seven pentassacharide and hexasaccharide alditols; the largest-size representatives are: [This symbol cannot be presented in ASCII format] Thereby, this approach afforded deeper insight into the structural heterogeneity displayed by the carbohydrate chains of bronchial mucins. [ABSTRACT FROM AUTHOR]

The urine of two patients with the infantile form of Gaucher's disease were analyzed for their carbohydrate content. Nine sialyl-oligosaccharides were isolated and characterized. Compounds with identical structures occur in the urine of patient with sialidosis. Furthermore, an abnormal excretion of mannosyl-glycoasparagines was observed. Three of these compounds were isolated and identified: This carbohydrate material, which is found also in the spleen of the patients, probably originates from obstruction of lysosomal functioning by the accumulation of an excessive amount of glucocerebroside. [ABSTRACT FROM AUTHOR]

In blood-group H substance from pig stomach cell linings, carbohydrate structures are known to occur which are O-glycosidically linked via GalNAc to Ser or Thr of the polypeptide backbone. They have in common the Gal(β1→3)GalNAc core unit, which bears one or more N-acetyllactosamine branches. The latter are terminated either by Fuc in (α1→2) linkage or by GlcNAc in (α1→4) linkage to Gal. Both terminal sequences are considered to be porcine blood-group H antigenic determinants. Employing 500-MHz 1H-NMR spectroscopy, the spectral features of the oligosaccharide-alditols corresponding to these chains were established. Insight could be gained into the microheterogeneity displayed by some of the alditol fractions as to the distribution of the terminating Fuc and GlcNAc residues over the various branches. Such information can hardly be obtained using other approaches. [ABSTRACT FROM AUTHOR]

Alkaline borohydride reductive cleavage (β-elimination) of hog submaxillary glycoproteins from three immunologically determined phenotypes, viz. A+, H+ and A-H-, resulted in the release of a series of neutral and acidic oligosaccharide-alditols. 360-MHz ¹H-NMR spectroscopy in combination with methylation analysis and mass spectrometry were used for reinvestigation of the structures of these oligosaccharide-alditols. All are partial structures representing the possible complete and biosynthetically incomplete stages of the chain of a pentasaccharide-N-acetylgalactosaminitol, present in the glycoprotein with blood-group-A activity: GalNAc(α1↵3)Gal(β1↵3)GalNAc-ol [This symbol cannot be presented in ASCII format] Fuc NeuGc. In this way, a prolonged argument about the occurrence of a NeuGc(α2↵6)Gal moiety in these carbohydrate chains, suggested by Aminoff et al. [Aminoff, D., Baig, M. M. and Gathmann, W. D. 11979) J. Biol. Chem. 254, 1788–1793 and 8909–8913] has been brought to a definite end. In the investigated oligosaccharide-alditols N-glycoloylneuraminic acid (NeuGc) is in no case attached to galactose (Gal), but, if present, it is (<2↵6)-linked to N-acetylgalactosaminitol (GalNAc-ol). [ABSTRACT FROM AUTHOR]

360-MHz proton nuclear magnetic resonance spectra were recorded of 10 sialyl-oligosaccharides isolated from urine of sialidosis patients. Their structures are related to the complex asparaginelinked glycan chains of glycoproteins. By correlation of these spectra and comparison with spectra of reference glycopeptides and sialyl-lactose isomers it was possible to assign all signals belonging to anomeric, mannose H-2, sialic acid H-3 and N-acetyl protons. The number of the constituting monosaccharide residues of the oligomers can be obtained by integration of the above-mentioned signals. The chemical shifts of the anomeric and mannose H-2 protons give information about the type of glycan structure (mono-, bi-, triantennary) and the presence of terminal sialic acid at each of the antennas. The chemical shifts of sialic acid H-3 protons are typical for sialic acid residues in 2 → 3 or 2 → 6 linkage to galactose. [ABSTRACT FROM AUTHOR]