Publications by authors named "Kamerling J"

HPLC analysis of sialic acids released from recombinant variants of human tissue plasminogen activator, human chimeric plasminogen activator, human erythropoietin, and human follitropin, expressed in Chinese hamster ovary cells, demonstrates for each glycoprotein the presence of N-acetylneuraminic acid and N-glycolylneuraminic acid in a ratio of 97:3. Structural analysis by 500 MHz1H-NMR spectroscopy, of the enzymatically released N-linked carbohydrate chains of chimeric plasminogen activator and of erythropoietin, showed that alpha 2-3 linked N-glycolylneuraminic acid can occur in different N-acetyllactosamine type antennary structures.

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Human beta-mannosidosis urine was fractionated by gel permeation chromatography on Bio-Gel P-2 and by high performance liquid chromatography on Partisil 10 SAX. Besides the disaccharide Man beta 1-4GlcNAc as the major component, a sialic acid-containing compound was detected in an amount of 10% compared to that of Man beta 1-4GlcNAc. Structural characterization of the oligosaccharide and of its reduced analogue by sugar composition analysis, methylation analysis, gas-liquid chromatography-mass spectrometry, and 500-MHz 1H NMR spectroscopy gave conclusive evidence for a novel urinary constituent: NeuAc alpha 2-6Man beta 1-4GlcNAc.

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O-alpha-d-Glucopyranosyl-(1----3)-alpha, beta-L-rhamnopyranose (15), O-alpha-D-galactopyranosyl-(1----3)-O-alpha-D-glucopyranosyl-(1----3)-al pha, beta-L-rhamnopyranose (17), O-alpha-D-galactopyranosyl-(1----3)-O-alpha-D-glucopyranosyl-(1----3)- O-alpha-L-rhamnopyranosyl-(1----3)-D-ribitol (23), and O-alpha-D-galactopyranosyl-(1----3)-O-alpha-D-glucopyranosyl-(1----3)- O-alpha-L-rhamnopyranosyl-(1----4)-D-ribitol (27), which are structural elements of the capsular polysaccharides of Streptococcus pneumoniae types 6A and 6B ([----2)-alpha-D-Galp-(1----3)-alpha-D-Glcp-(1----3)-alpha-L-Rhap- (1----X)- D-Rib-ol-(5-P----]n; 6A X = 3, 6B X = 4), have been synthesised. Ethyl 3-O-allyl-2,4,6-tri-O-benzyl-1-thio-beta-D-glucopyranoside (3) was coupled with benzyl 2,4-di-O-benzyl-alpha-L-rhamnopyranoside (4), and subsequent deallylation (----14) and debenzylation gave 15. Condensation of 14 with ethyl 2,3,4,6-tetra-O-benzyl-1-thio-beta-D-galactopyranoside (2) followed by debenzylation gave 17.

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Biologically active recombinant human follitropin has been expressed in Chinese hamster ovary cells. The carbohydrate chains of the recombinant glycoprotein hormone were enzymatically released by peptide-N4-(N-acetyl-beta-glucosaminyl)asparagine amidase F. The oligosaccharides were separated from the N-deglycosylated protein by gel-permeation chromatography on Bio-Gel P-100, and fractionated by a combination of FPLC on Mono Q and HPLC on Lichrosorb-NH2.

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The sugar composition and the electrophoretic mobility in SDS-polyacrylamide gel electrophoresis of the various lipopolysaccharides (LPS) from clinical isolates of Haemophilus influenzae type b (Hib) were determined to correlate epidemiologic data with compositional data. Rabbit sera specific in Ouchterlony immunodiffusion for 10 different LPS (LPS 1-10) reacted with 647 or 690 Hib strains isolated from patients with invasive disease in various continents. Serotype 1 was predominant and was found in 550 isolates (80%).

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Endoglucanase I from the filamentous fungus Trichoderma reesei catalyses hydrolysis and glycosyl-transfer reactions of cello-oligosaccharides. Initial bond-cleaving frequencies determined with 1-3H-labelled cello-oligosaccharides proved to be substrate-concentration-dependent. Using chromophoric glycosides and analysing the reaction products by h.

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1H- and 13C-NMR assignments for the carbohydrate part of the glycopeptide alpha-D-Man-(1----6)-[beta-D-Xyl-(1----2)]-beta-D-Man-(1----4)-beta-D- GlcNAc-(1----4)-[alpha-L-Fuc-(1----3)]-beta-D-GlcNAc-(1----N)-Asn approximately, derived from the proteolytic enzyme bromelain (EC 3.4.22.

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The carbohydrate moieties of equine chorionic gonadotropin alpha and beta subunits were released from the protein backbones by successive treatments with peptide-N4-(N-acetyl-beta-glucosaminyl)asparagine amidase F and alkaline borohydride and then fractionated by FPLC and HPLC. The major N- and O-linked glycans of the beta subunit were characterized by 500-MHz 1H-NMR spectroscopy, showing a remarkable structural heterogeneity for the N-glycosidically linked chains, comprising mono-, di-, tri- and tri'-antennary N-acetyllactosamine type of glycans, being partly alpha 1-6 fucosylated at the Asn-bound GlcNAc residue and having alpha 2-6 and alpha 2-3 linked N-acetyl- and N-acetyl-4-O-acetylneuraminic acid residues as sialic acid constituents. Significant differences in this respect were detected for the partially characterized glycans of the alpha subunit.

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4-Methylumbelliferyl-beta-D-galactopyranoside-6-sulphate was synthesized and used for the determination of galactose-6-sulphate sulphatase activity. Fibroblasts and leucocytes from 12 different Morquio A patients, showed 0.0-2.

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Cultured fibroblasts from two brothers with beta-mannosidosis were shown to accumulate the disaccharide mannosyl-beta(1----4)-N-acetylglucosamine in amounts of 52 and 68 nmol/mg protein. Structural identification of the Man beta(1----4)GlcNAc, isolated by HPLC, was performed by 500-MHz 1H-NMR spectroscopy and sugar composition analysis. Control fibroblasts did not contain a detectable amount of Man beta(1----4)GlcNAc.

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4-Methoxybenzyl 2,4-di-O-benzyl-3-O-[2,4,6-tri-O-benzyl-3-O-(3,4,6-tri-O-benzyl-alpha-D- galactopyranosyl)-alpha-D-glucopyranosyl]-alpha-L-rhamnopyranoside (22), a building block for the alpha-D-Galp-(1----3)-alpha-D-Glcp-(1----3)-alpha-L-Rhap fragment of the capsular polysaccharides of Streptococcus pneumoniae types 6A and 6B [----2)-alpha-D-Galp-(1----3)-alpha-D-Glcp-(1----3)-alpha-L-Rhap-( 1----X)-D- RibOH-(5-P----]n (6A, X = 3; 6B, X = 4) has been synthesised. Ethyl 3-O-allyl-2,4,6-tri-O-benzyl-1-thio-beta-D-glucopyranoside was coupled with 4-methoxybenzyl 2,4-di-O-benzyl-alpha-L-rhamnopyranoside in ether, using methyl triflate as promoter. The resulting alpha-D-Glcp-(1----3)-alpha-L-Rhap derivative was deallylated with KOBut in N,N-dimethylformamide followed by 0.

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The main constituent of the cell wall complex carbohydrate of the scaly green alga Tetraselmis striata Butcher is shown to be 3-deoxy-manno-2-octulosonic acid (42%). In addition two other 2-keto-sugar acids are present, namely, 3-deoxy-5-O-methyl-manno-2-octulosonic acid (7%), the first methylated derivative of 3-deoxy-manno-2-octulosonic acid found in nature, and 3-deoxy-lyxo-2-heptulosaric acid (11%). The characterization of the three 2-keto-sugar acids has been carried out on the corresponding methyl ester methyl glycosides using GLC-MS and 500-MHz 1H-NMR spectroscopy, and on the corresponding reduced alditol acetates using GLC-MS.

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A glycosylated form of recombinant human insulin-like growth factor I (IGF-I) expressed in Saccharomyces cerevisiae was shown to contain mannose as the only carbohydrate constituent. All oligosaccharide chains of the glycoprotein could be released by mild alkaline treatment, and separated from the protein by gel-permeation chromatography on Bio-Gel P-2. The structures of these O-linked carbohydrate chains were determined by 500-MHz 1H-NMR spectroscopy, affording the disaccharide Man alpha 1-2Man as the major component and the tetrasaccharide Man alpha 1-3Man alpha 1-2Man alpha 1-2Man as a minor component.

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The capsular polysaccharide of Streptococcus pneumoniae serotype 6B [----2)-alpha-D-Galp-(1----3)-alpha-D-Glcp-(1----3)-alpha-L-Rhap-( 1----4)- D-RibOH-(5-P----]n was depolymerised under alkaline (NaOH) and acidic (HF) conditions. The former treatment yielded, as the major component, alpha-2-P-Galp-(1----3)-alpha-Glcp-(1----3)-alpha-Rhap-(1----4)-5- P-RibOH. The latter treatment at -16 degrees gave alpha-Galp-(1----3)-alpha-Glcp-(1----3)-alpha-Rhap-(1----4)-Rib OH-(5-P----2)- alpha-Galp-(1----3)-alpha-Glcp-(1----3)-alpha-Rhap-(1----4)-Rib OH and at 4 degrees gave alpha-Galp-(1----3)-alpha-Glcp-(1----3)-alpha-Rhap-(1----4)-Rib OH.

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Hydrophobic conjugates of a series of aldoses have been prepared by reductive amination with octadecylamine and sodium cyanoborohydride, as model compounds for the analysis of reductively aminated oligosaccharides derived from capsular polysaccharides of Streptococcus pneumoniae. In the context of the methanolysis procedure for sugar analysis, g.l.

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Galactosialidosis urine was fractionated by gel-permeation chromatography on Bio-Gel P-6. The obtained sialic acid-containing carbohydrate fractions were purified by reversed-phase chromatography and separated according to charge by medium-pressure anion-exchange chromatography on Mono Q. The Mono Q fractions, being mixtures of sialyloligosaccharides differing mainly in sialic acid-linkage type (alpha 2-3/alpha 2-6), were subfractionated by high-performance liquid chromatography on Lichrosorb-NH2.

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The carbohydrate chains of equine fibrinogen were enzymatically released by peptide-N4-(N-acetyl-beta-glucosaminyl)asparagine amidase F. The oligosaccharides obtained were fractionated by a combination of FPLC and HPLC and analyzed by 500-MHz 1H-NMR spectroscopy. Four monosialo and four disialo diantennary N-acetyllactosamine type of carbohydrate chains occur: (formula; see text)

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The synthesis of the oligosaccharides beta-D-Xylp-(1----2)-beta-D-Manp-OMe (12), beta-D-Xylp-(1----2)-[alpha-D-Manp-(1----6)]-beta-D-Manp+ ++-OMe (17), beta-D-Xylp-(1----2)-[alpha-D-Manp-(1----3)]-beta-D-Manp+ ++-OMe (21), and beta-D-Xylp-(1----2)-[alpha-D-Manp-(1----3)] [alpha-D-Manp-(1----6)]-beta-D-Manp-OMe (25) is described. Methyl 3-O-benzyl-4,6-O-isopropylidene-beta-D-mannopyranoside (6) was prepared from the corresponding glucoepimer (4) by oxidation, followed by stereoselective reduction. Condensation of 6 with 2,3,4-tri-O-acetyl-alpha-D-xylopyranosyl bromide in the presence of mercuric cyanide gave a 1:9 mixture of methyl 3-O-benzyl-4,6-O-isopropylidene-2-O-(2,3,4- tri-O-acetyl-alpha- (7a) and -beta-D-xylopyranosyl)-beta-D-mannopyranoside (7), and then 7 was converted into the acetylated disaccharide-glycoside 11.

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Alkaline borohydride treatment of the cellulosome of Clostridium thermocellum yielded two major oligosaccharide-alditols, namely D-Galp-beta(1----4)-D-GalOH and (formula; see text) The compounds, isolated via gel permeation chromatography and high performance liquid chromatography, were analyzed by monosaccharide analysis, methylation analysis, gas-liquid chromatography/mass spectrometry, fast atom bombardment/mass spectrometry, and one- and two-dimensional 500-MHz (COSY, HOHAHA, ROESY) 1H NMR spectroscopy. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis combined with blotting technology indicated that the tetrasaccharide is mainly associated with one of the cellulosome subunits.

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Periodate oxidation of terminal N-acetyl- and N-glycoloylneuraminic acid residues in the mucins from edible bird nest substance and pig submandibular gland, respectively, can be carried out under conditions which exclusively give rise to the formation of the C-7 analogues of these sialic acids. In contrast, the C-8 compounds can be obtained in a maximum yield of about 40%. Under identical conditions, N-glycoloylneuraminic acid is oxidized about 1.

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N-linked carbohydrate chains of porcine thyroglobulin were released by the hydrazinolysis procedure. The resulting mixture of oligosaccharide-alditols was fractionated by high-voltage paper electrophoresis, the acidic fractions were further separated by high-performance liquid chromatography on Lichrosorb-NH2, and analyzed by 500-MHz 1H-NMR spectroscopy and, partially, by permethylation analysis. Of the acidic oligosaccharide-alditols, the following sulfated carbohydrate chains could be identified: NeuAc alpha 2----6Gal beta 1----4GlcNAc beta 1----2Man alpha 1----3[(SO3Na----3)Gal beta 1----4GlcNAc beta1----2-Mana alpha 1----6]Man beta 1----4GlcNAc beta 1----4[Fuc alpha 1----6]GlcNAc-ol and NeuAc alpha 2----6Gal beta 1----4(SO3Na----)0-1 GlcNAc beta 1----2-Man alpha 1----3[NeuAc alpha 2----6Gal beta 1----4(SO3Na----6)1-0GlcNAc beta 1----2Man alpha 1----6]Man beta 1----4GlcNAc beta 1----4[Fuc alpha 1----6]GlcNAc- ol.

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From the placenta of a human fetus with galactosialidosis, detected by prenatal diagnosis, sialyloligosaccharides were isolated by successively gel-permeation chromatography on Bio-Gel P-6, anion-exchange chromatography on Mono Q and high-performance liquid chromatography on Lichrosorb-NH2. 16 sialic acid-containing N-glycosidic N-acetyllactosamine type of structures were identified by sugar analysis and 500-MHz 1H-NMR spectroscopy. The fully sialylated oligosaccharides differ from each other in the type of branching (mono-, di-, tri-, tri'- and tetra-antennary) or sialic acid linkage types (alpha 2-3/alpha 2-6).

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The Cad antigen is a rare erythrocyte blood group antigen expressed on both sialoglycoprotein and ganglioside structures. It is related both serologically and biochemically to the Sda blood group antigen expressed on over 90% of Caucasian erythrocytes. We reported previously that Cad erythrocytes contain a novel ganglioside that binds Helix pomatia lectin and inhibits human anti-Sda antibody.

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Sialic acid-containing storage material was isolated from cultured human galactosialidosis fibroblasts, by a combination of gel filtration and anion-exchange chromatography on Mono Q. The obtained sialyloligosaccharides were analyzed by 500-MHz 1H-NMR spectroscopy in combination with sugar analysis and analytical HPLC. The storage material consisted of a series of completely sialylated N-acetyl-lactosamine type of structures having Man beta 1-4GlcNAc at the reducing terminus in common, similar to those recently reported for human sialidosis fibroblasts.

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