Ursula Dabrowski

Structures of acidic O-linked polylactosaminoglycans on human skim milk mucins.

O-Linked glycans were isolated from human skim milk mucins or mucin-derived high-molecular weight glycopeptides and fractionated by anion exchange chromatography into neutral and acidic alditols. Major oligosaccharides contained in the acidic fraction were purified by high performance liquid chromatography and structurally characterized by a combination of fast atom bombardment mass spectrometry, methylation analysis and 500 MHz1H-nuclear magnetic resonance spectroscopy. The structural aspects exhibited by these major species in the acidic fraction resemble those established previously for the neutral oligosaccharides from human skim milk mucins:1)the size of the alditols varies from tri- to decasaccharides2)the core structure is of the ubiquitous type 23)the backbone sequences are of the poly-N-acetyllactosamine type with a particular preponderance of linearly extended GlcNAcβ(1–3)Gal (major) or GlcNAcβ(1–6)Gal units (minor).N-Acetylneuraminic acid on monosialylated (mucin- or glycopeptide-derived) and disialylated glycans (glycopeptide derived) is linked predominantly to position C-3 of galactose.

MS and NMR analysis of the cross-reacting determinant glycan from Trypanosoma brucei brucei MITat 1.6 variant specific glycoprotein

The cross-reacting determinant glycan from Trypanosoma brucei brucei MITat 1.6 is known to contain galactose, mannose and non-acetylated glucosamine. The structural elucidation of this oligosaccharide has been impeded by an unusual non-glycosidic linkage to the peptide chain and a glycosidic linkage to inositol phosphate on either side of the oligosaccharide. Using two different approaches for the isolation of the glycan, namely hydrolysis to give the oligosaccharide directly or pronase digestion to yield the glycan-containing C-terminal glycophosphopeptide, the structure of this glycan was elucidated by mass spectrometry and 1H-NMR spectroscopy. There was evidence of heterogeneity in the glycan residue.

A two-dimensional 1H-n.m.r. (500 MHz) and 13C-n.m.r. (125 MHz) study of N-linked glycopeptides derived from calf fetuin

The oligosaccharide part of an N-linked triantennary glycopeptide from calf fetuin with fourteen carbohydrate residues and its smaller derivatives obtained by successive enzymic cleavage of the terminal residues were investigated using 2D 1H-n.m.r. (500 MHz) and 13C-n.m.r. (125 MHz) spectroscopy. Assignments have been made of the resonances of almost all the protons of the constituent carbohydrate residues in these glycopeptides. A comparison of the 1H chemical shifts and coupling constants, as determined from the cross-peaks, has shown the dependence of these parameters on the interactions of spatially related neighbouring carbohydrates. Small conformational changes take place upon elongation of the oligosaccharide side-chains.

Lipopolysaccharide core region of Hafnia alvei: structure elucidation using chemical methods, gas chromatography−mass spectrometry, and NMR spectroscopy

Sugar and methylation analysis with the use of gas chromatography-mass spectrometry and 1H NMR spectroscopy proved that the core oligosaccharides isolated from lipopolysaccharides of eight Hafnia alvei strains have the identical hexasaccharide skeleton. However, 1H, 31P heterocorrelated spectra showed that the phosphorylation pattern is not the same. The branched heptose for the ATCC 13337, 1187, 2, 1191, 1196, 1220, and 481L strains is phosphorylated as in the following formula, where P = -O-P(O)(O-)2 and P-PEtN = [-O-P(O)(O-)]2-O(CH2)2NH3+ [formula: see text] A different phosphorylation pattern was found for the 1211 strain, where the branched heptose residue is 6-substituted by a monophosphorylethanolamine group, ...-->3(-->7)(PEtN-->6)-alpha-LD-Hepp-(1-->3)..., where PEtN = -O-P(O)(O-)-O(CH2)2NH3+.

The determination of phytosphingosine-containing globotriaosylceramide from human kidney in the presence of lactosylceramide

Globotriaosylceramide, the natural substrate of alpha-galactosidase A (the enzyme deficient in Fabrys disease) was prepared from human kidney by repeated medium pressure chromatography on Lichroprep Si 60 (E. Merck) before and after peracetylation. The apparently homogeneous preparation migrating as a single band on HPTLC was analysed by fast atom bombardment mass spectrometry and 1H-NMR at 500 MHz. It was found that in this fraction two major molecular species were comigrating: Gal alpha 1-4Gal beta 1-4Glc beta 1-1ceramide with nervonic and lignoceric acid linked to phytosphingosine and Gal beta 1-4Glc beta 1-1 ceramide with palmitic acid linked to sphingosine.

Structure of the O-specific polysaccharide, containing a glycerol phosphate substituent, of Hafnia alvei strain 1220 lipopolysaccharide

The O-specific polysaccharide of the lipopolysaccharide produced by Hafnia alvei strain 1220 contained D-glucose, D-galactose, N-acetyl-D-glucosamine, N-acetyl-L-fucosamine (2-acetamido-2,6-dideoxy-L-galactose), glycerol, and phosphate. It was proved by composition and methylation analyses, Smith degradation, dephosphorylation, and one- and two-dimensional 1H NMR spectroscopy to be a teichoic acid-like polymer with a branched hexasaccharide repeating unit of the following structure. [sequence: see text]