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Very-high-field n.m.r. studies of bovine lung heparan sulphate oligosaccharides produced by nitrous acid deaminative cleavage. 13C-n.m.r. study of methylene resonances: degree and positions of C-6 sulphation.

ABSTRACT: Oligosaccharides with the general structure UA-(GlcNAc-GlcUA-)m-aManOH (m = 1-5) (where UA represents uronic acid, GlcNAc N-acetylglucosamine, GlcUA glucuronic acid and aManOH anhydromannitol) were prepared from low-sulphated heparan sulphates of bovine lung origin by nitrous acid deaminative cleavage followed by reduction. Analysis of the methylene signals in the 100 MHz 13C-n.m.r. spectrum of the tetrasaccharide (m = 1) shows that, whereas the extent of C-6 O-sulphation in the GlcNAc is approx. 65%, in the aManOH [formerly a GlcNSO3 (N-sulphoglucosamine) residue in the parent heparan sulphate] it is only approx. 10%. In the higher oligosaccharides (m = 2-5) the gross extent of C-6 O-sulphation of GlcNAc residues falls systematically with increasing oligosaccharide size, whereas that in the aManOH residues remains below 10%. There is also evidence that the C-6 O-sulphation of the GlcNAc residues is confined to the GlcNAc residue adjacent to the non-reducing terminal uronic acid residue. It is therefore tentatively proposed that the GlcNAc in the sequence -GlcNSO3-UA-GlcNAc- might be a favoured substrate for the 6-O-sulphotransferase. It is concluded that in the low-sulphated heparan sulphates GlcNSO3 residues that do not occur in (GlcNSO3-UA-)n blocks tend to have a significantly smaller extent of C-6 O-sulphation than do GlcNAc residues that occur in -GlcNSO3-UA-GlcNAc-GlcUA-GlcNSO3-sequences.

SUBMITTER: Sanderson PN

PROVIDER: S-EPMC1154413 | biostudies-other | 1983 Jun

REPOSITORIES: biostudies-other

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Very-high-field n.m.r. studies of bovine lung heparan sulphate tetrasaccharides produced by nitrous acid deaminative cleavage. Determination of saccharide sequence, uronate composition and degrees of sulphation.

Project description:Tetrasaccharides with the general structure UA-GlcNAc-GlcUA-aManOH (where UA represents uronate, GlcNAc N-acetylglucosamine, GlcUA glucuronate and aManOH anhydromannitol) were prepared from low-sulphated heparan sulphates of bovine lung origin by complete nitrous acid deaminative cleavage followed by reduction and fractionated by gel filtration. Ion-exchange chromatography of the tetrasaccharides yielded three major fractions in approximate yields of 37%, 45% and 14%. These were shown to be non-, mono- and di-sulphated respectively. Complete structural characterization of the tetrasaccharide fractions by quantitative high-field n.m.r. spectroscopy showed that each fraction contained only two discrete species and led to the following observations. (1) All of the uronate residues in the tetrasaccharides (and in larger oligosaccharides) are unsulphated, and hence sulphated iduronate [IdUA(2SO3)] must occur exclusively within -GlcNSO3-IdUA(2SO3)-GlcNSO3- sequences (where GlcNSO3 represents N-sulpho-glucosamine) in the parent polymers. (2) The GlcNAc residues in the tetrasaccharides are more highly C-6-O-sulphated than are the aManOH residues, and furthermore sulphation on the aManOH appears to occur only where the GlcNAc is also sulphated. (3) Where the GlcNAc is C-6-O-sulphated, iduronate is the major non-reducing terminal residue, whereas glucuronate predominates in this position if the GlcNAc is unsulphated. The quantitative data obtained are used to determine the degree of C-6-O-sulphation of glucosamine residues in specific sequences within the parent heparan sulphates.

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