Sequence analysis of heparan sulphate and heparin oligosaccharides

The biological activity of heparan sulphate (HS) and heparin largely depend s on internal oligosaccharide sequences that pro vide specific binding site s for an extensive range of proteins. Identification of such structures is crucial for the complete understanding of glycosaminoglycan (GAG)-protein i nteractions. We describe here a simple method of sequence analysis relying on the specific tagging of the sugar reducing end by H-3 radiolabelling, th e combination of chemical scission and specific enzymic digestion to genera te intermediate fragments, and the analysis of the generated products by st rong-anion-exchange HPLC. We present full sequence data on microgram quanti ties of four unknown oligosaccharides (three MS-derived hexasaccharides and one heparin-derived octasaccharide) which illustrate the utility and relat ive simplicity of the technique. The results clearly show that it is also p ossible to read sequences of inhomogeneous preparations. Application of thi s technique to biologically active oligosaccharides should accelerate progr ess in the understanding of HS and heparin structure-function relationships and provide new insights into the primary structure of these polysaccharid es.