Characterization of novel sequences containing 3-O-sulfated glucosamine in glomerular basement membrane heparan sulfate and localization of sulfated disaccharides to a peripheral domain.
Fragmentation of the heparan sulfate chains from bovine glomerular basement membrane (GBM) by hydrazine/nitrous acid treatment followed by NaB3H4-reduction yielded a mixture of six sulfated disaccharides containing D-glucuronic (GlcUA) or L-iduronic acid (IdUA) and terminating in 2,5-anhydro[3H]mannitol (AnManH2), in addition to the nonsulfated component GlcUA beta 1----4AnManH2. Among these products two novel disaccharide units were identified as IdUA alpha 1----4AnManH2(3-SO4) and IdUA(2-SO4)alpha 1----4AnManH2(3-SO4); these accounted for 22% of the total sulfated species indicating that there are 2-3 residues of 3-O-sulfated glucosamine/heparan sulfate chain. The disulfated disaccharide was shown through its release by direct nitrous acid treatment to be situated in a GlcNSO3-IdUA(2-SO4)-GlcNSO3(3-SO4) sequence which is distinct from that in which 3-O-sulfated glucosamine is located in the antithrombin-binding region of heparins. Analyses of heparan sulfate from lens capsule, a nonvascular basement membrane, indicated the absence of sequences containing 3-O-sulfated glucosamine, although otherwise the sulfated disaccharides produced by hydrazine/nitrous acid/Na-B3H4 treatment (GlcUA beta 1----4AnManH2(6-SO4), IdUA alpha 1----4AnManH2(6-SO4), IdUA(2-SO4)alpha 1----4AnManH2 and IdUA(2-SO4)alpha 1----4AnManH2(6-SO4] were the same as from GBM. Examination of the GBM heparan sulfate domains after nitrous acid treatment indicated that the O- as well as N-sulfate groups are clustered in an iduronic acid-rich 10-disaccharide peripheral segment, while the internal region (approximately 20 disaccharides) is composed primarily of repeating GlcUA beta 1----4GlcNAc units. The localization of chain diversity to the outer region may facilitate interactions of the heparan sulfate with other macromolecular components.
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Department of Biological Chemistry, Harvard Medical School, Boston, Massachusetts.
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D000818:Animals; D001485:Basement Membrane / Q000032:analysis; D002236:Carbohydrate Conformation; D002240:Carbohydrate Sequence; D002417:Cattle; D002848:Chromatography, DEAE-Cellulose; D002851:Chromatography, High Pressure Liquid; D002855:Chromatography, Thin Layer; D004187:Disaccharides / Q000302:isolation & purification*; D005944:Glucosamine / Q000031:analogs & derivatives* / Q000032:analysis; D006025:Glycosaminoglycans / Q000302:isolation & purification; D006497:Heparan Sulfate / Q000302:isolation & purification; D007908:Lens, Crystalline / Q000032:analysis; D008969:Molecular Sequence Data
D004187:Disaccharides; D005944:Glucosamine; D006025:Glycosaminoglycans; D006497:Heparan Sulfate; C065632:glucosamine 3-O-sulfate