Sulphate release from keratan sulphate and heparan sulphate by bovine N-acetylglucosamine-6-sulphate sulphatase.
The functions of sulphated monosaccharides within glycosaminoglycans (GAGs) and glycoproteins are being studied intensely, but progress is hindered by an inability to selectively desulphate glycoconjugates. We recently identified an N-acetylglucosamine-6-sulphate sulphatase (NG6SS) from bovine kidney that can remove sulphate from N-acetylglucosamine-6-sulphate (GlcNAc-6-SO4) within oligosaccharides and glycoproteins. However, the potential 'endosulphatase' activity of the NG6SS toward GAGs is not known. To test for this possibility, [3H]glucosamine-, [3H]galactose- and 35SO4-labelled keratan sulphate (KS) were separately prepared by metabolic radiolabelling of bovine cornea. NG6SS quantitatively removed sulphate from KS without release of sugar fragments. The enzyme had a Km of 4.7 mM toward free GlcNAc-6-SO4, but its Km for commercially available bovine corneal KS was found to be 9.1 microM. Analyses of both KS and heparan sulphate after treatment with NG6SS demonstrated significant loss of sulphate from GlcNAc-6-SO4 in both GAGs. These findings may be relevant for future studies aimed at defining the function(s) of GlcNAc-6-SO4 residues in GAGs and understanding the catabolism of GAGs, especially in regard to sulphatidoses, such as Sanfilippo D syndrome in humans, which involves a deficiency of NG6SS activity.
Shilatifard
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Department of Biochemistry and Molecular Biology, Oklahoma Center for Molecular Medicine, University of Oklahoma Health Sciences Center, Oklahoma City, 73104, USA.
Cummings
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D000818:Animals; D002240:Carbohydrate Sequence; D002417:Cattle; D002852:Chromatography, Ion Exchange; D002854:Chromatography, Paper; D006497:Heparan Sulfate / Q000378:metabolism*; D006868:Hydrolysis; D007632:Keratan Sulfate / Q000378:metabolism*; D008969:Molecular Sequence Data; D013429:Sulfatases / Q000378:metabolism*; D013431:Sulfates / Q000378:metabolism*
D006497:Heparan Sulfate; D007632:Keratan Sulfate; D013429:Sulfatases; D013431:Sulfates; C018719:N-acetylglucosamine-6-sulfatase