| Abstract: | Although 2-O-sulfated L-iduronic acid (IdoA) residues have been known to occur in heparin, 2-O-sulfated D-glucuronic acid (GlcA) residues have been reported only recently (Bienkowski, M. J., and Conrad, H. E. (1985) J. Biol. Chem. 250, 356-365). Disaccharides prepared by cleavage of heparin and N-deacetylated chondroitin 6-sulfate with nitrous acid were used to demonstrate a new sulfatase that catalyzed the removal of the 2-O-sulfate substituents from GlcA but not IdoA residues. The deamination products were labeled by NaB3H4 reduction to give disaccharides from heparin and chondroitin sulfate which had reducing terminal 2,5-anhydro-D-mannitol (3H]AManR) and 2,5-anhydro-D-talitol (3H]ATalR) residues, respectively. IdoA(2-SO4)-3H]AManR(6-SO4) from heparin and GlcA(2-SO4)-3H]ATalR(6-SO4) from chondroitin sulfate were purified for use as substrates. GlcA(2-SO4)-3H]AManR(6-SO4) was prepared by epimerization of IdoA(2-SO4)-3H]AManR(6-SO4) with hydrazine at 100 degrees C. Lysosomal enzyme preparations from chick embryo chondrocytes and from two normal human fibroblast cell lines catalyzed the removal of the 2-O-SO4 substituent from the uronic acid residues of IdoA(2-SO4)-3H]AManR(6-SO4), GlcA(2-SO4)-3H] AManR(6-SO4), and GlcA(2-SO4)-3H]ATalR(6-SO4). In contrast, a lysosomal enzyme preparation from a human fibroblast cell line deficient in idurono-2-sulfatase (Hunter's-syndrome), which had no activity on the IdoA(2-SO4)-3H]AManR(6-SO4), converted GlcA(2-SO4)-3H]AManR(6-SO4) to a mixture of GlcA-3H] AManR(6-SO4) and 3H]AManR(6-SO4). This enzyme also converted GlcA(2-SO4)-3H]ATalR(6-SO4) to a mixture of GlcA-3H]ATalR(6-SO4) and 3H]ATalR(6-SO4). Digestion of both GlcA(2-SO4)-3H]AManR(6-SO4) and GlcA(2-SO4)-3H]ATalR(6-SO4) was inhibited by 35SO2-4 and was arrested at the monosulfated disaccharide stage by 1,4-saccharolactone. The glucurono-2-sulfatase exhibited a pH optimum of 4. The results indicate that there exists a separate sulfatase for the removal of sulfate substituents from C-2 of GlcA residues in glycosaminoglycans. |
