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The HNK-1 carbohydrate epitope is expressed on several neural adhesion glycoproteins and as a glycolipid, and is involved in cell interactions. The structural element of the epitope common to glycoproteins and glycolipids has been determined to be sulfate-3-GlcA␤13 3Gal␤134GlcNAc. The glucuronyltransferase and sulfotransferase are considered to be the key enzymes in the biosynthesis of this epitope because the rest of the structure occurs often in glycoconjugates. Here we describe the isolation of the rat sulfotransferase cDNA via an expression cloning strategy. The clone finally isolated predicts a protein of 356 amino acids, with characteristics of a type II transmembrane protein and with no sequence similarity to other known sulfotransferases. Both the enzyme expressed as a soluble fusion protein and homogenates of cells transfected with the fulllength cDNA could transfer sulfate from a sulfate donor to acceptor substrates containing terminal glucuronic acid.The carbohydrate antigen recognized by the monoclonal antibody HNK-1 was originally described as a marker for human natural killer cells (1). Later it was shown to be expressed predominantly on glycolipids and glycoproteins from nervous tissue (2-5). The expression pattern of the HNK-1 carbohydrate in both the central and peripheral nervous system is spatially and developmentally regulated (6 -11). The HNK-1 carbohydrate epitope is carried by many, but not all, neural recognition glycoproteins and is involved in homo-and heterophilic binding of these proteins (for a review, see Ref. 12). Of special interest is the association of the epitope with Schwann cells myelinating motor but not sensory axons (10), where it may be involved in the preferential reinervation of muscle nerves by motor axons after lesion (13,14).Determination of the structure of the glycolipid (15) and glycoprotein (16) forms has shown that both carry sulfate-3-GlcA␤133Gal␤134GlcNAc at the nonreducing end. The minimal requirement for recognition by HNK-1 is unknown, but the antibody only binds to the sulfated form(17). Several other monoclonal antibodies have been isolated that recognize identical or similar structures(4, 18); of these, L2-412 is important for this study, because it also recognizes the non-sulfated form of the carbohydrate(19).The key enzymes in the biosynthesis of HNK-1 carbohydrates are a glucuronyltransferase (20, 21), transferring GlcA in ␤133 linkage to a terminal galactose, and a sulfotransferase (22), responsible for coupling sulfate to the C-3 position of this GlcA residue. A cDNA encoding the glucuronyltransferase involved in the biosynthesis of at least the HNK-1 glycoprotein epitope has recently been cloned (23). We describe here the cloning of a cDNA coding for a sulfotransferase active on terminal glucuronic acid residues and whose expression can render cells immunoreactive with HNK-1 antibody when cotransfected with a glucuronyltransferase cDNA. EXPERIMENTAL PROCEDURESCell Lines, Antibodies, and Plasmids-CHOP2 cells (24) were grown in minimal essential medi...

This work describes the first crystal structure ever reported of a histo-blood group carbohydrate antigen: Le(x). This study provides a detailed description of the conformation of two crystallographic independent molecules in a highly hydrated environment along with their hydrogen bonding properties and packing features. Some interactions observed between adjacent trisaccharides can provide the basis for involvement of Le(x)-Le(x) interactions in cell-cell adhesion.

SUMMARYNeutrophil recruitment into systemic inflammatory sites in viva is thought to be initiated by selectin-mediated endothelial adherence. The effect of fucoidan (natural sulfated polymer of L-fucose) on the selectin dependent PMN migration into rat peritoneum following the induction of inflammation by peptone injection was studied. Peritonitis was characterized by an increase in the total cell number (from 45.3 x 106to 91.6 x 106/rat), and by highly elevated PMN content (from 0.2% to 58%) in the rat peritoneal cavity 3 h after peptone injection. Intravenous administration of fucoidan was found to reduce, in a dose-dependent manner, neutrophil migration into peritoneum. Fucoidan in a dose as low as 0.8 mg per rat caused 96.8% reduction of neutrophil extravasation. The inhibitory effect of fucoidan was also dependent on the time intervals between the peptone and fucoidan injections. The maximal inhibitory effect of fucoidan was observed within the first 15 rain after the induction of peritonitis and it was ~maintained at a level of 80% during 1.5 h. Administration of fucoidan 2.5 h after peptone injection had practically no effect on PMN extravasation. Since P-selectin is known to play a key role at the earlier stages of PMN extravasation, it was suggested that the inhibitory effect of fucoidan was mostly due to its interaction with P-selectin. The in vitro experiments demonstrated the high affinity of fucoidan for both isolated P-selectin and P-selectin in plasma membranes of activated platelets.

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