Abstract
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-GlcAbeta1--> 3Galbeta1-->4GlcNAc. 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 full-length cDNA could transfer sulfate from a sulfate donor to acceptor substrates containing terminal glucuronic acid.
Highlights
The carbohydrate antigen recognized by the monoclonal antibody HNK-1 was originally described as a marker for human natural killer cells [1]
A cDNA encoding the glucuronyltransferase involved in the biosynthesis of at least the HNK-1 glycoprotein epitope has recently been cloned [23]
A single clone isolated from the last pool gave, upon cotransfection, HNK-1positive cells at about the same frequency as seen when cells were transfected with glucuronyltransferase cDNA alone and stained with antibody L2– 412 (Fig. 1, D and E)
Summary
The carbohydrate antigen recognized by the monoclonal antibody HNK-1 was originally described as a marker for human natural killer cells [1]. 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 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.
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