Polypeptide of N-acetylglucosamine-6-O-sulfotransferase and DNA encoding the same

Abstract

Apolypeptide of N-acetylglucosamine-6-O-sulfotransferase and a DNA encoding the peptide are provided. The polypeptide is (a) or (b) below: (a) a polypeptide consisting of an amino acid sequence represented by SEQ ID NO: 2; or (b) a polypeptide which comprises an amino acid sequence including substitution, deletion, insertion or transposition of one or few amino acids in the amino acid sequence of (a) and which has an enzymatic activity to transfer a sulfate group from a sulfate group donor to a hydroxyl group at 6 position of an N-acetylglucosamine residue located at a non-reducing end of an oligosaccharide represented by the formula I: GlcNAcβ1-3Galβ1-4GlcNAc  (I) wherein GlcNAc represents an N-acetylglucosamine residue, Gal represents a galactose residue, β1-3 represents a β1-3 glycosidic linkage, and β1-4 represents a β1-4 glycosidic linkage.

What is claimed is:  
     
         1 . An isolated polypeptide having an amino acid sequence shown as SEQ ID No. 2 or a variant thereof wherein the polypeptide has an enzymatic activity to transfer a sulfate group from a sulfate group donor to a hydroxyl group at 6 position of an N-acetylglucosamine residue located at a non-reducing end of an oligosaccharide represented by formula I:  
       GlcNAcβ11-3Galβ1-4GlcNAc  (I)  
       wherein GlcNAc represents an N-acetylglucosamine residue, Gal represents a galactose residue, β1-3 represents a β1-3 glycosidic linkage, and β1-4 represents a β1-4 glycosidic linkage.  
     
     
         2 . An isolated polypeptide having an amino acid sequence shown as SEQ ID No. 4 or a variant thereof wherein the polypeptide has an enzymatic activity to transfer a sulfate group from a sulfate group donor to a hydroxyl group at 6 position of an N-acetylglucosamine residue located at a non-reducing end of an oligosaccharide represented by formula I:  
       GlcNAcβ1-3Galβ1-4GlcNAc  (I)  
       wherein GlcNAc represents an N-acetylglucosamine residue, Gal represents a galactose residue, β1-3 represents a β1-3 glycosidic linkage, and β1-4 represents a β1-4 glycosidic linkage.  
     
     
         3 . An isolated polypeptide having the following properties: 
 (a) Action: a sulfate group is transferred from a sulfate group donor to a hydroxyl group at 6 position of an N-acetylglucosamine residue located at a non-reducing end of an oligosaccharide represented by the formula I:    GlcNAcβ1-3Galβ1-4GlcNAc  (I)    wherein GlcNAc represents an N-acetylglucosamine residue, Gal represents a galactose residue, β1-3 represents a β1-3 glycosidic linkage, and β1-4 represents a β1-4 glycosidic linkage.    (b) Substrate specificity: a sulfate group is not transferred to any of the substances selected from the group consisting of; chondroitin, chondroitin 4-sulfate, chondroitin 6-sulfate, dermatan sulfate, keratan sulfate, desulfated keratan sulfate, CDSNS-heparin, mucin from porcine stomach, mucin from bovine submaxillary gland, and an oligosaccharide represented by the formula II:    Galβ1-4 GlcNAcβ1-3Galβ1-4GlcNAc  (II)    wherein GlcNAc represents an N-acetylglucosamine residue, Gal represents a galactose residue, β1-3 represents a β1-3 glycosidic linkage, and β1-4 represents a β1-4 glycosidic linkage; and    (c) the N-terminal amino acid sequence comprises an amino acid sequence represented by amino acid numbers 1 to 48 in SEQ ID No: 2.    
     
     
         4 . A method of producing a sulfated sugar represented by the formula III:  
       (SO 4 -6)GlcNAc-R  (III)  wherein GlcNAc represents an N-acetylglucosamine residue; SO 4 -6 means that a hydroxyl group at position 6 is sulfated, and —R represents a hydrogen atom or a sugar residue bonded by a glycosidic linkage, said method comprising;    reacting the polypeptide described in  claim 1  with a sugar chain represented by the formula IV:    GlcNAc-R  (IV)    wherein GlcNAc represents an N-acetylglucosamine residue, and —R represents a hydrogen atom or a sugar residue bonded by a glycosidic linkage.    
     
     
         5 . An isolated antibody that reacts with 6-sulfated sialyl Lewis X but does not react with any of the substances selected from the group consisting of; 6′-sulfated sialyl Lewis X, 6,6-bis-sulfated sialyl Lewis X, 6-sulfated Lewis X and Lewis X.  
     
     
         6 . An isolated antibody that reacts with a sugar chain represented by formula V:  
       Galβ1-4(Fucα1-3)(SO 4 -6)GlcNAc  (V)  
     
     
         7 . An antibody that reacts with a sugar chain represented by formula VI:  
       NeuAcα2-3Galβ1-4(SO 4 -6)GlcNAc  (VI)  
     
     
         8 . A method of producing a sulfated sugar represented by the formula III:  
       (SO 4 -6)GlcNAc-R  (III)  wherein GIcNAc represents an N-acetylglucosamine residue; SO 4 -6 means that a hydroxyl group at position 6 is sulfated, and —R represents a hydrogen atom or a sugar residue bonded by a glycosidic linkage, said method comprising;    reacting the polypeptide described in  claim 2  with a sugar chain represented by the formula IV:    GlcNAc-R  (IV)    wherein GlcNAc represents an N-acetylglucosamine residue, and —R represents a hydrogen atom or a sugar residue bonded by a glycosidic linkage.    
     
     
         9 . A method of producing a sulfated sugar represented by the formula III:  
       (SO 4 -6)GlcNAc-R  (III)  wherein GlcNAc represents an N-acetylglucosamine residue; SO 4 -6 means that a hydroxyl group at position 6 is sulfated, and —R represents a hydrogen atom or a sugar residue bonded by a glycosidic linkage, said method comprising;    reacting the polypeptide described in  claim 3  with a sugar chain represented by the formula IV:    GlcNAc-R  (IV)    Wherein GlcNAc represents an N-acetylglucosamine residue, and —R represents a hydrogen atom or a sugar residue bonded by a glycosidic linkage.