cDNA cloning and expression of a novel human UDP-N-acetyl-alpha-D-galactosamine. Polypeptide N-acetylgalactosaminyltransferase, GalNAc-t3.
The glycosylation of serine and threonine residues during mucin-type O-linked protein glycosylation is carried out by a family of UDP-GalNAc:polypeptide N-acetylgalactosaminyltransferases (GalNAc-transferase). Previously two members, GalNAc-T1 and -T2, have been isolated and the genes cloned and characterized. Here we report the cDNA cloning and expression of a novel GalNAc-transferase termed GalNAc-T3. The gene was isolated and cloned based on the identification of a GalNAc-transferase motif (61 amino acids) that is shared between GalNAc-T1 and -T2 as well as a homologous Caenorhabditis elegans gene. The cDNA sequence has a 633-amino acid coding region indicating a protein of 72.5 kDa with a type II domain structure. The overall amino acid sequence similarity with GalNAc-T1 and -T2 is approximately 45%; 12 cysteine residues that are shared between GalNAc-T1 and -T2 are also found in GalNAc-T3. GalNAc-T3 was expressed as a soluble protein without the hydrophobic transmembrane domain in insect cells using a Baculo-virus vector, and the expressed GalNAc-transferase activity showed substrate specificity different from that previously reported for GalNAc-T1 and -T2. Northern analysis of human organs revealed a very restricted expression pattern of GalNAc-T3.
Bennett
E P
EP
Faculty of Health Sciences, School of Dentistry, DK-2200 Copenhagen, Denmark.
Hassan
H
H
Clausen
H
H
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D015152:Blotting, Northern; D017173:Caenorhabditis elegans; D002460:Cell Line; D003001:Cloning, Molecular; D017931:DNA Primers; D018076:DNA, Complementary; D005260:Female; D015870:Gene Expression; D015723:Gene Library; D006801:Humans; D007527:Isoenzymes / Q000096:biosynthesis* / Q000737:chemistry / Q000235:genetics; D008297:Male; D008969:Molecular Sequence Data; D017350:N-Acetylgalactosaminyltransferases / Q000096:biosynthesis* / Q000737:chemistry / Q000235:genetics; D009928:Organ Specificity; D016133:Polymerase Chain Reaction; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry; D015183:Restriction Mapping; D017386:Sequence Homology, Amino Acid; D018411:Spodoptera; D014162:Transfection; D000097763:Polypeptide N-acetylgalactosaminyltransferase
D017931:DNA Primers; D018076:DNA, Complementary; D007527:Isoenzymes; D017350:N-Acetylgalactosaminyltransferases; D011994:Recombinant Proteins; D000097763:Polypeptide N-acetylgalactosaminyltransferase