title: Cloning of a human UDP-N-acetyl-alpha-D-Galactosamine:polypeptide N-acetylgalactosaminyltransferase that complements other GalNAc-transferases in complete O-glycosylation of the MUC1 tandem repeat.
abstract: A fourth human UDP-GalNAc:polypeptide N-acetylgalactosaminyltransferase, designated GalNAc-T4, was cloned and expressed. The genomic organization of GalNAc-T4 is distinct from GalNAc-T1, -T2, and -T3, which contain multiple coding exons, in that the coding region is contained in a single exon. GalNAc-T4 was placed at human chromosome 12q21.3-q22 by in situ hybridization and linkage analysis. GalNAc-T4 expressed in Sf9 cells or in a stably transfected Chinese hamster ovary cell line exhibited a unique acceptor substrate specificity. GalNAc-T4 transferred GalNAc to two sites in the MUC1 tandem repeat sequence (Ser in GVTSA and Thr in PDTR) using a 24-mer glycopeptide with GalNAc residues attached at sites utilized by GalNAc-T1, -T2, and -T3 (TAPPAHGVTSAPDTRPAPGSTAPPA, GalNAc attachment sites underlined). Furthermore, GalNAc-T4 showed the best kinetic properties with an O-glycosylation site in the P-selectin glycoprotein ligand-1 molecule. Northern analysis of human organs revealed a wide expression pattern. Immunohistology with a monoclonal antibody showed the expected Golgi-like localization in salivary glands. A single base polymorphism, G1516A (Val to Ile), was identified (allele frequency 34%). The function of GalNAc-T4 complements other GalNAc-transferases in O-glycosylation of MUC1 showing that glycosylation of MUC1 is a highly ordered process and changes in the repertoire or topology of GalNAc-transferases will result in altered pattern of O-glycan attachments.
authors.affiliation: Faculty of Health Sciences, School of Dentistry, Copenhagen, Denmark.
authors.forename: E P
authors.initials: EP
authors.lastname: Bennett
authors.forename: H
authors.initials: H
authors.lastname: Hassan
authors.forename: U
authors.initials: U
authors.lastname: Mandel
authors.forename: E
authors.initials: E
authors.lastname: Mirgorodskaya
authors.forename: P
authors.initials: P
authors.lastname: Roepstorff
authors.forename: J
authors.initials: J
authors.lastname: Burchell
authors.forename: J
authors.initials: J
authors.lastname: Taylor-Papadimitriou
authors.forename: M A
authors.initials: MA
authors.lastname: Hollingsworth
authors.forename: G
authors.initials: G
authors.lastname: Merkx
authors.forename: A G
authors.initials: AG
authors.lastname: van Kessel
authors.forename: H
authors.initials: H
authors.lastname: Eiberg
authors.forename: R
authors.initials: R
authors.lastname: Steffensen
authors.forename: H
authors.initials: H
authors.lastname: Clausen
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D015152:Blotting, Northern; D016466:CHO Cells; D003001:Cloning, Molecular; D006224:Cricetinae; D018076:DNA, Complementary / Q000737:chemistry; D008040:Genetic Linkage; D006031:Glycosylation; D006801:Humans; D013058:Mass Spectrometry; D008969:Molecular Sequence Data; D018396:Mucin-1 / Q000378:metabolism*; D017350:N-Acetylgalactosaminyltransferases / Q000235:genetics* / Q000378:metabolism; D016415:Sequence Alignment; D018411:Spodoptera; D013363:Submandibular Gland / Q000201:enzymology; D013379:Substrate Specificity; D020080:Tandem Repeat Sequences; D013912:Threonine / Q000378:metabolism; D000097763:Polypeptide N-acetylgalactosaminyltransferase
chemical_list: D018076:DNA, Complementary; D018396:Mucin-1; D017350:N-Acetylgalactosaminyltransferases; D013912:Threonine; D000097763:Polypeptide N-acetylgalactosaminyltransferase