Cloning of a human UDP-N-acetyl-α-D-galactosamine:Polypeptide N- acetylgalactosaminyltransferase that complements other GalNAc-transferases in complete O-glycosylation of the MUC1 tandem repeat

Eric Paul Bennett, Helle Hassan, Ulla Mandel, Ekatarina Mirgorodskaya, Peter Roepstorff, Joy Burchell, Joyce Taylor-Papadimitriou, Michael A. Hollingsworth, Gerard Merkx, Ad Geurts Van Kessel, Hans Eiberg, Rudi Steffensen, Henrik Clausen

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Abstract

A fourth human UDP-GalNAc:polypeptide N-acetyl- galactosaminyltransferase, 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.

Original languageEnglish (US)
Pages (from-to)30472-30481
Number of pages10
JournalJournal of Biological Chemistry
Volume273
Issue number46
DOIs
StatePublished - Nov 13 1998

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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    Bennett, E. P., Hassan, H., Mandel, U., Mirgorodskaya, E., Roepstorff, P., Burchell, J., Taylor-Papadimitriou, J., Hollingsworth, M. A., Merkx, G., Van Kessel, A. G., Eiberg, H., Steffensen, R., & Clausen, H. (1998). Cloning of a human UDP-N-acetyl-α-D-galactosamine:Polypeptide N- acetylgalactosaminyltransferase that complements other GalNAc-transferases in complete O-glycosylation of the MUC1 tandem repeat. Journal of Biological Chemistry, 273(46), 30472-30481. https://doi.org/10.1074/jbc.273.46.30472