Substrate Specificities of Three Members of the Human UDP-N-Acetyl-α-d-galactosamine:Polypeptide N-Acetylgalactosaminyltransferase Family, GalNAc-T1, -T2, and -T3
Open accessAbstract
Mucin-type O-glycosylation is initiated by UDP-N-acetylgalactosamine:polypeptide N-acetylgalactosaminyltransferases (GalNAc-transferases). The role each GalNAc-transferase plays in O-glycosylation is unclear. In this report we characterized the specificity and kinetic properties of three purified recombinant GalNAc-transferases. GalNAc-T1, -T2, and -T3 were expressed as soluble proteins in insect cells and purified to near homogeneity. The enzymes have distinct but partly overlapping specificities with short peptide acceptor substrates. Peptides specifically utilized by GalNAc-T2 or -T3, or preferentially by GalNAc-T1 were identified. GalNAc-T1 and -T3 showed strict donor substrate specificities for UDP-GalNAc, whereas GalNAc-T2 also utilized UDP-Gal with one peptide acceptor substrate. Glycosylation of peptides based on MUC1 tandem repeat showed that three of five potential sites in the tandem repeat were glycosylated by all three enzymes when one or five repeat peptides were analyzed. However, analysis of enzyme kinetics by capillary electrophoresis and mass spectrometry demonstrated that the three enzymes react at different rates with individual sites in the MUC1 repeat. The results demonstrate that individual GalNAc-transferases have distinct activities and the initiation of O-glycosylation in a cell is regulated by a repertoire of GalNAc-transferases.
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UDP-GlcNAc is not a substrate for GalNAc-T2.
“As expected from previously published results [21], UDP-GlcNAc is not a substrate for GalNAc-T2.”
From High level in vivo mucin-type glycosylation in Escherichia coli (2018) · cites this paper as [21]
UDP-galactose has been postulated as an alternative sugar substrate for GalNAc-T2.
“Although UDP-galactose has been postulated as an alternative sugar substrate for GalNAc-T2 [21, 30], no sugar moieties attached to T7Muc10, which correlate with galactose residues were detected.”
From High level in vivo mucin-type glycosylation in Escherichia coli (2018) · cites this paper as [21]
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References 47 · Cited by 303
Research works printed but not linked: 6
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- Authors
- Wandall, H. H., Hassan, H., Mirgorodskaya, E., Kristensen, A. K., Roepstorff, P., Bennett, E. P., Nielsen, P. A., Hollingsworth, M. A., Burchell, J., Taylor-Papadimitriou, J., & Clausen, H.
- Affiliations
- University of Copenhagen (DK)
- University of Nebraska Medical Center (US)
- Corresponding author
- Clausen, Henrik
- ORCID
- Venue
- Journal of Biological Chemistry
- Publisher
- Elsevier BV
- Published
- 1997-09
- Volume
- 272
- Issue
- 38
- Pages
- 23503-23514
- ISSN
- 0021-9258 (ISSN-L)
- OpenAlex
- W2108168031
- Cited by
- 303
- References
- 47
- Citations by year
- 2026: 3 · 2025: 3 · 2024: 5 · 2023: 4 · 2022: 3 · 2021: 7 · 2020: 7 · 2019: 9 · 2018: 8 · 2017: 7 · 2016: 6 · 2015: 3 · 2014: 18 · 2013: 11 · 2012: 12
- FWCI
- 7.00
- Type
- journal article
- Language
- en
- Open access
- hybrid
- doi.org
- Licence
- Locations
- Journal of Biological Chemistry (publishedVersion, cc-by)
- PubMed (publishedVersion)
- Research at the University of Copenhagen (University of Copenhagen) (publishedVersion)
- Funders
- Velux Stiftung
- Kræftens Bekæmpelse
- Sundhed og Sygdom, Det Frie Forskningsråd
- Lundbeckfonden
- Novo Nordisk
- Novo Nordisk Fonden
- H. Lundbeck A/S
- National Institutes of Health (5r01ca066234-02)
- Medical Research Council
- National Cancer Institute (1RO1CA66234)
- Topics
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Glycosylation and Glycoproteins Research
- Physical Sciences › Chemistry › Organic Chemistry › Carbohydrate Chemistry and Synthesis
- Health Sciences › Medicine › Hematology › Blood groups and transfusion
- Keywords
- GalNAc-T3, N-acetylgalactosaminyltransferase, substrate specificity, mucin-type O-glycosylation, UDP-GalNAc, enzyme kinetics
- MeSH terms
- Amino Acid Sequence, Humans, Isoenzymes, Kinetics, Molecular Sequence Data, Peptides, Recombinant Proteins, Substrate Specificity, N-Acetylgalactosaminyltransferases
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