Abstract
Heparan sulfate proteoglycans are important modulators of cellular processes where the negatively charged polysaccharide chains interact with target proteins. The sulfation pattern of the heparan sulfate chains will determine the proteins that will bind and the affinity of the interactions. The N-deacetylase/N-sulfotransferase (NDST) enzymes are of key importance during heparan sulfate biosynthesis when the sulfation pattern is determined. In this chapter, metabolic labeling of heparan sulfate with [35S]sulfate or [3H]glucosamine in cell cultures is described, in addition to characterization of polysaccharide chain length and degree of N-sulfation. Methods to measure NDST enzyme activity are also presented.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
Similar content being viewed by others
References
Bishop JR, Schuksz M, Esko JD (2007) Heparan sulphate proteoglycans fine-tune mammalian physiology. Nature 446:1030–1037
Kreuger J, Kjellen L (2012) Heparan sulfate biosynthesis: regulation and variability. J Histochem Cytochem 60:898–907
Li F, Shi W, Capurro M, Filmus J (2011) Glypican-5 stimulates rhabdomyosarcoma cell proliferation by activating Hedgehog signaling. J Cell Biol 192:691–704
Ledin J, Staatz W, Li JP, Gotte M, Selleck S, Kjellen L et al (2004) Heparan sulfate structure in mice with genetically modified heparan sulfate production. J Biol Chem 279:42732–42741
Carlsson P, Presto J, Spillmann D, Lindahl U, Kjellen L (2008) Heparin/heparan sulfate biosynthesis: processive formation of N-sulfated domains. J Biol Chem 283:20008–20014
Kreuger J, Spillmann D, Li JP, Lindahl U (2006) Interactions between heparan sulfate and proteins: the concept of specificity. J Cell Biol 174:323–327
Humphries DE, Silbert CK, Silbert JE (1986) Glycosaminoglycan production by bovine aortic endothelial cells cultured in sulfate-depleted medium. J Biol Chem 261:9122–9127
Pikas DS, Eriksson I, Kjellen L (2000) Overexpression of different isoforms of glucosaminyl N-deacetylase/N-sulfotransferase results in distinct heparan sulfate N-sulfation patterns. Biochemistry 39:4552–4558
Ledin J, Ringvall M, Thuveson M, Eriksson I, Wilen M, Kusche-Gullberg M et al (2006) Enzymatically active N-deacetylase/N-sulfotrans-ferase-2 is present in liver but does not contribute to heparan sulfate N-sulfation. J Biol Chem 281:35727–35734
van den Born J, Pikas DS, Pisa BJ, Eriksson I, Kjellen L, Berden JH (2003) Antibody-based assay for N-deacetylase activity of heparan sulfate/heparin N-deacetylase/N-sulfotransferase (NDST): novel characteristics of NDST-1 and -2. Glycobiology 13:1–10
Shively JE, Conrad HE (1976) Formation of anhydrosugars in the chemical depolymerization of heparin. Biochemistry 15:3932–3942
Author information
Authors and Affiliations
Corresponding author
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2015 Springer Science+Business Media New York
About this protocol
Cite this protocol
Dagälv, A., Lundequist, A., Filipek-Górniok, B., Dierker, T., Eriksson, I., Kjellén, L. (2015). Heparan Sulfate Structure: Methods to Study N-Sulfation and NDST Action. In: Balagurunathan, K., Nakato, H., Desai, U. (eds) Glycosaminoglycans. Methods in Molecular Biology, vol 1229. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-1714-3_17
Download citation
DOI: https://doi.org/10.1007/978-1-4939-1714-3_17
Published:
Publisher Name: Humana Press, New York, NY
Print ISBN: 978-1-4939-1713-6
Online ISBN: 978-1-4939-1714-3
eBook Packages: Springer Protocols