Glycobiology Advance Access originally published online on March 19, 2004
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Glycobiology vol 14 no 5 pp. 467-479, 2004
Glycobiology vol. 14 no. 5 © Oxford University Press 2004; all rights reserved.
Mapping critical biological motifs and biosynthetic pathways of heparan sulfate
Department of Biology, Massachusetts Institute of Technology, Cambridge, MA 02139, and Division of Molecular and Vascular Medicine, BIDMC, Harvard Medical School, Boston, MA 02215
Received on November 20, 2003; revised on December 29, 2003; accepted on January 11, 2004
Heparan sulfate (HS) interacts with numerous proteins at the cell surface and orchestrates myriad biological events. Unraveling the mechanisms of these events at the molecular level calls for the structural analysis of these negatively charged and highly heterogeneous biopolymers. However, HS is often available only in small quantities, and the task of structural analysis necessitates the use of ultra-sensitive methods, such as mass spectrometry. Sequence heterogeneity within HS chains required us to identify critical functional groups and their spacing to determine structure–function relationships for HS. We carried out structural analysis of HS isolated from wild type, 3-OST-1, 3-OST-3A, or 3-OST-5 sulfotransferase-transduced Chinese hamster ovary cells and also from various tissues. In the context of tissue-specific HS, the data allowed us to map the biosynthetic pathways responsible for the placement of critical groups. As a means of determining the distance between critical groups within a motif, we determined the spacing of the rare GlcNAc-GlcA disaccharide sequence in the completely desulfated re-N-sulfated porcine intestinal heparin. These disaccharides are biosynthetic regulatory markers for 3-OST-1 modification and the partial structure of the antithrombin III binding site. They occur only at the distance of hexasaccharide, octasaccharide, decasaccharide, or dodecasaccharide. Thus this approach allowed us to map both the biosynthetic pathways for generating critical functional groups and their spacing within HS. Our new strategy removes two obstacles to rapid progress in this field of research.
1 These authors contributed equally to this article.
2 To whom correspondence should be addressed; e-mail: rdrrosen{at}mit.edu
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