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. 1999 May 1;339(Pt 3):767–773.

Sequence analysis of heparan sulphate and heparin oligosaccharides.

R R Vivès 1, D A Pye 1, M Salmivirta 1, J J Hopwood 1, U Lindahl 1, J T Gallagher 1
PMCID: PMC1220215  PMID: 10215618

Abstract

The biological activity of heparan sulphate (HS) and heparin largely depends on internal oligosaccharide sequences that provide specific binding sites for an extensive range of proteins. Identification of such structures is crucial for the complete understanding of glycosaminoglycan (GAG)-protein interactions. We describe here a simple method of sequence analysis relying on the specific tagging of the sugar reducing end by 3H radiolabelling, the combination of chemical scission and specific enzymic digestion to generate intermediate fragments, and the analysis of the generated products by strong-anion-exchange HPLC. We present full sequence data on microgram quantities of four unknown oligosaccharides (three HS-derived hexasaccharides and one heparin-derived octasaccharide) which illustrate the utility and relative simplicity of the technique. The results clearly show that it is also possible to read sequences of inhomogeneous preparations. Application of this technique to biologically active oligosaccharides should accelerate progress in the understanding of HS and heparin structure-function relationships and provide new insights into the primary structure of these polysaccharides.

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Selected References

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  1. Bielicki J., Hopwood J. J., Wilson P. J., Anson D. S. Recombinant human iduronate-2-sulphatase: correction of mucopolysaccharidosis-type II fibroblasts and characterization of the purified enzyme. Biochem J. 1993 Jan 1;289(Pt 1):241–246. doi: 10.1042/bj2890241. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Bourin M. C., Lindahl U. Glycosaminoglycans and the regulation of blood coagulation. Biochem J. 1993 Jan 15;289(Pt 2):313–330. doi: 10.1042/bj2890313. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Chai W., Hounsell E. F., Bauer C. J., Lawson A. M. Characterisation by LSI-MS and 1H NMR spectroscopy of tetra-, hexa-, and octa-saccharides of porcine intestinal heparin. Carbohydr Res. 1995 Apr 3;269(1):139–156. doi: 10.1016/0008-6215(94)00349-k. [DOI] [PubMed] [Google Scholar]
  4. Gallagher J. T., Lyon M., Steward W. P. Structure and function of heparan sulphate proteoglycans. Biochem J. 1986 Jun 1;236(2):313–325. doi: 10.1042/bj2360313. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Gallagher J. T., Turnbull J. E., Lyon M. Patterns of sulphation in heparan sulphate: polymorphism based on a common structural theme. Int J Biochem. 1992 Apr;24(4):553–560. doi: 10.1016/0020-711x(92)90326-v. [DOI] [PubMed] [Google Scholar]
  6. Guimond S., Maccarana M., Olwin B. B., Lindahl U., Rapraeger A. C. Activating and inhibitory heparin sequences for FGF-2 (basic FGF). Distinct requirements for FGF-1, FGF-2, and FGF-4. J Biol Chem. 1993 Nov 15;268(32):23906–23914. [PubMed] [Google Scholar]
  7. Ishihara M. Structural requirements in heparin for binding and activation of FGF-1 and FGF-4 are different from that for FGF-2. Glycobiology. 1994 Dec;4(6):817–824. doi: 10.1093/glycob/4.6.817. [DOI] [PubMed] [Google Scholar]
  8. Kjellén L., Lindahl U. Proteoglycans: structures and interactions. Annu Rev Biochem. 1991;60:443–475. doi: 10.1146/annurev.bi.60.070191.002303. [DOI] [PubMed] [Google Scholar]
  9. Lindahl U., Kusche-Gullberg M., Kjellén L. Regulated diversity of heparan sulfate. J Biol Chem. 1998 Sep 25;273(39):24979–24982. doi: 10.1074/jbc.273.39.24979. [DOI] [PubMed] [Google Scholar]
  10. Litjens T., Bielicki J., Anson D. S., Friderici K., Jones M. Z., Hopwood J. J. Expression, purification and characterization of recombinant caprine N-acetylglucosamine-6-sulphatase. Biochem J. 1997 Oct 1;327(Pt 1):89–94. doi: 10.1042/bj3270089. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Ludwigs U., Elgavish A., Esko J. D., Meezan E., Rodén L. Reaction of unsaturated uronic acid residues with mercuric salts. Cleavage of the hyaluronic acid disaccharide 2-acetamido-2-deoxy-3-O-(beta-D-gluco-4-enepyranosyluronic acid)-D-glucose. Biochem J. 1987 Aug 1;245(3):795–804. doi: 10.1042/bj2450795. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Lyon M., Deakin J. A., Gallagher J. T. Liver heparan sulfate structure. A novel molecular design. J Biol Chem. 1994 Apr 15;269(15):11208–11215. [PubMed] [Google Scholar]
  13. Lyon M., Deakin J. A., Mizuno K., Nakamura T., Gallagher J. T. Interaction of hepatocyte growth factor with heparan sulfate. Elucidation of the major heparan sulfate structural determinants. J Biol Chem. 1994 Apr 15;269(15):11216–11223. [PubMed] [Google Scholar]
  14. Maccarana M., Casu B., Lindahl U. Minimal sequence in heparin/heparan sulfate required for binding of basic fibroblast growth factor. J Biol Chem. 1993 Nov 15;268(32):23898–23905. [PubMed] [Google Scholar]
  15. Maccarana M., Sakura Y., Tawada A., Yoshida K., Lindahl U. Domain structure of heparan sulfates from bovine organs. J Biol Chem. 1996 Jul 26;271(30):17804–17810. doi: 10.1074/jbc.271.30.17804. [DOI] [PubMed] [Google Scholar]
  16. Pye D. A., Vives R. R., Turnbull J. E., Hyde P., Gallagher J. T. Heparan sulfate oligosaccharides require 6-O-sulfation for promotion of basic fibroblast growth factor mitogenic activity. J Biol Chem. 1998 Sep 4;273(36):22936–22942. doi: 10.1074/jbc.273.36.22936. [DOI] [PubMed] [Google Scholar]
  17. Rhomberg A. J., Ernst S., Sasisekharan R., Biemann K. Mass spectrometric and capillary electrophoretic investigation of the enzymatic degradation of heparin-like glycosaminoglycans. Proc Natl Acad Sci U S A. 1998 Apr 14;95(8):4176–4181. doi: 10.1073/pnas.95.8.4176. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Salmivirta M., Lidholt K., Lindahl U. Heparan sulfate: a piece of information. FASEB J. 1996 Sep;10(11):1270–1279. doi: 10.1096/fasebj.10.11.8836040. [DOI] [PubMed] [Google Scholar]
  19. Shively J. E., Conrad H. E. Formation of anhydrosugars in the chemical depolymerization of heparin. Biochemistry. 1976 Sep 7;15(18):3932–3942. doi: 10.1021/bi00663a005. [DOI] [PubMed] [Google Scholar]
  20. Tessler S., Rockwell P., Hicklin D., Cohen T., Levi B. Z., Witte L., Lemischka I. R., Neufeld G. Heparin modulates the interaction of VEGF165 with soluble and cell associated flk-1 receptors. J Biol Chem. 1994 Apr 29;269(17):12456–12461. [PubMed] [Google Scholar]
  21. Turnbull J. E., Fernig D. G., Ke Y., Wilkinson M. C., Gallagher J. T. Identification of the basic fibroblast growth factor binding sequence in fibroblast heparan sulfate. J Biol Chem. 1992 May 25;267(15):10337–10341. [PubMed] [Google Scholar]
  22. Turnbull J. E., Gallagher J. T. Distribution of iduronate 2-sulphate residues in heparan sulphate. Evidence for an ordered polymeric structure. Biochem J. 1991 Feb 1;273(Pt 3):553–559. doi: 10.1042/bj2730553. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Unger E. G., Durrant J., Anson D. S., Hopwood J. J. Recombinant alpha-L-iduronidase: characterization of the purified enzyme and correction of mucopolysaccharidosis type I fibroblasts. Biochem J. 1994 Nov 15;304(Pt 1):43–49. doi: 10.1042/bj3040043. [DOI] [PMC free article] [PubMed] [Google Scholar]

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