Skip to main content

Advertisement

SpringerLink
Log in

Preparation and characterization of heparin hexasaccharide library with N-unsubstituted glucosamine residues

  • Original Article
  • Published:
Glycoconjugate Journal Aims and scope Submit manuscript

Abstract

The rare N-unsubstituted glucosamine (GlcNH3 +) residues in heparan sulfate (HS) have important biological and pathophysiological roles. Therefore, the ability to chemically generate a series of oligosaccharides, which have a similar structure to the naturally-occurring, GlcNH3 + - containing oligosaccharides from HS, would greatly contribute to investigating their natural role in HS. In this study, a hexasaccharide library that possess GlcNH3 + residues were prepared from the chemical modification of the fully sulfated dp6. Chemical reaction conditions were optimized to generate different pattern of GlcNH3 + - containing oligosaccharides, then the structure of the library was detected by high-performance liquid chromatography-ion trap/time-of-flight mass spectrometry (LC/MS-ITTOF) analysis. EIC/MS and MS2 analysis showed different fragmentation patterns of dp6s with different GlcNH3 + residues. This provides a foundation for further identification and quantification of GlcNH3 + - oligosaccharides by mass spectrum analysis.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5

Similar content being viewed by others

Abbreviations

CyPB:

Cyclophilin-B

DMSO:

Dimethyl sulfoxide

dp:

Degree of polymerization (corresponds to the number of monosaccharide units, i.e. dp2 is a disaccharide)

EIC:

Extracted ion chromatography

GAGs:

Glycosaminoglycans

GlcA:

β-D-glucuronic acid

GlcNAc:

β-D-N-acetylglucosamine

GlcNH3 + :

β-D-N-unsubstituted glucosamine

GlcNS:

β-D-N-sulfoglucosamine

HS 3-OST:

Heparan sulfate 3-O-sulfotransferases

HS:

Heparan sulfate

HSV:

Herpes simplex virus

IdoA:

α-L-iduronic acid

IPRP-LC-ITTOF:

Ion-pair reversed-phase liquid chromatography-ion trap/time-of-flight mass spectrometry

LMWH:

Low molecular weight heparin

NDST:

N-deacetylase/N-sulfotransferase

PTA:

n-pentylamine

RT:

Retention time

SAX-HPLC:

Strong anion-exchange HPLC

SE-HPLC:

Size-exclusion HPLC

References

  1. Couchman, J.R.: Syndecans: proteoglycan regulators of cell-surface microdomains? Nat. Rev. Mol. Cell Biol. 4, 926–937 (2003)

    Article  CAS  PubMed  Google Scholar 

  2. Liu, D., Shriver, Z., Venkataraman, G., El Shabrawi, Y., Sasisekharan, R.: Tumor cell surface heparan sulfate as cryptic promoters or inhibitors of tumor growth and metastasis. Proc. Natl. Acad. Sci. U. S. A. 99, 568–573 (2002)

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  3. Perrimon, N., Bernfield, M.: Specificities of heparan sulphate proteoglycans in developmental processes. Nature 404, 725–728 (2000)

    Article  CAS  PubMed  Google Scholar 

  4. Robinson, C.J., Stringer, S.E.: The splice variants of vascular endothelial growth factor (VEGF) and their receptors. J. Cell Sci. 114, 853–865 (2001)

    CAS  PubMed  Google Scholar 

  5. Kamimura, K., Koyama, T., Habuchi, H., Ueda, R., Masu, M., Kimata, K., Nakato, H.: Specific and flexible roles of heparan sulfate modifications in Drosophila FGF signaling. J. Cell Biol. 174, 773–778 (2006)

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  6. Kreuger, J., Spillmann, D., Li, J.P., Lindahl, U.: Interactions between heparan sulfate and proteins: the concept of specificity. J. Cell Biol. 174, 323–327 (2006)

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  7. Esko, J.D., Lindahl, U.: Molecular diversity of heparan sulfate. J. Clin. Invest. 108, 169–173 (2001)

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  8. Lindahl, U., Kusche-Gullberg, M., Kjellén, L.: Regulated diversity of heparan sulfate. J. Biol. Chem. 273, 24979–24982 (1998)

    Article  CAS  PubMed  Google Scholar 

  9. Esko, J.D., Selleck, S.B.: Order out of chaos: assembly of ligand binding sites in heparan sulfate. Annu. Rev. Biochem. 71, 435–471 (2002)

    Article  CAS  PubMed  Google Scholar 

  10. Casu, B., Lindahl, U.: Structure and biological interactions of heparin and heparan sulfate. Adv. Carbohydr. Chem. Biochem. 57, 159–206 (2001)

    Article  CAS  PubMed  Google Scholar 

  11. Cheng, F., Svensson, G., Fransson, L.-Å., Mani, K.: Non-conserved, S-nitrosylated cysteines in glypican-1 react with N-unsubstituted glucosamines in heparan sulfate and catalyze deaminative cleavage. Glycobiology 11, 1480–1486 (2012)

    Article  Google Scholar 

  12. Shukla, D., Liu, J., Blaiklock, P., Shworak, N.W., Bai, X., Esko, J.D., Cohen, G.H., Eisenberg, R.J., Rosenberg, R.D., Spear, P.G.: A novel role for 3-O-sulfated heparan sulfate in herpes simplex virus 1 entry. Cell 99, 13–22 (1999)

    Article  CAS  PubMed  Google Scholar 

  13. Liu, J., Shriver, Z., Pope, R.M., Thorp, S.C., Duncan, M.B., Copeland, R.J., Raska, C.S., Yoshida, K., Eisenberg, R.J., Cohen, G., Linhardt, R.J., Sasisekharan, R.: Characterization of a heparan sulfate octasaccharide that binds to herpes simplex virus type 1 glycoprotein D. J. Biol. Chem. 277, 33456–33467 (1997)

    Article  Google Scholar 

  14. Liu, J., Shriver, Z., Blaiklock, P., Yoshida, K., Sasisekharan, R., Rosenberg, R.D.: Heparan sulfate D-glucosaminyl 3-O-sulfotransferase-3A sulfates N-unsubstituted glucosamine residues. J. Biol. Chem. 274, 38155–38162 (1999)

    Article  CAS  PubMed  Google Scholar 

  15. Fu, L., Li, L., Cai, C., Li, G., Zhang, F., Linhardt, R.J.: Heparin stability by determining unsubstituted amino groups using HILIC-MS. Anal. Biochem. 461, 46–48 (2014)

    Article  CAS  PubMed  Google Scholar 

  16. Beaudet, J.M., Weyers, A., Solakyildirim, K., Yang, B., Takieddin, M., Mousa, S., Zhang, F., Linhardt, R.J.: Impact of autoclave sterilization on the activity and structure of formulated heparin. J. Pharm. Sci. 100, 3396–3404 (2011)

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  17. Nadanaka, S., Purunomo, E., Takeda, N., Tamura, J.I., Kitagawa, H.: Heparan sulfate containing unsubstituted glucosamine residues: biosynthesis and heparanase-inhibitory activity. J. Biol. Chem. 289, 15231–15243 (2014)

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  18. Shi, X., Zaia, J.: Organ-specific heparan sulfate structural phenotypes. J. Biol. Chem. 284, 11806–11814 (2009)

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  19. Wei, Z., Lyon, M., Gallagher, J.T.: Distinct substrate specificities of bacterial heparinases against N-unsubstituted glucosamine residues in heparan sulfate. J. Biol. Chem. 280, 15742–15748 (2005)

    Article  CAS  PubMed  Google Scholar 

  20. Rees, M.D., Pattison, D.I., Davies, M.J.: Oxidation of heparan sulphate by hypochlorite: role of N-chloro derivatives and dichloramine-dependent fragmentation. Biochem. J. 391, 125–134 (2005)

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  21. Westling, C., Lindahl, U.: Location of N-unsubstituted glucosamine residues in heparan sulfate. J. Biol. Chem. 277, 49247–49255 (2002)

    Article  CAS  PubMed  Google Scholar 

  22. Toida, T., Yoshida, H., Toyoda, H., Koshiishi, I., Imanari, T., Hileman, R.E., Fromm, J.R., Linhardt, R.J.: Structural differences and the presence of unsubstituted amino groups in heparan sulphate from different tissues and species. Biochem. J. 322, 499–506 (1997)

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  23. Wei, Z., Deakin, A.J., Blaum, B.S., Uhrín, D., Gallagher, J.T., Lyon, M.: Preparation of heparin / heparan sulfate oligosaccharides with internal N-unsubstituted glucosamine residues for functional studies. Glycoconj. J. 28, 525–535 (2011)

    Article  CAS  PubMed  Google Scholar 

  24. Liang, Q.T., Xiao, X.X., Lin, J.H., Wei, Z.: A new sequencing approach for N-unsubstituted heparin/heparan sulfate oligosaccharides. Glycobiology 25(7), 714–725 (2015)

    Article  PubMed  Google Scholar 

  25. Huang, R., Liu, J., Sharp, J.S.: An approach for separation and complete structural sequencing of heparin/heparan sulfate-like oligosaccharides. Anal. Chem. 85, 5787–5795 (2013)

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  26. Wolff, J.J., Chi, L., Linhardt, R.J., Amster, I.J.: Distinguishing glucuronic from iduronic acid in glycosaminoglycan tetrasaccharides by using electron detachment dissociation. Anal. Chem. 79, 2015–2022 (2007)

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  27. Schenauer, M.R., Meissen, J.K., Seo, Y.J., Ames, J.B., Leary, J.A.: Heparan sulfate separation, sequencing, and isomeric differentiation: ion mobility spectrometry reveals specific iduronic and glucuronic scid-containing hexasaccharides. Anal. Chem. 81, 10179–10185 (2009)

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  28. Shi, X., Huang, Y., Mao, Y., Naimy, H., Zaia, J.: Tandem mass spectrometry of heparan sulfate negative ions: sulfate loss patterns and chemical modification methods for improvement of product ion profiles. J. Am. Soc. Mass Spectrom. 23, 1498–1511 (2012)

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  29. Hu, H., Huang, Y., Mao, Y., Yu, X., Xu, Y., Liu, J., Zong, C., Boons, G.J., Lin, C., Xia, Y., Zaia, J.: A computational framework for heparan sulfate sequencing using high-resolution tandem mass spectra. Mol. Cell. Proteomics 13, 2490–2502 (2014)

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  30. Domon, B., Costello, C.E.: A systematic nomenclature for carbohydrate fragmentations in FAB-MS/MS spectra of glycoconjugates. Glycoconj. J. 5, 397–409 (1988)

    Article  CAS  Google Scholar 

  31. Liu, J., Shriver, Z., Pope, M., Thorp, S.C., Duncan, M.B., Copeland, R.J., Sasisekharan, R.: Characterization of a heparan sulfate octasaccharide that binds to herpes simplex virus type 1 glycoprotein D. J. Biol. Chem. 277, 33456–33467 (2002)

    Article  CAS  PubMed  Google Scholar 

  32. Vanpouille, C., Deligny, A., Delehedde, M., Denys, A., Melchior, A., Liénard, X., Lyon, M., Mazurier, J., Fernig, D.G., Allain, F.: The heparin/heparan sulfate sequence that interacts with cyclophilin B contains a 3-O-sulfated N-unsubstituted glucosamine residue. J. Biol. Chem. 282, 24416–24429 (2007)

    Article  CAS  PubMed  Google Scholar 

  33. Doneanu, C.E., Chen, W.B., Gebler, J.C.: Analysis of oligosaccharides derived from heparin by ion-pair reversed-phase chromatography/mass spectrometry. Anal. Chem. 81, 3485–3499 (2009)

    Article  CAS  PubMed  Google Scholar 

  34. Zaia, J., Costello, C.E.: Tandem mass spectrometry of sulfated heparin-like glycosaminoglycan oligosaccharides. Anal. Chem. 75, 2445–2455 (2003)

    Article  CAS  PubMed  Google Scholar 

Download references

Acknowledgments

This work was supported by Grants 20773023 and 21343013 from National Natural Science Foundation of China.

Author information

Authors and Affiliations

  1. Institute of Glycobiochemistry, National Engineering Research Centre of Chemical Fertilizer Catalyst, Fu Zhou University, Fu Zhou, 350002, People’s Republic of China

    Qun Tao Liang, Jia Yan Du, Qing Fu, Jiang Hui Lin & Zheng Wei

Authors
  1. Qun Tao Liang
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Jia Yan Du
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Qing Fu
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Jiang Hui Lin
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Zheng Wei
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Zheng Wei.

Electronic supplementary material

Below is the link to the electronic supplementary material.

ESM 1

(PDF 117 kb)

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Liang, Q.T., Du, J.Y., Fu, Q. et al. Preparation and characterization of heparin hexasaccharide library with N-unsubstituted glucosamine residues. Glycoconj J 32, 643–653 (2015). https://doi.org/10.1007/s10719-015-9612-8

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s10719-015-9612-8

Keywords

Search

Navigation