Preparation and structure of heparin lyase-derived heparan sulfate oligosaccharides

Ronald E. Hileman, April E. Smith, Toshihiko Toida, Robert J. Linhardt

Research output: Contribution to journalArticlepeer-review

53 Scopus citations

Abstract

Porcine intestinal mucosal heparan sulfate was exhaustively depolymerized on a large scale using heparin lyase II (heparinase II) or heparin lyase III (heparitinase, EC 4.2.2.8). The oligosaccharide mixtures formed with each enzyme were fractionated by low pressure gel permeation chromatography. Size-uniform mixtures of disaccharides, tetrasaccharides, and hexasaccharides were obtained. Each size-fractionated mixture was then purified on the basis of charge by repetitive semipreparative strong-anion-exchange high-performance liquid chromatography. This approach has led to the isolation of 13 homogenous oligosaccharides. The purity of each oligosaccharide was demonstrated by the presence of a single peak on analytical strong-anion-exchange high-performance liquid chromatography and reversed polarity capillary electrophoresis. The structures of these oligosaccharides were established using 500 MHz one- and two-dimensional nuclear magnetic resonance spectroscopy. Three of the thirteen structures that were solved were novel while the remaining 10 have been previously described. All of the structures obtained using heparin lyase In: contained a ΔUAp residue (where ΔUAp is 4-deoxy-α-L-threo-hex-4-eno-pyranosyluronic acid) at their nonreducing termini. Structures obtained using heparin lyase II contained both ΔUAp and ΔUAp2S (where S is sulfate) at their nonreducing termini. These results are consistent with the reported specificity of both enzymes.

Original languageEnglish
Pages (from-to)231-239
Number of pages9
JournalGlycobiology
Volume7
Issue number2
DOIs
StatePublished - Mar 1997
Externally publishedYes

Keywords

  • 1D and 2D NMR
  • Heparan sulfate
  • Heparin lyase
  • Heparinase
  • Sulfated oligosaccharides

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