Heparinase 1 selectivity for the 3,6-di-O-sulfo-2-deoxy-2-sulfamido-a-D- glucopyranose (1,4) 2-O-sulfo-a-Lidopyranosyluronic acid (GlcNS3S6S-IdoA2S) linkages

Zhongping Xiao, Wenjing Zhao, Bo Yang, Zhenqing Zhang, Huashi Guan, Robert J. Linhardt

Research output: Contribution to journalArticlepeer-review

38 Scopus citations

Abstract

Porcine intestinal mucosa heparin was partially depolymerized by recombinant heparinase 1 (heparin lyase 1, originating from Flavobacterium heparinum and expressed in Escherichia coli) and then fractionated, leading to the isolation of 22 homogeneous oligosaccharides with sizes ranging from disaccharide to hexadecasaccharide. The purity of these oligosaccharides was determined by gel electrophoresis, strong anion exchange and reversed-phase ionpairing high-performance liquid chromatography. The molecular mass of oligosaccharides was determined using electrospray ionization-mass spectrometry and their structures were elucidated using one- and two-dimensional nuclear magnetic resonance spectroscopy at 600 MHz. Five of the characterized oligosaccharides represent new compounds. The most prominent oligosaccharide comprises the common repeating unit of heparin, ?UA2S-[-GlcNS6SIdoA2S-] n-GlcNS6S, where ?UA is 4-deoxy-a-L-threo-hex-4-eno-pyranosyluronic acid, GlcN is 2-deoxy-2-amino-Dglucopyranose, IdoA is L-idopyranosyluronic acid, S is sulfate and n = 0-7. A second prominent heparin oligosaccharide motif corresponds to?UA2S-[GlcNS6S-IdoA2S]n- GlcNS6S-IdoA-GlcNAc6S-GlcA-GlcNS3S6S (where n = 0-5 and GlcA is D-glucopyranosyluronic acid), a fragment of the antithrombin III binding site in heparin. The prominence of this second set of oligosaccharides and the absence of intact antithrombin III binding sites suggest that the -GlcNS3S6S-IdoA2S- linkage is particularly susceptible toheparinase 1.

Original languageEnglish
Pages (from-to)13-22
Number of pages10
JournalGlycobiology
Volume21
Issue number1
DOIs
StatePublished - Jan 2011
Externally publishedYes

Keywords

  • Antithrombin III
  • Enzyme specificity
  • Heparin
  • Heparin lyase
  • Oligosaccharide structure

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