Relative Susceptibilities of the Glucosamine−Glucuronic Acid and N-Acetylglucosamine−Glucuronic Acid Linkages to Heparin Lyase III†
Identifieur interne : 003085 ( Main/Exploration ); précédent : 003084; suivant : 003086Relative Susceptibilities of the Glucosamine−Glucuronic Acid and N-Acetylglucosamine−Glucuronic Acid Linkages to Heparin Lyase III†
Auteurs : Wengang Chai [Suède, Royaume-Uni] ; Christine Leteux [Suède] ; Camilla Westling [Suède] ; Ulf Lindahl [Suède] ; Ten Feizi [Suède]Source :
- Biochemistry [ 0006-2960 ] ; 2004.
Abstract
Heparin lyases are valuable tools for generating oligosaccharide fragments and in sequence determination of heparan sulfate (HS). Heparin lyase III is known to cleave the linkages between N-acetylglucosamine (GlcNAc) or N-sulfated glucosamine (GlcNS) and glucuronic acid (GlcA) as the primary sites and the linkages between GlcNAc, GlcNAc(6S), or GlcNS and iduronic acid as secondary sites. N-Unsubstituted glucosamine (GlcN) occurs as a minor component in HS, and it has been associated with various bioactivities. Here we investigate the specificity of heparin lyase III toward the GlcN−GlcA linkage using a recombinant enzyme of high purity and as substrates the partially de-N-acetylated polysaccharide of Escherichia coli K5 strain and derived hexasaccharides. The specificity of lyase III toward the GlcN−GlcA linkage is deduced by sequencing of the oligosaccharide products using electrospray mass spectrometry with collision-induced dissociation and MS/MS scanning. The results demonstrate that under controlled conditions for partial digestion, lyase III does not act at the GlcN−GlcA linkage, whereas GlcNAc−GlcA is cleaved. Even under forced conditions for exhaustive digestion, the GlcN−GlcA linkage is only partly cleaved. It is this property of lyase III that has enabled the isolation of a unique, nonsulfated antigenic determinant ΔUA−GlcN−UA−GlcNAc from HS and from partially de-N-acetylated K5 polysaccharide. It was unexpected that pentasaccharide fragments were also detected among the digestion products of the K5 polysaccharide used. It is possible that these are products of an additional glycosidase activity of lyase III, although other mechanisms cannot be completely ruled out.
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DOI: 10.1021/bi036250k
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<hi rend="italic">N</hi>-Acetylglucosamine−Glucuronic Acid Linkages to Heparin Lyase III<ref type="bib" target="#bi036250kAF2">†</ref></title><author><name sortKey="Chai, Wengang" sort="Chai, Wengang" uniqKey="Chai W" first="Wengang" last="Chai">Wengang Chai</name><affiliation wicri:level="4"><country xml:lang="fr">Suède</country><wicri:regionArea>MRC Glycosciences Laboratory, Imperial College Faculty of Medicine, Northwick Park Hospital Campus, Watford Road,Harrow, Middlesex HA1 3UJ, U.K., and Department of Medical Biochemistry and Microbiology, Uppsala University,The Biomedical Center, P.O. 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Heparin lyase III is known to cleave the linkages between N-acetylglucosamine (GlcNAc) or N-sulfated glucosamine (GlcNS) and glucuronic acid (GlcA) as the primary sites and the linkages between GlcNAc, GlcNAc(6S), or GlcNS and iduronic acid as secondary sites. N-Unsubstituted glucosamine (GlcN) occurs as a minor component in HS, and it has been associated with various bioactivities. Here we investigate the specificity of heparin lyase III toward the GlcN−GlcA linkage using a recombinant enzyme of high purity and as substrates the partially de-N-acetylated polysaccharide of Escherichia coli K5 strain and derived hexasaccharides. The specificity of lyase III toward the GlcN−GlcA linkage is deduced by sequencing of the oligosaccharide products using electrospray mass spectrometry with collision-induced dissociation and MS/MS scanning. The results demonstrate that under controlled conditions for partial digestion, lyase III does not act at the GlcN−GlcA linkage, whereas GlcNAc−GlcA is cleaved. Even under forced conditions for exhaustive digestion, the GlcN−GlcA linkage is only partly cleaved. It is this property of lyase III that has enabled the isolation of a unique, nonsulfated antigenic determinant ΔUA−GlcN−UA−GlcNAc from HS and from partially de-N-acetylated K5 polysaccharide. It was unexpected that pentasaccharide fragments were also detected among the digestion products of the K5 polysaccharide used. It is possible that these are products of an additional glycosidase activity of lyase III, although other mechanisms cannot be completely ruled out.</div></front></TEI><affiliations><list><country><li>Royaume-Uni</li><li>Suède</li></country><region><li>East Middle Sweden</li><li>Svealand</li></region><settlement><li>Uppsala</li></settlement><orgName><li>Université d'Uppsala</li></orgName></list><tree><country name="Suède"><region name="Svealand"><name sortKey="Chai, Wengang" sort="Chai, Wengang" uniqKey="Chai W" first="Wengang" last="Chai">Wengang Chai</name></region><name sortKey="Feizi, Ten" sort="Feizi, Ten" uniqKey="Feizi T" first="Ten" last="Feizi">Ten Feizi</name><name sortKey="Leteux, Christine" sort="Leteux, Christine" uniqKey="Leteux C" first="Christine" last="Leteux">Christine Leteux</name><name sortKey="Lindahl, Ulf" sort="Lindahl, Ulf" uniqKey="Lindahl U" first="Ulf" last="Lindahl">Ulf Lindahl</name><name sortKey="Westling, Camilla" sort="Westling, Camilla" uniqKey="Westling C" first="Camilla" last="Westling">Camilla Westling</name></country><country name="Royaume-Uni"><noRegion><name sortKey="Chai, Wengang" sort="Chai, Wengang" uniqKey="Chai W" first="Wengang" last="Chai">Wengang Chai</name></noRegion></country></tree></affiliations></record>Pour manipuler ce document sous Unix (Dilib)
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