Characterisation by LSI-MS and 1H NMR spectroscopy of tetra-, hexa-, and octa-saccharides of porcine intestinal heparin.
The characterisation of oligosaccharide fragments isolated from enzymatically depolymerised porcine intestinal heparin is required in order to probe structure/function relationships of heparin in anticoagulation, antiangiogenesis and antiviral activity. We have used both LSI-MS and 600-MHz 1H NMR with chemical shift assignment by comprehensive 1H-1H TOCSY experiments to fully characterise the major oligosaccharide components including 4 tetrasaccharides, 3 hexasaccharides, and 2 octasaccharides. One of the octasaccharides has not been identified previously and has the structure: delta UA(2S)-GlcNS(6S)-IdoA(2S)-GlcNS(6S)-IdoA(2S)- GlcNS(6S)-GlcA-GlcNS(6S), where delta UA is 4,5-unsaturated uronic acid (4-deoxy-alpha-L-threo-hex-4-enopyranosyluronic acid), GlcN is --> 4)-alpha-D-glucosamine, IdoA is --> 4)-alpha-L-iduronic acid, GlcA is --> 4)-beta-D-glucuronic acid, and 2-O-, 6-O-, and 2-N-sulfate are abbreviated to 2S, 6S, and NS, respectively. Nearly complete NMR proton chemical shifts are reported for this data set. In addition a novel approach involving oxymercuration-lipid conjugation was used to independently assign sulfate substitution on the delta UA residues.
Chai
W
W
Clinical Mass Spectrometry Section, MRC Clinical Research Centre, Harrow, Middx, United Kingdom.
Hounsell
E F
EF
Bauer
C J
CJ
Lawson
A M
AM
D000818:Animals; D002240:Carbohydrate Sequence; D002851:Chromatography, High Pressure Liquid; D006493:Heparin / Q000737:chemistry*; D007422:Intestines / Q000737:chemistry*; D009682:Magnetic Resonance Spectroscopy; D008969:Molecular Sequence Data; D009844:Oligosaccharides / Q000737:chemistry* / Q000302:isolation & purification; D017421:Sequence Analysis; D018629:Spectrometry, Mass, Secondary Ion; D013552:Swine
D009844:Oligosaccharides; D006493:Heparin