Generating heparan sulfate saccharide libraries for glycomics applications.
Natural and semi-synthetic heparan sulfate (HS) saccharide libraries are a valuable resource for investigating HS structure-function relationships, enabling high-throughput glycomics studies. Owing to the difficulty of chemical or in vitro enzymatic synthesis of HS saccharides, the structural diversity displayed in saccharides from tissue or cell sources cannot be readily accessed. In contrast, saccharide libraries can be generated by partial digestion of tissue-derived HS polysaccharide chains and chromatographic fractionation of the resulting saccharide mixtures. Fractionation is initially on the basis of hydrodynamic volume, using size exclusion chromatography. Further fractionation, on the basis of charge using strong anion exchange, can subsequently be applied. Desalting and sample concentration follows each fractionation step. Chromatographic fractions are generated that contain purified, or partially purified, saccharides. Here we describe a comprehensive protocol for generation of structurally diverse natural saccharide libraries from HS variants that is fast (approximately 3 weeks) and reproducible.
Powell
Andrew K
AK
Centre for Glycobiology, School of Biological Sciences, University of Liverpool, Liverpool, UK.
Ahmed
Yassir A
YA
Yates
Edwin A
EA
Turnbull
Jeremy E
JE
D002845:Chromatography; D054794:Glycomics / Q000379:methods*; D019763:Heparin Lyase; D006497:Heparan Sulfate / Q000737:chemistry* / Q000302:isolation & purification; D015394:Molecular Structure; D009005:Monosaccharides / Q000737:chemistry / Q000302:isolation & purification; D009844:Oligosaccharides / Q000737:chemistry / Q000302:isolation & purification
D019763:Heparin Lyase; D006497:Heparan Sulfate; D009005:Monosaccharides; D009844:Oligosaccharides