Separation and identification of neisserial lipooligosaccharide oligosaccharides using high-performance anion-exchange chromatography with pulsed amperometric detection.
We determined the optimal conditions for high-performance anion-exchange chromatography with pulsed amperometric detection (HPAE-PAD) of oligosaccharides (OS) released from neisserial lipooligosaccharides (LOS) by mild acid hydrolysis. We efficiently obtained detailed composition, sequence, and linkage information about high Mr LOS. We found that HPAE-PAD can discriminate isobaric (same Mr) molecules of different structure, for example, nLc4 and Gb4, distinguish alpha from beta chain extensions, and determine the number of phosphoethanolamine (PEA) substituents. HPAE-PAD provided quantitative information that could be used to compare the relative abundances of OS. We used HPAE-PAD to identify all of the known LOS alpha chain antennae. When used with antibody-binding profiles and exoglycosidase digestion results, HPAE-PAD can provide nearly complete structures rapidly.
Swanson
Karen V
KV
Centre for Immunochemistry, VA Medical Center (111W1), Department of Laboratory Medicine, University of California San Francisco, 4150 Clement Street, San Francisco, CA 94121, USA.
Griffiss
J McLeod
JM
D000143:Acids; D000838:Anions; D002236:Carbohydrate Conformation; D002240:Carbohydrate Sequence; D002851:Chromatography, High Pressure Liquid; D002852:Chromatography, Ion Exchange; D004563:Electrochemistry; D004983:Ethanolamines / Q000737:chemistry; D007887:Fabaceae / Q000201:enzymology; D005696:Galactosidases / Q000378:metabolism; D006868:Hydrolysis; D008070:Lipopolysaccharides / Q000737:chemistry*; D008969:Molecular Sequence Data; D009344:Neisseria gonorrhoeae / Q000737:chemistry*; D009844:Oligosaccharides / Q000737:chemistry* / Q000302:isolation & purification*; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
D000143:Acids; D000838:Anions; D004983:Ethanolamines; D008070:Lipopolysaccharides; D009844:Oligosaccharides; C023023:lipid-linked oligosaccharides; C005448:phosphorylethanolamine; D005696:Galactosidases