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Gewählte Publikation:

Pabst, M; Bondili, JS; Stadlmann, J; Mach, L; Altmann, F; .
(2007): Mass + retention time = structure: a strategy for the analysis of N-glycans by carbon LC-ESI-MS and its application to fibrin N-glycans.
Anal Chem. 2007; 79(13):5051-5057 FullText FullText_BOKU

Abstract:
Analysis of the numerous possible, often isobaric structures of protein-bound oligosaccharides calls for a high-performance two-dimensional method that combines liquid chromatography's ability to separate isomers and mass spectrometry's ability to determine glycan composition. Here we investigate the usefulness of porous graphitic carbon columns coupled to ESI-MS for the separation of N-glycans with two or more sialic acids. Internal standards helped to rectify retention time fluctuations and thus allowed elution times to play an essential role in the structural assignment of peaks. For generation of a retention time library, standards representing the possible isomers of diantennary non-, mono-, and disialylated N-glycans, differing in the linkage of galactose and sialic acids as well as isobaric hybrid-type N-glycans, were produced using recombinant glycosyltransferases. Once the retention times library was established, isomers could be identified by LC-ESI-MS in the positive mode without additional MS/MS experiments. The method was applied for the detailed structural analysis of fibrin(ogen) N-glycans from various species (human, cow, pig, mouse, rat, cat, dog, Chinese hamster, horse, sheep, and chicken). All fibrins contained diantennary N-glycans. They differed in the occurrence of beta 1,3-linked galactose, alpha 2,3-linked sialic acids, and N-glycolylneuraminic acid, in the mono/diantennary glycan ratio, and in the O-acetylation of neuraminic acids. The separation system's potential for analyzing tri- and tetrasialylated N-glycans was demonstrated.
Autor*innen der BOKU Wien:
Altmann Friedrich
Mach Lukas
Pabst Martin
Stadlmann Johannes
BOKU Gendermonitor:

Find related publications in this database (using NML MeSH Indexing)
Acetylation;Animals;Carbohydrate Sequence;Carbon/chemistry*;Chromatography, Liquid/methods*;Fibrin/analogs & derivatives;Fibrin/analysis*;Galactose/chemistry;Glycosyltransferases/metabolism;Humans;Isomerism;Molecular Sequence Data;Polysaccharides/analysis*;Sialic Acids/chemistry;Species Specificity;Spectrometry, Mass, Electrospray Ionization/methods*;Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization/methods;Time Factors;Transferrin/analysis;



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