Characterization of Glycoconjugates by Low- and High-Energy CID Mass Spectrometry

Description

The analysis of carbohydrate structures is a challenging task due to the possible variations in sequence and linkages between the individual monomer units and the general tendency of glycans to branch. Mass spectrometry (MS) and particularly low- and high-energy collision-induced dissociation (CID) analysis has become a powerful tool for the investigation this class of analytes. The characterization of glycoconjugates can be even more sophisticated, since the fragmentation of the glycan-part typically takes place at different collision energies compared to the non-glycan part. Several examples of the detailed structural characterization of glyconjugates are presented.

Multistage matrix-assisted laser desorption/ionization - ion trap/reflectron time-of-flight mass spectrometry (MALDI-IT/RTOF-MS) was applied to the analysis of Flavomycin, a commercially available antimicrobial growth promoter. The structurally closely related phosphoglycolipids, named moenomycins, that constitue this product are the only known antibiotically active compunds to date that directly interact with the bacterial transglycosylases and thus represent an interesting template for the design of new antibiotic drugs. Based on the derived CID fragmentation pathway of the five known major moenomycins, four not yet described moenomycin-type constituents could be characterized.

The same technique together with ESI-IT-MS was utilized for the analysis of the site specific glycosylaton pattern of human antithrombin, a serine-protease-inhibitor that is importantly involved in the regulation of blood coagulation. The glycopeptides were isolated by lectin-affinity chromatography after tryptic digestion of the highly purified protein. Our results show that the variability in the carbohydrate structures differs site-specifically.