Glycomics using mass spectrometry

Abstract

Mass spectrometry plays an increasingly important role in structural glycomics. This review provides an overview on currently used mass spectrometric approaches such as the characterization of glycans, the analysis of glycopeptides obtained by proteolytic cleavage of proteins and the analysis of glycosphingolipids. The given examples are demonstrating the application of mass spectrometry to study glycosylation changes associated with congenital disorders of glycosylation, lysosomal storage diseases, autoimmune diseases and cancer.

Fig. 1

Permethylated serum N-glycans measured by MALDI-linear ion trap-MS (a). Tandem mass spectrum of the sodiated precursor ion at m/z 2968. Fragments at m/z 1022, 1143, and 2690 are indicative of antenna fucosylation, whereas fragments at m/z 1113, 1317, and 2516 mark core fucosylation (b). The inset in (b) shows an MS3 experiment of m/z 1022 confirming the proposed structurewith terminal sialic acid and antenna fucosylation. Filled square, GlcNAc, empty circle, galactose; filled circle, mannose; filled triangle, fucose; filled diamond, N-acetylneuraminic acid. Taken from [39] with permission

Fig. 2

IgG Fc-glycosylation profiling by LC-quadrupole-TOF-MS. Long-term stability of MS detection in nanoLC-MS is achieved using a sheath-flow ESI spray with a sheath flow of 2 μl/min 50 % isopropanol, 20 % propionic acid (a). The extracted ion chromatograms of IgG1, IgG4 and IgG2/3 tryptic Fc glycopeptide species (b), and the mass spectrum of the IgG1 Fc glycopeptides elution range is shown in (c). Glycopeptide signals observed below m/z 1200 are triple protonated, and signals above m/z 1200 are double protonated. The excellent repeatability of the overall sample preparation and analysis method is demonstrated in (d) for IgG1. Blue square, N-acetylglucosamine; red triangle, fucose; green circle, mannose; yellow circle, galactose; purple diamond, N-acetylneuraminic acid

Fig. 3

High performance-anion exchange chromatography with online MS detection of urinary oligosaccharides of a GM1-gangliosidosis patient (a). Next to the total ion chromatogram (TIC) specific extracted ion chromatograms are given labeled with the composition of the oligosaccharide in terms of hexoses (H) and N-acetylhexosamines (N). The ion trap tandem mass spectra obtained for the two detected H3N2 isomers are shown in (b) and (c). Green circle, mannose; yellow circle, galactose; blue square, N-acetylglucosamine. Fragment ions are assigned according to Domon and Costello [74]. Taken from [32] with permission

Fig. 4

Low core-fucosylation of anti-HPA-3a alloantibodies. Fc glycosylation of total serum IgG1 (a) and anti-HPA3a alloantibodies from a patient with pregnancy complications (fetal and neonatal alloimmune thrombocytopenia; FNAIT) (b). Glycopeptides were detected in triple protonated form by nanoLC-ESI-ion trap-MS carrying neutral N-glycan chains (left panels) and acidic N-glycan chains (right panels). In (b) the assigned structures representing afucosylated glycoforms are highlighted in red. Blue square, N-acetylglucosamine; yellow circle, galactose, green circle, mannose; red triangle, fucose; purple diamond, N-acetylneuraminic acid; pep, tryptic peptide moiety; asterisk, non-glycopeptide signal. Taken from [82] with permission