Glycosaminoglycan characterization by electrospray ionization mass spectrometry including fourier transform mass spectrometry

Abstract

Electrospray ionization mass spectrometry (ESI MS) is a versatile analytical technique in glycomics of glycosaminoglycans (GAGs). Combined with enzymology, ESI MS is used for assessing changes in disaccharide composition of GAGs biosynthesized under different environmental or physiological conditions. ESI coupled with high-resolution mass analyzers such as a Fourier transform mass spectrometer (FTMS) permits accurate mass measurement of large oligosaccharides and intact GAGs as well as structural characterization of GAG oligosaccharides using information-rich fragmentation methods such as electron detachment dissociation. The first part of this chapter describes methods for disaccharide compositional profiling using ESI MS and the second part is dedicated to FTMS and tandem MS methods of GAG compositional and structural analysis.

Figure 3.1

The Domon-Costello nomenclature for naming hexose fragments. Glycosidic cleavages are denoted with B or C if they contain the nonreducing end (NRE), and Y or Z if they contain the reducing end (RE). Cross-ring cleavages are denoted by A or X with A containing the NRE and X containing the RE. Cross-ring superscripts indicate the cleaved bonds and subscripts indicated the position along the oligosaccharide. Complementary cleavage subscripts add up to the length of the oligosaccharide, for example, B₃ + Y₁ indicate a tetrasaccharide.

Figure 3.2

A scheme of electron-based activation of multiply charged anions during the EDD experiment.

Figure 3.3

Structures, monoisotopic masses, and observed m/z of eight CS/DS disaccharide standards.

Figure 3.4

Elution profiles of CS/DS disaccharides separated by IP RP HPLC: (A) total ion chromatogram, (B) extracted ion chromatogram for m/z 481, 662, 843, and 1024, and (C) absorbance trace at 232 nm.

Figure 3.5

Positive-ion ESI mass spectra of CS/DS disaccharides: (A) unsulfated disaccharide, (B) monosulfated disaccharide, (C) disulfated disaccharide, and (D) trisulfated disaccharide. Characteristic loss of 79 mass units can be attributed to HXA/Na exchange, (−101 + 22).

Figure 3.6

Structures, monoisotopic masses, and observed m/z of eight HS disaccharide standards.

Figure 3.7

Elution profiles of HS disaccharides separated by IP RP HPLC: (A) Extracted negative-ion chromatogram for m/z 378, 416, 458, 496, 538, and 576, and (B) absorbance trace at 232 nm.

Figure 3.8

PAGE and the negative ion ESI FTMS analysis of a bikunin GAG fraction isolated using continuous-elution PAGE: an example of data interpretation.

Figure 3.9

An illustration of the MS signal improvement achieved with quadrupole mass filter: ESI FT ICR MS analysis of intact bikunin GAG mixture (A) without mass filter, and (B) with a 20 m/z mass window. Overlapping small windows can be combined into a full m/z range mass spectrum with improved S/N.

Figure 3.10

A diagram of the Bruker Apex QeFTMS instrument.

Figure 3.11

An example of the tandem MS spectra, IRMPD and EDD, of an HS tetrasaccharide (ΔUAGlcNAc6S-GlcAGlcNAc6S) with the assigned fragments.