Mass spectrometry of glycoproteins is
an emerging field in proteomics, poised to meet the
technical demand for elucidation of the structural complexity
and functions of the oligosaccharide components of molecules.
Considering the divergence of the mass spectrometric
methods employed for oligosaccharide analysis in recent
publications, it is necessary to establish technical
standards and demonstrate capabilities. In the present
study of the HUPO HGPI (Human Proteome Organisation
Human Disease Glycomics/Proteome Initiative), the same
samples of transferrin and immunoglobulin-G, both of
which have N-linked oligosaccharides, were analyzed
for oligosaccharides and their relative abundances in
20 laboratories, and the chromatographic and mass spectrometric
analysis results were evaluated. In general, matrix-assisted
laser desorption/ionization (MALDI) time-of-flight mass
spectrometry (MS) of permethylated oligosaccharide mixtures
carried out in six laboratories yielded good quantitation
based on the signal intensity of [M+Na]+ ions, and the
results can be correlated to those of chromatography
of reductive amination derivatives. For underivatized
oligosaccharide alditols, graphitized carbon-liquid
chromatography (LC)/electrospray ionization (ESI) MS
detecting deprotonated molecules in the negative ion
mode provided acceptable quantitation. Detailed analyses
of tryptic glycopeptides by MS were submitted by a few
laboratories employing either nanoLC/ESI MS/MS or MALDI
MS, and the capability to determine site-specific or
subclass-specific glycan profiles in these samples was
excellent. Taking into account the variety of MS technologies
and options for distinct protocols used in this study,
the results of this multi-institutional study indicate
that MS-based analysis appears as the efficient method
for identification and quantitation of oligosaccharides
in glycomic studies and endorse the power of MS for
glycopeptide characterization with high sensitivity
in proteomic programs. Together glycomic and glycoproteomic
methodologies of the type assessed in this study have
the potential to provide large volumes of quantitation
data for diagnostics and, more broadly, for underpinning
functional glycomics and systems biology research.
Wada, et al. (Glycobiology 2007 17 (4): 411-22) |
