Quantitative proteome profiling using stable isotope protein
tagging and automated tandem mass spectrometry (MS/MS) is an emerging
technology with great potential for the functional analysis of biological
systems and for the detection of clinical diagnostic or prognostic marker
proteins. Owing to the enormous complexity of proteomes, their comprehensive
analysis is an as-yet-unresolved technical challenge. However, biologically or
clinically important information can be obtained if specific, information-rich
protein classes, or sub-proteomes, are isolated and analyzed. Glycosylation is
the most common post-translational modification. Here we describe a method for
the selective isolation, identification and quantification of peptides that
contain N-linked carbohydrates. It is based on the conjugation of glycoproteins
to a solid support using hydrazide chemistry, stable isotope labeling of
glycopeptides and the specific release of formerly N-linked glycosylated
peptides via peptide- N-glycosidase F (PNGase F). The recovered peptides are
then identified and quantified by MS/MS. We applied the approach to the
analysis of plasma membrane proteins and proteins contained in human blood
serum.
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REFERENCE Glycoproteins Nature Encyclopaedia of Life Sciences
