Title :
Sialic acid-focused quantitative mouse serum glycoproteomics by multiple reaction monitoring assay
Abstract :
- Despite increasing importance of protein glycosylation, most of the large-scale glycoproteomics have been limited to profiling the sites of N-glycosylation
- However, in-depth knowledge of protein glycosylation to uncover functions and their clinical applications requires quantitative glycoproteomics eliciting both peptide and glycan sequences concurrently
- Here we describe a novel strategy for the multiplexed quantitative mouse serum glycoproteomics based on a specific chemical ligation, namely, reverse glycoblotting technique, focusing sialic acids and multiple reaction monitoring (MRM)
- LC-MS/MS analysis of de-glycosylated peptides identified 270 mouse serum peptides ( 95 glycoproteins ) as sialylated glycopeptides , of which 67 glycopeptides were fully characterized by MS/MS analyses in a straightforward manner
- We revealed the importance of a fragment ion containing innermost N-acetylglucosamine (GlcNAc) residue as MRM transitions regardless the sequence of the peptides
- Versatility of the reverse glycoblotting-assisted MRM assays was demonstrated by quantitative comparison of 25 targeted glycopeptides from 16 proteins between mice with homo and hetero types of diabetes disease model