Glycopeptide Enrichment Using a Combination of ZIC-HILIC and Cotton Wool for Exploring the Glycoproteome of Wheat Flour Albumins

Hydrophilic
liquid chromatography (HILIC) is used extensively as
a sample preparation step for glycopeptide enrichment in proteome
research. Here, we have applied cotton wool and a zwitterionic HILIC
(ZIC-HILIC) resin in solid-phase extraction microcolumns to provide
a higher loading capacity and broader specificity for glycopeptide
enrichment. This strategy was applied to tryptic digests of wheat
flour albumin extracts followed by simulataneous site-specific 18O labeling and deglycosylation using peptide-N-glycosidase A (PNGase A) in H218O. Subsequent
LC–MS/MS analysis allowed for assignment of 78 N-glycosylation
sites in 67 albumin proteins. Bioinformatic analysis revealed that
several of the identified glycoproteins show sequence similarity to
known food allergens. In addition, the potential impact of some of
the identified glycoproteins on wheat beer quality is discussed.