title: Mapping site-specific protein N-glycosylations through liquid chromatography/mass spectrometry and targeted tandem mass spectrometry.
abstract: Glycosylation is one of the most common posttranslational modifications (PTMs) of proteins, the characterization of which is commonly achieved through proteomic protocol, involving trypsin digestion followed by liquid chromatography/tandem mass spectrometry (LC/MS/MS). However, it is often not possible to characterize all glycopeptides in a complex sample because of the high complexity of glycoproteomic samples, and the relative lower abundances of glycopeptides in comparison to the unmodified peptides. We present here a targeted MS/MS analysis approach, which utilizes a previously developed computational tool, GlyPID, to guide multiple experiments, thus permitting a complete characterization of all N-glycosylation sites of glycoproteins present in a complex sample. We have tested our approach using model glycoproteins analyzed by high-resolution LTQ-FT MS. The results demonstrate a potential use of our method for a high-throughput characterization of complex mixtures of glycosylated proteins.
authors.affiliation: School of Informatics and Computing, Indiana University, Bloomington, IN 47408, USA.
authors.forename: Yin
authors.initials: Y
authors.lastname: Wu
authors.forename: Yehia
authors.initials: Y
authors.lastname: Mechref
authors.forename: Iveta
authors.initials: I
authors.lastname: Klouckova
authors.forename: Anoop
authors.initials: A
authors.lastname: Mayampurath
authors.forename: Milos V
authors.initials: MV
authors.lastname: Novotny
authors.forename: Haixu
authors.initials: H
authors.lastname: Tang
mesh_terms: D000465:Algorithms; D000818:Animals; D002417:Cattle; D002853:Chromatography, Liquid / Q000379:methods*; D045424:Complex Mixtures / Q000737:chemistry; D019295:Computational Biology / Q000379:methods*; D006020:Glycopeptides / Q000737:chemistry* / Q000378:metabolism; D006031:Glycosylation; D006801:Humans; D010449:Peptide Mapping / Q000379:methods*; D011499:Protein Processing, Post-Translational; D015203:Reproducibility of Results; D012984:Software; D053719:Tandem Mass Spectrometry / Q000379:methods*; D000509:alpha-Fetoproteins / Q000737:chemistry
chemical_list: D045424:Complex Mixtures; D006020:Glycopeptides; D000509:alpha-Fetoproteins