title: Isotope-targeted glycoproteomics (IsoTaG): a mass-independent platform for intact N- and O-glycopeptide discovery and analysis.
abstract: Protein glycosylation is a heterogeneous post-translational modification (PTM) that plays an essential role in biological regulation. However, the diversity found in glycoproteins has undermined efforts to describe the intact glycoproteome via mass spectrometry (MS). We present IsoTaG, a mass-independent chemical glycoproteomics platform for characterization of intact, metabolically labeled glycopeptides at the whole-proteome scale. In IsoTaG, metabolic labeling of the glycoproteome is combined with (i) chemical enrichment and isotopic recoding of glycopeptides to select peptides for targeted glycoproteomics using directed MS and (ii) mass-independent assignment of intact glycopeptides. We structurally assigned 32 N-glycopeptides and over 500 intact and fully elaborated O-glycopeptides from 250 proteins across three human cancer cell lines and also discovered unexpected peptide sequence polymorphisms (pSPs). The IsoTaG platform is broadly applicable to the discovery of PTM sites that are amenable to chemical labeling, as well as previously unknown protein isoforms including pSPs.
authors.affiliation: Department of Chemistry, University of California, Berkeley, California, USA.
authors.forename: Christina M
authors.initials: CM
authors.lastname: Woo
authors.affiliation: QB3/Chemistry Mass Spectrometry Facility, University of California, Berkeley, California, USA.
authors.forename: Anthony T
authors.initials: AT
authors.lastname: Iavarone
authors.affiliation: Department of Chemistry, University of California, Berkeley, California, USA.
authors.forename: David R
authors.initials: DR
authors.lastname: Spiciarich
authors.affiliation: Department of Chemistry, University of California, Berkeley, California, USA.
authors.forename: Krishnan K
authors.initials: KK
authors.lastname: Palaniappan
authors.affiliation: 1] Department of Chemistry, University of California, Berkeley, California, USA. [2] Department of Molecular and Cell Biology, University of California, Berkeley, California, USA. [3] Howard Hughes Medical Institute, Berkeley, California, USA.
authors.forename: Carolyn R
authors.initials: CR
authors.lastname: Bertozzi
mesh_terms: D045744:Cell Line, Tumor; D006023:Glycoproteins / Q000737:chemistry* / Q000378:metabolism; D006031:Glycosylation; D006801:Humans; D007553:Isotope Labeling; D015394:Molecular Structure; D020543:Proteome; D040901:Proteomics / Q000379:methods*; D053719:Tandem Mass Spectrometry / Q000379:methods
chemical_list: D006023:Glycoproteins; D020543:Proteome