title: One-Step Selective Exoenzymatic Labeling (SEEL) Strategy for the Biotinylation and Identification of Glycoproteins of Living Cells.
abstract: Technologies that can visualize, capture, and identify subsets of biomolecules that are not encoded by the genome in the context of healthy and diseased cells will offer unique opportunities to uncover the molecular mechanism of a multitude of physiological and disease processes. We describe here a chemical reporter strategy for labeling of cell surface glycoconjugates that takes advantage of recombinant glycosyltransferases and a corresponding sugar nucleotide functionalized by biotin. The exceptional efficiency of this method, termed one-step selective exoenzymatic labeling, or SEEL, greatly improved the ability to enrich and identify large numbers of tagged glycoproteins by LC-MS/MS. We further demonstrated that this labeling method resulted in far superior enrichment and detection of glycoproteins at the plasma membrane compared to a sulfo-NHS-activated biotinylation or two-step SEEL. This new methodology will make it possible to profile cell surface glycoproteomes with unprecedented sensitivity in the context of physiological and disease states.
authors.affiliation: Complex Carbohydrate Research Center, University of Georgia, 315 Riverbend Road, Athens, Georgia 30602, United States.|Department of Chemistry, University of Georgia, 315 Riverbend Road, Athens, Georgia 30602, United States.
authors.forename: Tiantian
authors.initials: T
authors.lastname: Sun
authors.affiliation: Complex Carbohydrate Research Center, University of Georgia, 315 Riverbend Road, Athens, Georgia 30602, United States.
authors.forename: Seok-Ho
authors.initials: SH
authors.lastname: Yu
authors.affiliation: Complex Carbohydrate Research Center, University of Georgia, 315 Riverbend Road, Athens, Georgia 30602, United States.
authors.forename: Peng
authors.initials: P
authors.lastname: Zhao
authors.affiliation: Complex Carbohydrate Research Center, University of Georgia, 315 Riverbend Road, Athens, Georgia 30602, United States.
authors.forename: Lu
authors.initials: L
authors.lastname: Meng
authors.affiliation: Complex Carbohydrate Research Center, University of Georgia, 315 Riverbend Road, Athens, Georgia 30602, United States.|Department of Biochemistry and Molecular Biology, University of Georgia, 315 Riverbend Road, Athens, Georgia 30602, United States.
authors.forename: Kelley W
authors.initials: KW
authors.lastname: Moremen
authors.affiliation: Complex Carbohydrate Research Center, University of Georgia, 315 Riverbend Road, Athens, Georgia 30602, United States.|Department of Biochemistry and Molecular Biology, University of Georgia, 315 Riverbend Road, Athens, Georgia 30602, United States.
authors.forename: Lance
authors.initials: L
authors.lastname: Wells
authors.affiliation: Complex Carbohydrate Research Center, University of Georgia, 315 Riverbend Road, Athens, Georgia 30602, United States.|Department of Biochemistry and Molecular Biology, University of Georgia, 315 Riverbend Road, Athens, Georgia 30602, United States.
authors.forename: Richard
authors.initials: R
authors.lastname: Steet
authors.affiliation: Complex Carbohydrate Research Center, University of Georgia, 315 Riverbend Road, Athens, Georgia 30602, United States.|Department of Chemistry, University of Georgia, 315 Riverbend Road, Athens, Georgia 30602, United States.|Department of Chemical Biology and Drug Discovery, Utrecht Institute for Pharmaceutical Sciences, and Bijvoet Center for Biomolecular Research, Utrecht University, Universiteitsweg 99, 3584 CG Utrecht, The Netherlands.
authors.forename: Geert-Jan
authors.initials: GJ
authors.lastname: Boons
mesh_terms: D019921:Biotinylation; D002470:Cell Survival; D004798:Enzymes / Q000378:metabolism*; D006023:Glycoproteins / Q000378:metabolism*; D006367:HeLa Cells; D006801:Humans; D008247:Lysosomes / Q000378:metabolism; D009711:Nucleotides / Q000737:chemistry / Q000378:metabolism; D040901:Proteomics; D013194:Staining and Labeling; D013379:Substrate Specificity
chemical_list: D004798:Enzymes; D006023:Glycoproteins; D009711:Nucleotides