IDAWG: Metabolic incorporation of stable isotope labels for quantitative glycomics of cultured cells. J Proteome Res 2009 Aug;8(8):3816-23
Date
05/20/2009Pubmed ID
19449840Pubmed Central ID
PMC4141490DOI
10.1021/pr8010028Scopus ID
2-s2.0-68549133585 97 CitationsAbstract
Robust quantification is an essential component of comparative -omic strategies. In this regard, glycomics lags behind proteomics. Although various isotope-tagging and direct quantification methods have recently enhanced comparative glycan analysis, a cell culture labeling strategy, that could provide for glycomics the advantages that SILAC provides for proteomics, has not been described. Here, we report the development of IDAWG, Isotopic Detection of Aminosugars With Glutamine, for the incorporation of differential mass tags into the glycans of cultured cells. In this method, culture media containing amide-(15)N-Gln is used to metabolically label cellular aminosugars with heavy nitrogen. Because the amide side chain of Gln is the sole source of nitrogen for the biosynthesis of GlcNAc, GalNAc, and sialic acid, we demonstrate that culturing mouse embryonic stems cells for 72 h in the presence of amide-(15)N-Gln media results in nearly complete incorporation of (15)N into N-linked and O-linked glycans. The isotopically heavy monosaccharide residues provide additional information for interpreting glycan fragmentation and also allow quantification in both full MS and MS/MS modes. Thus, IDAWG is a simple to implement, yet powerful quantitative tool for the glycomics toolbox.
Author List
Orlando R, Lim JM, Atwood JA 3rd, Angel PM, Fang M, Aoki K, Alvarez-Manilla G, Moremen KW, York WS, Tiemeyer M, Pierce M, Dalton S, Wells LAuthor
Kazuhiro Aoki PhD Associate Professor in the Cell Biology, Neurobiology and Anatomy department at Medical College of WisconsinMESH terms used to index this publication - Major topics in bold
AnimalsCarbohydrate Sequence
Cells, Cultured
Embryonic Stem Cells
Fourier Analysis
Glycomics
Isotope Labeling
Mass Spectrometry
Mice
Molecular Sequence Data
Nitrogen Isotopes
Polysaccharides
