title: Combining amine metabolomics and quantitative proteomics of cancer cells using derivatization with isobaric tags.
abstract: Quantitative metabolomics and proteomics technologies are powerful approaches to explore cellular metabolic regulation. Unfortunately, combining the two technologies typically requires different LC-MS setups for sensitive measurement of metabolites and peptides. One approach to enhance the analysis of certain classes of metabolites is by derivatization with various types of tags to increase ionization and chromatographic efficiency. We demonstrate here that derivatization of amine metabolites with tandem mass tags (TMT), typically used in multiplexed peptide quantitation, facilitates amino acid analysis by standard nanoflow reversed-phase LC-MS setups used for proteomics. We demonstrate that this approach offers the potential to perform experiments at the MS1-level using duplex tags or at the MS2-level using novel 10-plex reporter ion-containing isobaric tags for multiplexed amine metabolite analysis. We also demonstrate absolute quantitative measurements of amino acids conducted in parallel with multiplexed quantitative proteomics, using similar LC-MS setups to explore cellular amino acid regulation. We further show that the approach can also be used to determine intracellular metabolic labeling of amino acids from glucose carbons.
authors.affiliation: Department of Cell Biology, Harvard Medical School , Boston, Massachusetts 02115, United States.
authors.forename: J Patrick
authors.initials: JP
authors.lastname: Murphy
authors.forename: Robert A
authors.initials: RA
authors.lastname: Everley
authors.forename: Jonathan L
authors.initials: JL
authors.lastname: Coloff
authors.forename: Steven P
authors.initials: SP
authors.lastname: Gygi
mesh_terms: D000588:Amines / Q000378:metabolism*; D006801:Humans; D055432:Metabolomics; D009369:Neoplasms / Q000378:metabolism / Q000473:pathology*; D040901:Proteomics
chemical_list: D000588:Amines