title: Analysis of quantitative proteomic data generated via multidimensional protein identification technology.
abstract: We describe the analysis of quantitative proteomic samples via multidimensional protein identification technology (MudPIT). Ratio amounts of the soluble portion of the S. cerevisiae proteome from cultures of S. cerevisiae strain S288C grown in either 14N minimal media or 15N-enriched minimal media were mixed and digested into a complex peptide mixture. A 1 x 14N/1 x 15N complex peptide mixture was analyzed by single-dimensional reversed-phase chromatography and electrospray ionization quadrapole time-of-flight mass spectrometry in order to demonstrate the replacement of 14N by 15N under the growth conditions used. After conformation of the incorporation of 15N into the labeled sample, three separate samples consisting of a 1 x 14N/1 x 15N complex peptide mixture, a 5 x 14N/1 x 15N complex peptide mixture, and a 10 x 14N/1 x 15N complex peptide mixture were analyzed via MudPIT. We demonstrate the dynamic range of the system by analyzing a 1:1, 5:1, and 10:1 data set using the soluble portion from S. cerevisiae grown in either 14N or 15N-enriched minimal media. The method described provides an accurate way to undertake a large-scale quantitative proteomic study.
authors.affiliation: Proteomics, Torrey Mesa Research Institute, San Diego, California 92121, USA. michael.washburn@syngenta.com
authors.forename: Michael P
authors.initials: MP
authors.lastname: Washburn
authors.forename: Ryan
authors.initials: R
authors.lastname: Ulaszek
authors.forename: Cosmin
authors.initials: C
authors.lastname: Deciu
authors.forename: David M
authors.initials: DM
authors.lastname: Schieltz
authors.forename: John R
authors.initials: JR
authors.lastname: Yates
mesh_terms: D002851:Chromatography, High Pressure Liquid; D009587:Nitrogen Isotopes; D010446:Peptide Fragments / Q000032:analysis; D011506:Proteins / Q000032:analysis; D020543:Proteome / Q000032:analysis* / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000737:chemistry; D021241:Spectrometry, Mass, Electrospray Ionization; D013223:Statistics as Topic
chemical_list: D009587:Nitrogen Isotopes; D010446:Peptide Fragments; D011506:Proteins; D020543:Proteome