title: Modified MuDPIT separation identified 4488 proteins in a system-wide analysis of quiescence in yeast.
abstract: A modified multidimensional protein identification technology (MudPIT) separation was coupled to an LTQ Orbitrap Velos mass spectrometer and used to rapidly identify the near-complete yeast proteome from a whole cell tryptic digest. This modified online two-dimensional liquid chromatography separation consists of 39 strong cation exchange steps followed by a short 18.5 min reversed-phase (RP) gradient. A total of 4269 protein identifications were made from 4189 distinguishable protein families from yeast during log phase growth. The "Micro" MudPIT separation performed as well as a standard MudPIT separation in 40% less gradient time. The majority of the yeast proteome can now be routinely covered in less than a days' time with high reproducibility and sensitivity. The newly devised separation method was used to detect changes in protein expression during cellular quiescence in yeast. An enrichment in the GO annotations "oxidation reduction", "catabolic processing" and "cellular response to oxidative stress" was seen in the quiescent cellular fraction, consistent with their long-lived stress resistant phenotypes. Heterogeneity was observed in the stationary phase fraction with a less dense cell population showing reductions in KEGG pathway categories of "Ribosome" and "Proteasome", further defining the complex nature of yeast populations present during stationary phase growth. In total, 4488 distinguishable protein families were identified in all cellular conditions tested.
authors.affiliation: Department of Chemical Physiology, The Scripps Research Institute, 10550 North Torrey Pines Road, La Jolla, California 92037, USA.
authors.forename: Kristofor J
authors.initials: KJ
authors.lastname: Webb
authors.forename: Tao
authors.initials: T
authors.lastname: Xu
authors.forename: Sung Kyu
authors.initials: SK
authors.lastname: Park
authors.forename: John R
authors.initials: JR
authors.lastname: Yates
mesh_terms: D059447:Cell Cycle Checkpoints; D002851:Chromatography, High Pressure Liquid; D002852:Chromatography, Ion Exchange; D020543:Proteome / Q000302:isolation & purification / Q000378:metabolism*; D040901:Proteomics; D012441:Saccharomyces cerevisiae / Q000166:cytology / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000302:isolation & purification / Q000378:metabolism*; D053719:Tandem Mass Spectrometry
chemical_list: D020543:Proteome; D029701:Saccharomyces cerevisiae Proteins
references.citation: Bioinformatics. 2012 Jun 15;28(12):1652-4
references.pmid: 22539673
references.citation: Anal Chem. 1984 Oct;56(12):1258A-1260A, 1262A, 1264A passim
references.pmid: 6507850
references.citation: J Proteomics. 2011 Nov 18;74(12):2837-45
references.pmid: 21782986
references.citation: J Bacteriol. 1994 Sep;176(18):5802-13
references.pmid: 8083172
references.citation: Microbiol Mol Biol Rev. 2004 Jun;68(2):187-206
references.pmid: 15187181
references.citation: Nat Methods. 2012 May 30;9(6):524-5
references.pmid: 22669647
references.citation: Subcell Biochem. 2012;57:123-43
references.pmid: 22094420
references.citation: Cell Div. 2011 Dec 09;6(1):20
references.pmid: 22152110
references.citation: Rapid Commun Mass Spectrom. 2004;18(18):2162-8
references.pmid: 15317041
references.citation: Mol Biol Cell. 2008 Mar;19(3):1271-80
references.pmid: 18199684
references.citation: Mol Cell Proteomics. 2005 Sep;4(9):1265-72
references.pmid: 15958392
references.citation: Mol Cell Proteomics. 2012 Mar;11(3):M111.013722
references.pmid: 22021278
references.citation: Anal Chem. 2010 Oct 15;82(20):8696-702
references.pmid: 20845935
references.citation: BMC Bioinformatics. 2008 Jul 21;9:316
references.pmid: 18644148
references.citation: Nature. 2008 Oct 30;455(7217):1251-4
references.pmid: 18820680
references.citation: Anal Chem. 2001 Dec 1;73(23):5683-90
references.pmid: 11774908
references.citation: Nature. 2003 Oct 16;425(6959):737-41
references.pmid: 14562106
references.citation: Mol Biosyst. 2007 May;3(5):354-60
references.pmid: 17460794
references.citation: Nature. 2013 Feb 14;494(7436):266-70
references.pmid: 23334424
references.citation: Nat Biotechnol. 2001 Mar;19(3):242-7
references.pmid: 11231557
references.citation: J Cell Biol. 2008 Jun 2;181(5):737-45
references.pmid: 18504300
references.citation: J Cell Biol. 2006 Jul 3;174(1):89-100
references.pmid: 16818721
references.citation: J Am Soc Mass Spectrom. 2002 Jun;13(6):659-69
references.pmid: 12056566
references.citation: J Proteome Res. 2002 Jan-Feb;1(1):21-6
references.pmid: 12643522
references.citation: FEMS Microbiol Rev. 2012 Mar;36(2):306-39
references.pmid: 21658086
references.citation: Anal Chem. 2004 Jul 15;76(14):4193-201
references.pmid: 15253663
references.citation: Nat Methods. 2010 Jan;7(1):43-6
references.pmid: 19966807
references.citation: Mol Biol Cell. 2011 Apr;22(7):988-98
references.pmid: 21289090
references.citation: Anal Chem. 2009 Sep 15;81(18):7757-65
references.pmid: 19689114