The impact of peptide abundance and dynamic range on stable-isotope-based quantitative proteomic analyses.
Recently, mass spectrometry has been employed in many studies to provide unbiased, reproducible, and quantitative protein abundance information on a proteome-wide scale. However, how instruments' limited dynamic ranges impact the accuracy of such measurements has remained largely unexplored, especially in the context of complex mixtures. Here, we examined the distribution of peptide signal versus background noise (S/N) and its correlation with quantitative accuracy. With the use of metabolically labeled Jurkat cell lysate, over half of all confidently identified peptides had S/N ratios less than 10 when examined using both hybrid linear ion trap-Fourier transform ion cyclotron resonance and Orbitrap mass spectrometers. Quantification accuracy was also highly correlated with S/N. We developed a mass precision algorithm that significantly reduced measurement variance at low S/N beyond the use of highly accurate mass information alone and expanded it into a new software suite, Vista. We also evaluated the interplay between mass measurement accuracy and S/N; finding a balance between both parameters produced the greatest identification and quantification rates. Finally, we demonstrate that S/N can be a useful surrogate for relative abundance ratios when only a single species is detected.
Bakalarski
Corey E
CE
Department of Cell Biology, Harvard Medical School, Boston, Massachusetts 02115, USA.
Elias
Joshua E
JE
Villén
Judit
J
Haas
Wilhelm
W
Gerber
Scott A
SA
Everley
Patrick A
PA
Gygi
Steven P
SP
D000465:Algorithms; D045424:Complex Mixtures / Q000032:analysis / Q000737:chemistry; D017357:Cyclotrons; D005583:Fourier Analysis; D006801:Humans; D007553:Isotope Labeling; D007554:Isotopes / Q000032:analysis; D019169:Jurkat Cells; D013058:Mass Spectrometry / Q000379:methods*; D010455:Peptides / Q000032:analysis* / Q000737:chemistry; D020543:Proteome / Q000032:analysis*; D040901:Proteomics / Q000379:methods*; D012984:Software
D045424:Complex Mixtures; D007554:Isotopes; D010455:Peptides; D020543:Proteome
Methods. 2003 Feb;29(2):124-30
12606218
J Proteome Res. 2007 Apr;6(4):1482-91
17311443
J Proteome Res. 2006 May;5(5):1224-31
16674112
Science. 2007 Aug 3;317(5838):660-3
17673661
Nat Biotechnol. 2004 Nov;22(11):1459-66
15529173
Nat Biotechnol. 2006 Oct;24(10):1285-92
16964243
Mol Cell Proteomics. 2003 Nov;2(11):1198-204
14506205
Mol Cell. 2006 Apr 21;22(2):193-204
16630889
Nat Methods. 2007 Mar;4(3):207-14
17327847
Mol Cell Proteomics. 2005 Mar;4(3):246-54
15542864
Science. 2007 May 25;316(5828):1160-6
17525332
Mol Cell Proteomics. 2006 Jul;5(7):1326-37
16635985
Anal Chem. 2002 Oct 15;74(20):5383-92
12403597
J Proteome Res. 2007 Mar;6(3):1190-7
17330950
Anal Bioanal Chem. 2007 Nov;389(5):1409-19
17874083
Anal Chem. 2006 Jan 15;78(2):493-500
16408932
Electrophoresis. 1999 Dec;20(18):3551-67
10612281
J Am Soc Mass Spectrom. 1994 Nov;5(11):976-89
24226387
Proc Natl Acad Sci U S A. 2007 Jan 30;104(5):1488-93
17242355
Mol Cell Proteomics. 2005 Feb;4(2):214-23
15608339
J Proteome Res. 2004 May-Jun;3(3):621-6
15253445
J Mass Spectrom. 2005 Apr;40(4):430-43
15838939
J Am Soc Mass Spectrom. 2002 Jun;13(6):659-69
12056566
Nat Biotechnol. 1999 Oct;17(10):994-9
10504701
Mol Cell Proteomics. 2002 May;1(5):376-86
12118079
Cell. 2006 Nov 3;127(3):635-48
17081983
Nat Methods. 2007 Sep;4(9):677-8
17762871
Nat Biotechnol. 2007 May;25(5):566-8
17450129