Simultaneous qualitative and quantitative analysis of the Escherichia coli proteome: a sweet tale.
We describe a novel LCMS approach to the relative quantitation and simultaneous identification of proteins within the complex milieu of unfractionated Escherichia coli. This label-free, LCMS acquisition method observes all detectable, eluting peptides and their corresponding fragment ions. Postacquisition data analysis methods extract both the chromatographic and the mass spectrometric information on the tryptic peptides to provide time-resolved, accurate mass measurements, which are subsequently used for quantitation and identification of constituent proteins. The response of E. coli to carbon source variation is well understood, and it is thus commonly used as a model biological system when validating an analytical method. Using this LCMS approach, we characterized proteins isolated from E. coli grown in glucose, lactose, and acetate. The change in relative abundance of the corresponding proteins was measured from peptides common to both conditions. Protein identities were also determined for those peptides that were unique to each condition, and these identities were found to be consistent with the underlying biochemical restrictions imposed by the growth conditions. The relative change in abundance of the characterized proteins ranged from 0.1- to 90-fold among the three binary comparisons. The overall coverage of the characterized proteins ranged from 10 to 80%, consisting of one to 34 peptides per protein. The quantitative results obtained from our study were comparable to other existing proteomic and transcriptional profiling approaches. This study illustrates the robustness of this novel LCMS approach for the simultaneous quantitative and comprehensive qualitative analysis of proteins in complex mixtures.
Silva
Jeffrey C
JC
Waters Corporation, Milford, Massachusetts 01757-3696, USA. jeff_silva@waters.com
Denny
Richard
R
Dorschel
Craig
C
Gorenstein
Marc V
MV
Li
Guo-Zhong
GZ
Richardson
Keith
K
Wall
Daniel
D
Geromanos
Scott J
SJ
D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000737:chemistry*; D029968:Escherichia coli Proteins / Q000737:chemistry*; D013058:Mass Spectrometry / Q000379:methods*; D020543:Proteome
D029968:Escherichia coli Proteins; D020543:Proteome