Correlation of relative abundance ratios derived from peptide ion chromatograms and spectrum counting for quantitative proteomic analysis using stable isotope labeling.
In this study, S. cerevisiae crude membrane fractions were prepared using the acid-labile detergent RapiGest from cells grown under rich and minimal media conditions using 14N and 15N ammonium sulfate as the sole nitrogen source. Four independent MudPIT analyses of 1:1 mixtures of sample were prepared and analyzed via quantitative multidimensional protein identification technology on a two-dimensional ion trap mass spectrometer. Using the method described in this study, low-abundance integral membrane proteins with up to 14 transmembrane domains were identified and their protein expression determined when sufficient spectrum counting and ion chromatogram information was generated. We demonstrate that spectrum counting and mass spectrometry derived ion chromatograms strongly correlate for determining quantitative changes in protein expression. Spectrum counting proved more reproducible and has a wider dynamic range contributing to the deviation of the two quantitative approaches from a perfect positive correlation.
Zybailov
Boris
B
Stowers Institute for Medical Research, 1000 East 50th Street, Kansas City, Missouri 64110, USA.
Coleman
Michael K
MK
Florens
Laurence
L
Washburn
Michael P
MP
D000595:Amino Acid Sequence; D002853:Chromatography, Liquid / Q000379:methods*; D007553:Isotope Labeling; D013058:Mass Spectrometry / Q000379:methods*; D008969:Molecular Sequence Data; D010455:Peptides / Q000032:analysis* / Q000737:chemistry* / Q000378:metabolism; D040901:Proteomics / Q000379:methods*; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism
D010455:Peptides