应用电子捕获解离质谱对蛋白质磷酸化位点的定位.pptVIP

Large Scale Localization of ProteinPhosphorylation by Use of Electron Capture Dissociation Mass Spectrometry;Steve M. M. Sweet?§, Christopher M. Bailey?§, Debbie L. Cunningham?§,John K. Heath?§, and Helen J. Cooper§?;Content;1. Introduction of background;2. Experiment procedures;3. Analysis of Results;FIG. 1. Overlap between DTAs leading to identifications by ECD and CID. All 4763 identifications (IDs) are from paired CID/ECD events .FIG. 2. Overlap between distinct phosphopeptides identified by ECD and CID. All 1220 identifications are from paired CID/ECDevents. ;FIG. 3. Binned distribution of SLoMo scores. The x axis has log scale. All identifications are from paired CID/ECD events. Identifications with only one possible site of localization are not included.;FIG. 4. Overlap between distinct, well localized (SLoMo 19) phosphopeptides identified by ECD and CID. All 725 identifications are from paired CID/ECD events. Identifications with only one possible localization are not included.;;FIG. 5. Example of isomeric phosphopeptide co-elution. a, ECD mass spectrum showing first and third serine phosphorylation (SLoMo score, 21.5). b, CID mass spectrum showing evidence for threonine phosphorylation (SLoMo score, 24.3). The site of phosphorylation indicated by SLoMo is shown uppermost inset in b.;FIG. 6. Multiply phosphorylated peptides identified and localized from ECD and CID mass spectra and sorted by charge state. All identifications are from paired events. Identifications with only one possible localization are not included.;4. Conclusion;In conclusion, our results indicate that combined ECD and CID analysis results in high confidence phosphopeptide identifications and phosphorylation site localization. Hybrid mass spectrometers, such as the LTQ-FT used in this work, are primarily used to measure precursor ions with high mass accuracy while carrying out rapid CID (at lower resolution). We showed that there are potential advantages to using both parts of the hybrid