PXD007145 is an
original dataset announced via ProteomeXchange.
Dataset Summary
Title | Benchmarking LFQ, SILAC and MS2/MS3-based TMT quantification strategies for large-scale phosphoproteomics |
Description | Comprehensive mass spectrometry (MS)-based proteomics is now feasible, but reproducible and multiplexed quantification remains challenging especially for analysis of post-translational modifications (PTMs), such as phosphorylation. Here we compared the most popular quantification techniques for phosphoproteomics in context of cell-signaling studies: label-free quantification (LFQ), stable isotope labeling by amino acids in cell culture (SILAC) and MS2- and MS3-measured tandem mass tags (TMT). In a mixed species comparison with fixed phosphopeptide-ratios, we found LFQ and SILAC to be the most accurate techniques. MS2-based TMT suffered from substantial ratio compression, which MS3-based TMT could partly rescue. However, when analyzing phosphoproteome changes in the DNA damage response (DDR), we found that MS3-based TMT was outperformed by MS2-based TMT as it identified most significantly regulated phosphopeptides due to its higher precision and higher number of identifications. Finally, we show that the high accuracy of MS3-based TMT is crucial for determination of phosphorylation site stoichiometry using a novel multiplexing-dependent algorithm. |
HostingRepository | PRIDE |
AnnounceDate | 2018-03-13 |
AnnouncementXML | Submission_2018-03-19_01:45:30.xml |
DigitalObjectIdentifier | |
ReviewLevel | Peer-reviewed dataset |
DatasetOrigin | Original dataset |
RepositorySupport | Unsupported dataset by repository |
PrimarySubmitter | Alexander Hogrebe |
SpeciesList | scientific name: Homo sapiens (Human); NCBI TaxID: 9606; scientific name: Saccharomyces cerevisiae (Baker's yeast); NCBI TaxID: 4932; |
ModificationList | TMT6plex-126 reporter+balance reagent acylated residue; phosphorylated residue; iodoacetamide derivatized residue |
Instrument | Orbitrap Fusion Lumos |
Dataset History
Revision | Datetime | Status | ChangeLog Entry |
0 | 2017-07-27 07:09:56 | ID requested | |
1 | 2018-03-13 01:50:09 | announced | |
⏵ 2 | 2018-03-19 01:45:31 | announced | Updated publication reference for PubMed record(s): 29535314. |
Publication List
Hogrebe A, von Stechow L, Bekker-Jensen DB, Weinert BT, Kelstrup CD, Olsen JV, Benchmarking common quantification strategies for large-scale phosphoproteomics. Nat Commun, 9(1):1045(2018) [pubmed] |
Keyword List
curator keyword: Technical |
submitter keyword: phosphoproteomics, technical, benchmark, quantification, LFQ, SILAC, TMT, MS2, MS3 |
Contact List
Jesper V. Olsen |
contact affiliation | Novo Nordisk Foundation Center for Protein Research, Proteomics Program, University of Copenhagen, Denmark |
contact email | jesper.olsen@cpr.ku.dk |
lab head | |
Alexander Hogrebe |
contact affiliation | NNF Center for Protein Research, Copenhagen, Denmark |
contact email | alexander.hogrebe@cpr.ku.dk |
dataset submitter | |
Full Dataset Link List
Dataset FTP location
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PRIDE project URI |
Repository Record List
[ + ]
[ - ]
- PRIDE
- PXD007145
- Label: PRIDE project
- Name: Benchmarking LFQ, SILAC and MS2/MS3-based TMT quantification strategies for large-scale phosphoproteomics