Project : PXD000357

PRIDE Assigned Tags:
Biomedical Dataset



Identification of ERK1 direct substrates using stable isotope labeled kinase assay linked phosphoproteomics


Following the kinase assay linked phosphoproteomics strategy (KALIP) (Xue, L et al., 2012), we used extracellular signal-regulated kinases 1 (ERK1) to phosphorylate the HEK293 cell lysate under the in vitro kinase assay condition. The phosphorylated proteins were then isolated and identified by mass spectrometry. The in vitro phosphorylated proteins with new phosphates were further overlapped with reported in vivo ERK1-dependent phosphoproteomics data for the identification of bona fide direct substrates of ERK1. In total, we identified 27 direct substrates of ERK1. Data analysis procedure: Raw MS files from the LTQ-Orbitrap-Velos were analyzed by Proteome Discoverer 1.3. MS/MS spectra were searched against the IPI-human database (version 3.83) containing both forward and reverse protein sequences by the SEQUEST search engine. The false discovery rate (FDR) was set to 0.01 on the peptide level. Ingenuity Pathway Analysis (IPA) was applied for the functional annotation.

Sample Processing Protocol

See details in reference PMID : 25022875

Data Processing Protocol

See details in reference PMID : 25022875


Liang Xue, Biochemistry

Submission Date



Not available


LTQ Orbitrap Velos


Not available


phosphorylated residue


Not available

Experiment Type

Bottom-up proteomics


    Xue L, Wang P, Cao P, Zhu JK, Tao WA. Identification of ERK1 Direct Substrates using Stable Isotope Labeled Kinase Assay-Linked Phosphoproteomics. Mol Cell Proteomics. 2014 Jul 14. pii: mcp.O114.038588 PubMed(s) : 25022875

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