Ultra-deep human phosphoproteome reveals different regulatory nature of Tyr and Ser/Thr-based signaling

ABSTRACT: Regulatory protein phosphorylation controls nearly every normal and pathophysiological signaling system in eukaryotic cells. Despite great advances in mass spectrometry based-proteomics, the total number, localization and site-specific stoichiometry of this post-translational modification (PTM) are unknown. Here we develop stringent experimental and computational workflow, capable of mapping more than 50,000 distinct phosphorylated peptides in a single human cancer cell line. Label-free quantitation determined very high stoichiometries in mitosis or growth factor signaling and more than three-quarters of cellular proteins were detected as phosphoproteins. The proportion of phospho-Tyr drastically decreases as coverage of the phosphoproteome increases, whereas Ser/Thr sites only saturate for technical reasons. Tyrosine phosphorylation is maintained at especially low stoichiometric levels in the absence of specific signaling events. Unexpectedly, it is statistically enriched on higher abundance proteins and this correlates with the substrate Km values of tyrosine kinases. Our data suggests that P-Tyr should be considered a functionally separate PTM of eukaryotic proteomes.

INSTRUMENT(S): Q Exactive

ORGANISM(S): Homo Sapiens (ncbitaxon:9606)

SUBMITTER: Matthias Mann  

PROVIDER: MSV000080701 | MassIVE | Tue Mar 28 21:28:00 BST 2017

SECONDARY ACCESSION(S): PXD000612

REPOSITORIES: MassIVE