Format

Send to

Choose Destination
See comment in PubMed Commons below
Nat Methods. 2011 Jul 3;8(8):677-83. doi: 10.1038/nmeth.1636.

A large-scale method to measure absolute protein phosphorylation stoichiometries.

Author information

1
Department of Cell Biology, Harvard Medical School, Boston, Massachusetts, USA.

Abstract

The functional role of protein phosphorylation is impacted by its fractional stoichiometry. Thus, a comprehensive strategy to study phosphorylation dynamics should include an assessment of site stoichiometry. Here we report an integrated method that relies on phosphatase treatment and stable-isotope labeling to determine absolute stoichiometries of protein phosphorylation on a large scale. This approach requires the measurement of only a single ratio relating phosphatase-treated and mock-treated samples. Using this strategy we determined stoichiometries for 5,033 phosphorylation sites in triplicate analyses from Saccharomyces cerevisiae growing through mid-log phase. We validated stoichiometries at ten sites that represented the full range of values obtained using synthetic phosphopeptides and found excellent agreement. Using bioinformatics, we characterized the biological properties associated with phosphorylation sites with vastly differing absolute stoichiometries.

PMID:
21725298
PMCID:
PMC3146562
DOI:
10.1038/nmeth.1636
[Indexed for MEDLINE]
Free PMC Article
PubMed Commons home

PubMed Commons

0 comments
How to join PubMed Commons

    Supplemental Content

    Full text links

    Icon for Nature Publishing Group Icon for PubMed Central
    Loading ...
    Support Center