biostudies-literatureBeausoleil SANHGRI NIH HHSNIGMS NIH HHS12130-5https://www.ebi.ac.uk/biostudies/studies/S-EPMC514446biostudies-literatureUnknown101(33)Determining the site of a regulatory phosphorylation event is often essential for elucidating specific kinase-substrate relationships, providing a handle for understanding essential signaling pathways and ultimately allowing insights into numerous disease pathologies. Despite intense research efforts to elucidate mechanisms of protein phosphorylation regulation, efficient, large-scale identification and characterization of phosphorylation sites remains an unsolved problem. In this report we describe an application of existing technology for the isolation and identification of phosphorylation sites. By using a strategy based on strong cation exchange chromatography, phosphopeptides were enriched from the nuclear fraction of HeLa cell lysate. From 967 proteins, 2,002 phosphorylation sites weProceedings of the National Academy of Sciences of the United States of AmericaLarge-scale characterization of HeLa cell nuclear phosphoproteins.PMC514446GM67945R01 GM056203GMS6203T32 HG000041HG00041R01 GM067945K22 HG000041Schwartz DLi JBeausoleil SACohn MAVillen JCantley LCGygi SPJedrychowski MElias JEfalseLarge-scale characterization of HeLa cell nuclear phosphoproteins.Determining the site of a regulatory phosphorylation event is often essential for elucidating specific kinase-substrate relationships, providing a handle for understanding essential signaling pathways and ultimately allowing insights into numerous disease pathologies. Despite intense research efforts to elucidate mechanisms of protein phosphorylation regulation, efficient, large-scale identification and characterization of phosphorylation sites remains an unsolved problem. In this report we describe an application of existing technology for the isolation and identification of phosphorylation sites. By using a strategy based on strong cation exchange chromatography, phosphopeptides were enriched from the nuclear fraction of HeLa cell lysate. From 967 proteins, 2,002 phosphorylation sites we2004-01-01T00:00:00Z2004 Aug2025-04-05T15:26:27.285Z2019-03-27T00:50:42ZS-EPMC5144461530293510.1073/pnas.0404720101