A probability-based approach for high-throughput protein phosphorylation analysis and site localization.
Article Details
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Beausoleil SA, Villen J, Gerber SA, Rush J, Gygi SP
A probability-based approach for high-throughput protein phosphorylation analysis and site localization.
Nat Biotechnol. 2006 Oct;24(10):1285-92. Epub 2006 Sep 10.
- PubMed ID
- 16964243 [ View in PubMed]
- Abstract
Data analysis and interpretation remain major logistical challenges when attempting to identify large numbers of protein phosphorylation sites by nanoscale reverse-phase liquid chromatography/tandem mass spectrometry (LC-MS/MS) (Supplementary Figure 1 online). In this report we address challenges that are often only addressable by laborious manual validation, including data set error, data set sensitivity and phosphorylation site localization. We provide a large-scale phosphorylation data set with a measured error rate as determined by the target-decoy approach, we demonstrate an approach to maximize data set sensitivity by efficiently distracting incorrect peptide spectral matches (PSMs), and we present a probability-based score, the Ascore, that measures the probability of correct phosphorylation site localization based on the presence and intensity of site-determining ions in MS/MS spectra. We applied our methods in a fully automated fashion to nocodazole-arrested HeLa cell lysate where we identified 1,761 nonredundant phosphorylation sites from 491 proteins with a peptide false-positive rate of 1.3%.
DrugBank Data that Cites this Article
- Polypeptides
Name UniProt ID Pyruvate kinase PKM P14618 Details DNA ligase 3 P49916 Details Bifunctional glutamate/proline--tRNA ligase P07814 Details DNA ligase 1 P18858 Details Serine/threonine-protein kinase A-Raf P10398 Details Abelson tyrosine-protein kinase 2 P42684 Details cGMP-inhibited 3',5'-cyclic phosphodiesterase A Q14432 Details Chromodomain-helicase-DNA-binding protein 1 O14646 Details Choline-phosphate cytidylyltransferase A P49585 Details Serine/threonine-protein kinase B-raf P15056 Details Cytosolic phospholipase A2 P47712 Details Pyrroline-5-carboxylate reductase 2 Q96C36 Details DNA topoisomerase 2-alpha P11388 Details Glucocorticoid receptor P04150 Details CTP synthase 1 P17812 Details Signal transducer and activator of transcription 5B P51692 Details Fatty acid synthase P49327 Details Deoxyuridine 5'-triphosphate nucleotidohydrolase, mitochondrial P33316 Details Thymidine kinase, cytosolic P04183 Details Kinesin-like protein KIF11 P52732 Details Keratin, type II cytoskeletal 8 P05787 Details Pyridoxal-dependent decarboxylase domain-containing protein 1 Q6P996 Details Molybdenum cofactor sulfurase Q96EN8 Details Multifunctional protein ADE2 P22234 Details CAD protein P27708 Details Protein farnesyltransferase/geranylgeranyltransferase type-1 subunit alpha P49354 Details DNA topoisomerase 2-beta Q02880 Details Ribonucleoside-diphosphate reductase subunit M2 P31350 Details NAD-dependent protein deacetylase sirtuin-1 Q96EB6 Details Gephyrin Q9NQX3 Details Focal adhesion kinase 1 Q05397 Details Integrin beta-4 P16144 Details Far upstream element-binding protein 1 Q96AE4 Details Heat shock protein beta-1 P04792 Details Ribosome-binding protein 1 Q9P2E9 Details Microtubule-associated protein 4 P27816 Details Nuclear receptor coactivator 2 Q15596 Details Proteasome subunit alpha type-3 P25788 Details Proteasome subunit alpha type-5 P28066 Details Ribosomal protein S6 kinase alpha-1 Q15418 Details SRSF protein kinase 2 P78362 Details Glycylpeptide N-tetradecanoyltransferase 1 P30419 Details Mediator of RNA polymerase II transcription subunit 1 Q15648 Details Epsin-1 Q9Y6I3 Details Vimentin P08670 Details Uridine-cytidine kinase-like 1 Q9NWZ5 Details Hepatocyte growth factor-regulated tyrosine kinase substrate O14964 Details Ribosomal protein S6 kinase alpha-3 P51812 Details Dihydropyrimidinase-related protein 2 Q16555 Details Nucleolar and coiled-body phosphoprotein 1 Q14978 Details 60 kDa heat shock protein, mitochondrial P10809 Details Hepatoma-derived growth factor P51858 Details Cyclin-dependent kinase inhibitor 1 P38936 Details Heterogeneous nuclear ribonucleoprotein K P61978 Details Far upstream element-binding protein 2 Q92945 Details Cysteine and glycine-rich protein 1 P21291 Details Filamin-A P21333 Details 60S ribosomal protein L18 Q07020 Details Splicing factor 1 Q15637 Details Zyxin Q15942 Details Myosin-9 P35579 Details Splicing factor 3B subunit 1 O75533 Details Serine/threonine-protein kinase TAO1 Q7L7X3 Details AP2-associated protein kinase 1 Q2M2I8 Details Casein kinase I isoform alpha P48729 Details Mitogen-activated protein kinase kinase kinase 11 Q16584 Details Serine/threonine-protein kinase MARK2 Q7KZI7 Details Serine/threonine-protein kinase Nek9 Q8TD19 Details Serine/threonine-protein kinase PAK 1 Q13153 Details Serine/threonine-protein kinase N1 Q16512 Details Serine/threonine-protein kinase N2 Q16513 Details Serine/threonine-protein kinase 10 O94804 Details Histone-lysine N-methyltransferase EZH2 Q15910 Details Exportin-1 O14980 Details POU domain, class 2, transcription factor 1 P14859 Details Tyrosine-protein phosphatase non-receptor type 12 Q05209 Details Molybdopterin synthase catalytic subunit O96007 Details Proliferation marker protein Ki-67 P46013 Details