Large-scale proteomics analysis of the human kinome.

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Oppermann FS, Gnad F, Olsen JV, Hornberger R, Greff Z, Keri G, Mann M, Daub H

Large-scale proteomics analysis of the human kinome.

Mol Cell Proteomics. 2009 Jul;8(7):1751-64. doi: 10.1074/mcp.M800588-MCP200. Epub 2009 Apr 15.

PubMed ID
19369195 [ View in PubMed
]
Abstract

Members of the human protein kinase superfamily are the major regulatory enzymes involved in the activity control of eukaryotic signal transduction pathways. As protein kinases reside at the nodes of phosphorylation-based signal transmission, comprehensive analysis of their cellular expression and site-specific phosphorylation can provide important insights into the architecture and functionality of signaling networks. However, in global proteome studies, low cellular abundance of protein kinases often results in rather minor peptide species that are occluded by a vast excess of peptides from other cellular proteins. These analytical limitations create a rationale for kinome-wide enrichment of protein kinases prior to mass spectrometry analysis. Here, we employed stable isotope labeling by amino acids in cell culture (SILAC) to compare the binding characteristics of three kinase-selective affinity resins by quantitative mass spectrometry. The evaluated pre-fractionation tools possessed pyrido[2,3-d]pyrimidine-based kinase inhibitors as immobilized capture ligands and retained considerable subsets of the human kinome. Based on these results, an affinity resin displaying the broadly selective kinase ligand VI16832 was employed to quantify the relative expression of more than 170 protein kinases across three different, SILAC-encoded cancer cell lines. These experiments demonstrated the feasibility of comparative kinome profiling in a compact experimental format. Interestingly, we found high levels of cytoplasmic and low levels of receptor tyrosine kinases in MV4-11 leukemia cells compared with the adherent cancer lines HCT116 and MDA-MB-435S. The VI16832 resin was further exploited to pre-fractionate kinases for targeted phosphoproteomics analysis, which revealed about 1200 distinct phosphorylation sites on more than 200 protein kinases. This hitherto largest survey of site-specific phosphorylation across the kinome significantly expands the basis for functional follow-up studies on protein kinase regulation. In conclusion, the straightforward experimental procedures described here enable different implementations of kinase-selective proteomics with considerable potential for future signal transduction and kinase drug target analysis.

DrugBank Data that Cites this Article

Polypeptides
NameUniProt ID
Tyrosine-protein kinase ABL1P00519Details
Cystine/glutamate transporterQ9UPY5Details
7-dehydrocholesterol reductaseQ9UBM7Details
Insulin receptorP06213Details
Pyridoxal kinaseO00764Details
Adenine phosphoribosyltransferaseP07741Details
Receptor-type tyrosine-protein kinase FLT3P36888Details
Ribosyldihydronicotinamide dehydrogenase [quinone]P16083Details
Large neutral amino acids transporter small subunit 1Q01650Details
5'-AMP-activated protein kinase subunit beta-1Q9Y478Details
Activin receptor type-1Q04771Details
Peroxisomal multifunctional enzyme type 2P51659Details
Tyrosine-protein kinase ABL2P42684Details
Mast/stem cell growth factor receptor KitP10721Details
Acetyl-CoA carboxylase 1Q13085Details
cAMP and cAMP-inhibited cGMP 3',5'-cyclic phosphodiesterase 10AQ9Y233Details
Monocarboxylate transporter 1P53985Details
5'-AMP-activated protein kinase catalytic subunit alpha-1Q13131Details
Fibroblast growth factor receptor 2P21802Details
Epidermal growth factor receptorP00533Details
Activated CDC42 kinase 1Q07912Details
5'-AMP-activated protein kinase subunit beta-2O43741Details
Proto-oncogene tyrosine-protein kinase SrcP12931Details
Tyrosine-protein kinase FynP06241Details
Tyrosine-protein kinase YesP07947Details
Ephrin type-A receptor 2P29317Details
Tyrosine-protein kinase LckP06239Details
Macrophage colony-stimulating factor 1 receptorP07333Details
Insulin-like growth factor 1 receptorP08069Details
Dual specificity mitogen-activated protein kinase kinase 1Q02750Details
Protein-tyrosine kinase 2-betaQ14289Details
Protein kinase C beta typeP05771Details
Cyclin-dependent kinase 5Q00535Details
Hepatocyte growth factor receptorP08581Details
Mitogen-activated protein kinase 1P28482Details
Phosphatidylinositol 3-kinase regulatory subunit alphaP27986Details
Fatty acid synthaseP49327Details
Insulin receptor substrate 1P35568Details
CalnexinP27824Details
Mitogen-activated protein kinase 14Q16539Details
Mitogen-activated protein kinase 3P27361Details
Glycogen synthase kinase-3 betaP49841Details
Cyclin-dependent kinase 2P24941Details
Cyclin-dependent kinase 1P06493Details
Phosphatidylserine synthase 1P48651Details
Epithelial discoidin domain-containing receptor 1Q08345Details
Inhibitor of nuclear factor kappa-B kinase subunit betaO14920Details
Thymidine kinase, cytosolicP04183Details
Tyrosine-protein kinase BTKQ06187Details
Tyrosine-protein kinase SYKP43405Details
Protein kinase C theta typeQ04759Details
Tyrosine-protein kinase HCKP08631Details
Dual specificity protein kinase CLK1P49759Details
TGF-beta receptor type-2P37173Details
Cyclin-dependent kinase 7P50613Details
cGMP-dependent 3',5'-cyclic phosphodiesteraseO00408Details
Keratin, type II cytoskeletal 8P05787Details
Cyclin-dependent kinase 9P50750Details
Cyclin-dependent kinase 6Q00534Details
Cyclin-dependent kinase 4P11802Details
Cyclin-dependent kinase 8P49336Details
Bifunctional phosphoribosylaminoimidazole carboxylase/phosphoribosylaminoimidazole succinocarboxamide synthetaseP22234Details
Phosphatidylserine synthase 2Q9BVG9Details
Phosphatidylinositol 3-kinase regulatory subunit betaO00459Details
DNA-dependent protein kinase catalytic subunitP78527Details
Serine/threonine-protein kinase mTORP42345Details
Phosphatidylinositol 4,5-bisphosphate 3-kinase catalytic subunit delta isoformO00329Details
Proto-oncogene tyrosine-protein kinase receptor RetP07949Details
Protein kinase C delta typeQ05655Details
Ephrin type-B receptor 4P54760Details
Prostaglandin E synthase 3Q15185Details
Deoxycytidine kinaseP27707Details
Focal adhesion kinase 1Q05397Details
Integrin beta-4P16144Details
Casein kinase I isoform gamma-2P78368Details
Serine/threonine-protein kinase Chk1O14757Details
Mitogen-activated protein kinase 11Q15759Details
Mitogen-activated protein kinase 13O15264Details
Aurora kinase BQ96GD4Details
cAMP-dependent protein kinase type II-alpha regulatory subunitP13861Details
cAMP-dependent protein kinase type I-alpha regulatory subunitP10644Details
Mitogen-activated protein kinase kinase kinase 1Q13233Details
Heat shock protein beta-1P04792Details
Microtubule-associated protein 4P27816Details
Tyrosine-protein kinase JAK3P52333Details
Casein kinase I isoform epsilonP49674Details
Cation-independent mannose-6-phosphate receptorP11717Details
Phosphatidylinositol 3-kinase catalytic subunit type 3Q8NEB9Details
Cyclin-A2P20248Details
Serine/threonine-protein kinase PLK1P53350Details
cAMP-dependent protein kinase catalytic subunit alphaP17612Details
Mitogen-activated protein kinase 9P45984Details
TGF-beta receptor type-1P36897Details
Mitogen-activated protein kinase 8P45983Details
Dual specificity protein kinase TTKP33981Details
Calcium/calmodulin-dependent protein kinase type II subunit betaQ13554Details
Serine/threonine-protein kinase Nek2P51955Details
Inner centromere proteinQ9NQS7Details
Large ribosomal subunit protein eL15P61313Details
Large ribosomal subunit protein uL3P39023Details
Serine/threonine-protein kinase/endoribonuclease IRE1O75460Details
Casein kinase I isoform gamma-3Q9Y6M4Details
Casein kinase II subunit alpha'P19784Details
STE20-like serine/threonine-protein kinaseQ9H2G2Details
Cyclin-dependent kinase 16Q00536Details
Serine/threonine-protein kinase Sgk1O00141Details
Calcium/calmodulin-dependent protein kinase type II subunit deltaQ13557Details
Serine/threonine-protein kinase 26Q9P289Details
Tyrosine-protein kinase JAK1P23458Details
Non-receptor tyrosine-protein kinase TYK2P29597Details
DestrinP60981Details
Cofilin-1P23528Details
Tyrosine-protein kinase MerQ12866Details
Ribosomal protein S6 kinase alpha-1Q15418Details
Tyrosine-protein kinase LynP07948Details
Serine/threonine-protein kinase 17BO94768Details
Mediator of RNA polymerase II transcription subunit 1Q15648Details
Calcium/calmodulin-dependent protein kinase type II subunit gammaQ13555Details
Mitogen-activated protein kinase kinase kinase 9P80192Details
Ephrin type-B receptor 2P29323Details
Transitional endoplasmic reticulum ATPaseP55072Details
Spectrin beta chain, non-erythrocytic 1Q01082Details
Ribosomal protein S6 kinase alpha-4O75676Details
Serine/threonine-protein kinase TAO2Q9UL54Details
Discoidin domain-containing receptor 2Q16832Details
Dual specificity mitogen-activated protein kinase kinase 2P36507Details
Mitogen-activated protein kinase kinase kinase 2Q9Y2U5Details
LIM domain kinase 1P53667Details
RalA-binding protein 1Q15311Details
Serine-protein kinase ATMQ13315Details
Ribosomal protein S6 kinase alpha-3P51812Details
Membrane-associated progesterone receptor component 1O00264Details
5'-AMP-activated protein kinase catalytic subunit alpha-2P54646Details
5'-AMP-activated protein kinase subunit gamma-2Q9UGJ0Details
Dual specificity calcium/calmodulin-dependent 3',5'-cyclic nucleotide phosphodiesterase 1CQ14123Details
Casein kinase II subunit betaP67870Details
Tyrosine-protein kinase FgrP09769Details
Mitogen-activated protein kinase kinase kinase 20Q9NYL2Details
Heterogeneous nuclear ribonucleoproteins A2/B1P22626Details
Heterogeneous nuclear ribonucleoprotein H3P31942Details
Hepatoma-derived growth factorP51858Details
Phosphatidylinositol 4-kinase type 2-betaQ8TCG2Details
Beclin-1Q14457Details
Heterogeneous nuclear ribonucleoprotein FP52597Details
Heat shock protein 105 kDaQ92598Details
DesmoplakinP15924Details
HornerinQ86YZ3Details
Far upstream element-binding protein 2Q92945Details
Heterogeneous nuclear ribonucleoprotein D0Q14103Details
NucleophosminP06748Details
Large ribosomal subunit protein eL14P50914Details
Small ribosomal subunit protein eS17P08708Details
Survival motor neuron proteinB4DP61Details
Myosin-9P35579Details
Bone morphogenetic protein receptor type-2Q13873Details
Serine/threonine-protein kinase 24Q9Y6E0Details
Serine/threonine-protein kinase 3Q13188Details
Tyrosine-protein kinase TecP42680Details
Serine/threonine-protein kinase TAO1Q7L7X3Details
Serine/threonine-protein kinase TAO3Q9H2K8Details
Serine/threonine-protein kinase TBK1Q9UHD2Details
Serine/threonine-protein kinase tousled-like 1Q9UKI8Details
Serine/threonine-protein kinase tousled-like 2Q86UE8Details
TRAF2 and NCK-interacting protein kinaseQ9UKE5Details
Non-receptor tyrosine-protein kinase TNK1Q13470Details
Serine/threonine-protein kinase ULK1O75385Details
AP2-associated protein kinase 1Q2M2I8Details
BMP-2-inducible protein kinaseQ9NSY1Details
Dual specificity protein kinase CLK2P49760Details
Dual specificity protein kinase CLK4Q9HAZ1Details
Casein kinase I isoform alphaP48729Details
eIF-2-alpha kinase GCN2Q9P2K8Details
Ephrin type-A receptor 1P21709Details
Ephrin type-A receptor 4P54764Details
Tyrosine-protein kinase FerP16591Details
Tyrosine-protein kinase Fes/FpsP07332Details
Cyclin-G-associated kinaseO14976Details
Glycogen synthase kinase-3 alphaP49840Details
Homeodomain-interacting protein kinase 2Q9H2X6Details
Serine/threonine-protein kinase ICKQ9UPZ9Details
Inhibitor of nuclear factor kappa-B kinase subunit epsilonQ14164Details
Interleukin-1 receptor-associated kinase 3Q9Y616Details
Serine/threonine-protein kinase SIK3Q9Y2K2Details
Serine/threonine-protein kinase LATS1O95835Details
LIM domain kinase 2P53671Details
Dual specificity mitogen-activated protein kinase kinase 5Q13163Details
Mitogen-activated protein kinase kinase kinase 10Q02779Details
Mitogen-activated protein kinase kinase kinase 11Q16584Details
Mitogen-activated protein kinase kinase kinase 12Q12852Details
Mitogen-activated protein kinase kinase kinase 15Q6ZN16Details
Mitogen-activated protein kinase kinase kinase 3Q99759Details
Mitogen-activated protein kinase kinase kinase 4Q9Y6R4Details
Mitogen-activated protein kinase kinase kinase 6O95382Details
Mitogen-activated protein kinase kinase kinase kinase 1Q92918Details
Mitogen-activated protein kinase kinase kinase kinase 2Q12851Details
Mitogen-activated protein kinase kinase kinase kinase 3Q8IVH8Details
Mitogen-activated protein kinase kinase kinase kinase 4O95819Details
Mitogen-activated protein kinase 15Q8TD08Details
Mitogen-activated protein kinase 7Q13164Details
Serine/threonine-protein kinase MARK1Q9P0L2Details
Serine/threonine-protein kinase MARK2Q7KZI7Details
MAP/microtubule affinity-regulating kinase 3P27448Details
MAP/microtubule affinity-regulating kinase 4Q96L34Details
Megakaryocyte-associated tyrosine-protein kinaseP42679Details
Maternal embryonic leucine zipper kinaseQ14680Details
Misshapen-like kinase 1Q8N4C8Details
Serine/threonine-protein kinase Nek1Q96PY6Details
Serine/threonine-protein kinase Nek3P51956Details
Serine/threonine-protein kinase Nek9Q8TD19Details
NUAK family SNF1-like kinase 1O60285Details
NUAK family SNF1-like kinase 2Q9H093Details
Serine/threonine-protein kinase PAK 4O96013Details
Cyclin-dependent kinase 17Q00537Details
Phosphatidylinositol 5-phosphate 4-kinase type-2 betaP78356Details
Serine/threonine-protein kinase N2Q16513Details
Serine/threonine-protein kinase PLK4O00444Details
Serine/threonine-protein kinase D1Q15139Details
Serine/threonine-protein kinase RIO2Q9BVS4Details
Receptor-interacting serine/threonine-protein kinase 2O43353Details
Serine/threonine-protein kinase SIK2Q9H0K1Details
Serine/threonine-protein kinase 10O94804Details
Serine/threonine-protein kinase 17AQ9UEE5Details
Mitogen-activated protein kinase 6Q16659Details
Plasma membrane ascorbate-dependent reductase CYBRD1Q53TN4Details
ATP-citrate synthaseP53396Details
Receptor tyrosine-protein kinase erbB-3P21860Details
Receptor protein-tyrosine kinaseQ504U8Details
Lysine-specific histone demethylase 1AO60341Details
Proliferation marker protein Ki-67P46013Details
Dual specificity mitogen-activated protein kinase kinase 4P45985Details