IntAct European Bioinformatics Institute IntAct intact http://www.ebi.ac.uk/ http://www.ebi.ac.uk/intact/query/${ac} EBI-[0-9]+|IA:[0-9]+ INTerAction database (IntAct) provides an open source database and toolkit for the storage, presentation and analysis of molecular interactions. http://www.ebi.ac.uk/intact European Bioinformatics Institute; Wellcome Trust Genome Campus; Hinxton, Cambridge; CB10 1SD; United Kingdom http://www.ebi.ac.uk/intact/ http://www.ebi.ac.uk/ yadavilli-2009-3 Ribosomal protein S3: A multi-functional protein that interacts with both p53 and MDM2 through its KH domain. in vitro In vitro pull down pull down anti tag western anti tag western blot anti tag western Yadavilli S., Mayo L.D., Higgins M., Lain S., Hegde V., Deutsch W.A. Affinomics - Interactions curated for the Affinomics consortium. His-p53, His-MDM2 and GST-RPS3 were found to be expressed in E.coli and MDM2 tags were N-terminal. (From Reference 11 -Mayo et al., 1997; Ref 16 Hegde at al.,2001.) deutscwa@pbrc.edu DNA Repair (Amst.) (1568-7864) 2009 Only protein-protein interactions imex curation yadavilli-2009-4 Ribosomal protein S3: A multi-functional protein that interacts with both p53 and MDM2 through its KH domain. in vitro In vitro pull down pull down western blot western blot Immuno blot Immuno blot Yadavilli S., Mayo L.D., Higgins M., Lain S., Hegde V., Deutsch W.A. Affinomics - Interactions curated for the Affinomics consortium. His-p53, His-MDM2 and GST-RPS3 were found to be expressed in E.coli and MDM2 tags were N-terminal. (From Reference 11 -Mayo et al., 1997; Ref 16 Hegde at al.,2001.) Figure 5 interactions from a "8-oxodg oligonucleotide pull-down assay" have not been included as the sequence of the 8-oxodg oligonucleotide is not given. deutscwa@pbrc.edu DNA Repair (Amst.) (1568-7864) 2009 Only protein-protein interactions imex curation yadavilli-2009-5 Ribosomal protein S3: A multi-functional protein that interacts with both p53 and MDM2 through its KH domain. human-293 Homo sapiens transformed primary embryonal kidney cells Human 293 Transformed human primary embryonal kidney cells. The Cell type named 293 is synonymous with HEK-293. fret fluorescent resonance energy transfer FRET RET FRET analysis FRET RET FRET analysis fret tag fluorescence tag visualisation by fluorescence tag fluorescence Yadavilli S., Mayo L.D., Higgins M., Lain S., Hegde V., Deutsch W.A. Affinomics - Interactions curated for the Affinomics consortium. deutscwa@pbrc.edu DNA Repair (Amst.) (1568-7864) 2009 Only protein-protein interactions imex curation yadavilli-2009-6 Ribosomal protein S3: A multi-functional protein that interacts with both p53 and MDM2 through its KH domain. human-293 Homo sapiens transformed primary embryonal kidney cells Human 293 Transformed human primary embryonal kidney cells. The Cell type named 293 is synonymous with HEK-293. pla proximity ligation assay pELISA pELISA PLA antibody detection identification by antibody antibody detection Yadavilli S., Mayo L.D., Higgins M., Lain S., Hegde V., Deutsch W.A. Affinomics - Interactions curated for the Affinomics consortium. deutscwa@pbrc.edu DNA Repair (Amst.) (1568-7864) 2009 Only protein-protein interactions imex curation yadavilli-2009-7 Ribosomal protein S3: A multi-functional protein that interacts with both p53 and MDM2 through its KH domain. in vitro In vitro ubiquitinase assay ubiquitinase assay ubiquitinase assay western blot western blot Immuno blot Immuno blot Yadavilli S., Mayo L.D., Higgins M., Lain S., Hegde V., Deutsch W.A. Affinomics - Interactions curated for the Affinomics consortium. E1 and E2 enzymes were present in these assays as well, but no origins are provided, so these proteins cannot be entered as participants. deutscwa@pbrc.edu DNA Repair (Amst.) (1568-7864) 2009 Only protein-protein interactions imex curation yadavilli-2009-1 Ribosomal protein S3: A multi-functional protein that interacts with both p53 and MDM2 through its KH domain. in vitro In vitro pull down pull down fingerprinting peptide massfingerprinting fingerprinting Yadavilli S., Mayo L.D., Higgins M., Lain S., Hegde V., Deutsch W.A. Affinomics - Interactions curated for the Affinomics consortium. deutscwa@pbrc.edu DNA Repair (Amst.) (1568-7864) 2009 Only protein-protein interactions imex curation yadavilli-2009-2 Ribosomal protein S3: A multi-functional protein that interacts with both p53 and MDM2 through its KH domain. human-u2-os Human U2-OS osteosarcoma cell line u2-os U2-OS osteocaroma cell line Not in CAPRI and no primary literature reference anti bait coip anti bait coimmunoprecipitation anti bait coip anti tag western anti tag western blot anti tag western Yadavilli S., Mayo L.D., Higgins M., Lain S., Hegde V., Deutsch W.A. Affinomics - Interactions curated for the Affinomics consortium. deutscwa@pbrc.edu DNA Repair (Amst.) (1568-7864) 2009 Only protein-protein interactions imex curation mdm2_human E3 ubiquitin-protein ligase Mdm2 MDM2 p53-binding protein Mdm2 Oncoprotein Mdm2 Double minute 2 protein RING-type E3 ubiquitin transferase Mdm2 protein protein human Homo sapiens Human MCNTNMSVPTDGAVTTSQIPASEQETLVRPKPLLLKLLKSVGAQKDTYTMKEVLFYLGQYIMTKRLYDEKQQHIVYCSNDLLGDLFGVPSFSVKEHRKIYTMIYRNLVVVNQQESSDSGTSVSENRCHLEGGSDQKDLVQELQEEKPSSSHLVSRPSTSSRRRAISETEENSDELSGERQRKRHKSDSISLSFDESLALCVIREICCERSSSSESTGTPSNPDLDAGVSEHSGDWLDQDSVSDQFSVEFEVESLDSEDYSLSEEGQELSDEDDEVYQVTVYQAGESDTDSFEEDPEISLADYWKCTSCNEMNPPLPSHCNRCWALRENWLPEDKGKDKGEISEKAKLENSTQAEEGFDVPDCKKTIVNDSRESCVEENDDKITQASQSQESEDYSQPSTSSSIIYSSQEDVKEFEREETQDKEESVESSLPLNAIEPCVICQGRPKNGCIVHGKTGHLMACFTCAKKLKKRNKPCPVCRQPIQMIVLTYFP rs3_human 40S ribosomal protein S3 RPS3 Small ribosomal subunit protein uS3 OK/SW-cl.26 protein protein human Homo sapiens Human MAVQISKKRKFVADGIFKAELNEFLTRELAEDGYSGVEVRVTPTRTEIIILATRTQNVLGEKGRRIRELTAVVQKRFGFPEGSVELYAEKVATRGLCAIAQAESLRYKLLGGLAVRRACYGVLRFIMESGAKGCEVVVSGKLRGQRAKSMKFVDGLMIHSGDPVNYYVDTAVRHVLLRQGVLGIKVKIMLPWDPTGKIGPKKPLPDHVSIVEPKDEILPTTPISEQKGGKPEPPAMPQPVPTA p53_human Cellular tumor antigen p53 TP53 P53 Tumor suppressor p53 Phosphoprotein p53 Antigen NY-CO-13 protein protein human Homo sapiens Human MEEPQSDPSVEPPLSQETFSDLWKLLPENNVLSPLPSQAMDDLMLSPDDIEQWFTEDPGPDEAPRMPEAAPPVAPAPAAPTPAAPAPAPSWPLSSSVPSQKTYQGSYGFRLGFLHSGTAKSVTCTYSPALNKMFCQLAKTCPVQLWVDSTPPPGTRVRAMAIYKQSQHMTEVVRRCPHHERCSDSDGLAPPQHLIRVEGNLRVEYLDDRNTFRHSVVVPYEPPEVGSDCTTIHYNYMCNSSCMGGMNRRPILTIITLEDSSGNLLGRNSFEVRVCACPGRDRRTEEENLRKKGEPHHELPPGSTKRALPNNTSSSPQPKKKPLDGEYFTLQIRGRERFEMFRELNEALELKDAQAGKEPGGSRAHSSHLKSKKGQSTSRHKKLMFKTEGPDSD rs20_human 40S ribosomal protein S20 RPS20 Small ribosomal subunit protein uS10 protein protein human Homo sapiens Human MAFKDTGKTPVEPEVAIHRIRITLTSRNVKSLEKVCADLIRGAKEKNLKVKGPVRMPTKTLRITTRKTPCGEGSKTWDRFQMRIHKRLIDLHSPSEIVKQITSISIEPGVEVEVTIADA mdm2-rps20-2 1532964 n/a 1532971 unspecified role unspecified role bait bait n_terminus his tag his tag Hexa-His-tag 6-His-tag Histidine-tag Hexa-His-tag Histidine-tag 6-His-tag n-term range n-terminal range n-term range n-term range n-terminal range n-term range false ecoli Escherichia coli (strain K12) n/a 1532972 unspecified role unspecified role prey prey region gst tag glutathione s tranferase tag glutathione S-tranferase tag glutathione S-tranferase tag gst tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined false ecoli Escherichia coli (strain K12) direct interaction direct interaction false false false Figure 2A mdm2-rps3-1 1532964 n/a 1532972 unspecified role unspecified role prey prey region gst tag glutathione s tranferase tag glutathione S-tranferase tag glutathione S-tranferase tag gst tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined false ecoli Escherichia coli (strain K12) n/a 1532973 unspecified role unspecified role bait bait region his tag his tag Hexa-His-tag 6-His-tag Histidine-tag Hexa-His-tag Histidine-tag 6-His-tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined false ecoli Escherichia coli (strain K12) direct interaction direct interaction false false false Figure 2B mdm2-rps20-3 1532965 n/a 1532972 unspecified role unspecified role prey prey human-h1299 Homo sapiens lung lymph node carcinoma h1299 Human lung lymph node carcinoma p53 null. These cells have also been reported to secrete the peptide hormone neuromedin B (NMB), but not gastrin releasing peptide (GRP). Also known as NCI H1299 or CRL-5803. http://www.lgcpromochem.com/atcc/ ATCC number: CRL-5803. ATCC number of the cell line is CRL-5803. n/a 1532971 unspecified role unspecified role bait bait region sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain false Q00987:p.Glu250_Glu252delinsAlaAlaAla mutation disrupting strength mutation disrupting interaction strength mutation disrupting strength certain certain sequence position certain certain certain sequence position certain false n_terminus biotin tag biotin tag n-term range n-terminal range n-term range n-term range n-terminal range n-term range false chemical synthesis Chemical synthesis (Chemical synthesis) physical association physical association false false false When full length MDM2 possessing the acidic domain mutation was used as bait, binding to RPS3 was not affected, suggesting the presence of another binding site outside of the acidic domain. Figure 1C -RHS rps3-mdm2-1 1532965 n/a 1532973 unspecified role unspecified role prey prey region his tag his tag Hexa-His-tag 6-His-tag Histidine-tag Hexa-His-tag Histidine-tag 6-His-tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined false human-293 Homo sapiens transformed primary embryonal kidney cells Human 293 Transformed human primary embryonal kidney cells. The Cell type named 293 is synonymous with HEK-293. n/a 1532972 unspecified role unspecified role bait bait region gst tag glutathione s tranferase tag glutathione S-tranferase tag glutathione S-tranferase tag gst tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined false k_homology_domain required to bind necessary binding region required to bind certain certain sequence position certain certain certain sequence position certain false ecoli Escherichia coli (strain K12) n/a 1532971 unspecified role unspecified role prey prey human-293 Homo sapiens transformed primary embryonal kidney cells Human 293 Transformed human primary embryonal kidney cells. The Cell type named 293 is synonymous with HEK-293. association association false false false Figure 2C mdm2-rps3-2 1532966 n/a 1532971 unspecified role unspecified role fluorescence donor fluorescence donor n_terminus cfp tag cyan fluorescent protein tag CFP cyan fluorescent protein cyan fluorescent protein CFP cfp tag n-term range n-terminal range n-term range n-term range n-terminal range n-term range false n/a 1532972 unspecified role unspecified role fluorescence accept fluorescence acceptor fluorescence accept n_terminus yfp tag yellow fluorescent protein tag YFP yellow fluorescent protein yellow fluorescent protein YFP yfp tag n-term range n-terminal range n-term range n-term range n-terminal range n-term range false proximity proximity close-contact colocalization neighbourhood interaction false false false Figure 3 oxidative stress i.e.hydrogen peroxide - but only in the nucleus. No interaction was observed in the nucleus for the phosphorylation-defective mutant YFP-T42A, as well as a lack of interaction in the cytoplasm for the phosphorylation mimic YFP-T42D. p53-rps3-3 1532966 n/a 1532972 unspecified role unspecified role fluorescence accept fluorescence acceptor fluorescence accept n_terminus yfp tag yellow fluorescent protein tag YFP yellow fluorescent protein yellow fluorescent protein YFP yfp tag n-term range n-terminal range n-term range n-term range n-terminal range n-term range false n/a 1532973 unspecified role unspecified role fluorescence donor fluorescence donor n_terminus cfp tag cyan fluorescent protein tag CFP cyan fluorescent protein cyan fluorescent protein CFP cfp tag n-term range n-terminal range n-term range n-term range n-terminal range n-term range false proximity proximity close-contact colocalization neighbourhood interaction false false false Figure 3 oxidative stress i.e. hydrogen peroxide Nutlin-3 -a cis-imidazoline analog which inhibits the interaction between mdm2 and tumour suppressor p53, but as shown here, it inhibits only under conditions of oxidative stress. The RPS3 mutant -T42D no longer was able to interact with cytoplasmic CFP-p53, but did maintain the ability to interact with nuclear CFP-p53. RPS3 mutant YFP-T42A only interacted with cytoplasmic CFP-p53. rps3-tp53 1532967 n/a 1532972 unspecified role unspecified role neutral component neutral component n/a 1532973 unspecified role unspecified role neutral component neutral component proximity proximity close-contact colocalization neighbourhood interaction false false false Figure 4 conditions of oxidative stress- hydrogen peroxide rps3-tp53-1 1532967 n/a 1532972 unspecified role unspecified role neutral component neutral component n/a 1532971 unspecified role unspecified role neutral component neutral component proximity proximity close-contact colocalization neighbourhood interaction false false false Figure 4 conditions of oxidative stress- hydrogen peroxide mdm2-tp53-22 1532968 n/a 1532971 enzyme enzyme neutral component neutral component Q00987:p.Cys464Ser mutation disrupting rate mutation disrupting interaction rate mutation disrupting rate certain certain sequence position certain certain certain sequence position certain false ecoli Escherichia coli (strain K12) n/a 1532973 enzyme target enzyme target substrate substrate neutral component neutral component region his tag his tag Hexa-His-tag 6-His-tag Histidine-tag Hexa-His-tag Histidine-tag 6-His-tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined false ecoli Escherichia coli (strain K12) ubiquitination ubiquitination reaction ubiquitination false false false E1 and E2 enzymes were present in these assays as well, but no origins are provided, so these proteins cannot be entered as participants. Figure 6 mdm2-rps3-3 1532968 n/a 1532972 competitor competitor competitor neutral component neutral component region gst tag glutathione s tranferase tag glutathione S-tranferase tag glutathione S-tranferase tag gst tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined false ecoli Escherichia coli (strain K12) n/a 1532973 enzyme target enzyme target substrate substrate neutral component neutral component region his tag his tag Hexa-His-tag 6-His-tag Histidine-tag Hexa-His-tag Histidine-tag 6-His-tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined false ecoli Escherichia coli (strain K12) n/a 1532971 enzyme enzyme neutral component neutral component Q00987:p.Cys464Ser mutation disrupting rate mutation disrupting interaction rate mutation disrupting rate certain certain sequence position certain certain certain sequence position certain false ecoli Escherichia coli (strain K12) ubiquitination ubiquitination reaction ubiquitination false false false E1 and E2 enzymes were present in these assays as well, but no origins are provided, so these proteins cannot be entered as participants. Figure 6i mdm2-rps20 1532969 n/a 1532972 unspecified role unspecified role prey prey human-h1299 Homo sapiens lung lymph node carcinoma h1299 Human lung lymph node carcinoma p53 null. These cells have also been reported to secrete the peptide hormone neuromedin B (NMB), but not gastrin releasing peptide (GRP). Also known as NCI H1299 or CRL-5803. http://www.lgcpromochem.com/atcc/ ATCC number: CRL-5803. ATCC number of the cell line is CRL-5803. n/a 1532974 unspecified role unspecified role prey prey human-h1299 Homo sapiens lung lymph node carcinoma h1299 Human lung lymph node carcinoma p53 null. These cells have also been reported to secrete the peptide hormone neuromedin B (NMB), but not gastrin releasing peptide (GRP). Also known as NCI H1299 or CRL-5803. http://www.lgcpromochem.com/atcc/ ATCC number: CRL-5803. ATCC number of the cell line is CRL-5803. n/a 1532971 unspecified role unspecified role bait bait Q00987:p.Glu250_Glu252delinsAlaAlaAla mutation disrupting strength mutation disrupting interaction strength mutation disrupting strength certain certain sequence position certain certain certain sequence position certain false n_terminus biotin tag biotin tag n-term range n-terminal range n-term range n-term range n-terminal range n-term range false region sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain false chemical synthesis Chemical synthesis (Chemical synthesis) association association false false false When full length MDM2 possessing the acidic domain mutation was used as bait, binding to RPS3 was not affected, suggesting the presence of another binding site outside of the acidic domain. Figure 1C mdm2-rps3 1532970 n/a 1532971 unspecified role unspecified role bait bait n/a 1532972 unspecified role unspecified role prey prey physical association physical association false false false Figure 1Ei When full length MDM2 possessing the acidic domain mutation was used as bait, binding to RPS3 was not affected, suggesting the presence of another binding site outside of the acidic domain.