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/ HDM2-binding partners: interaction with translation elongation factor EF1alpha. HDM2-binding partners: interaction with translation elongation factor EF1alpha. J. Proteome Res. (1535-3893) 2007 imex curation Frum R., Busby S.A., Ramamoorthy M., Deb S., Shabanowitz J., Hunt D.F., Deb S.P. Apoptosis - Interactions involving proteins with a function related to apoptosis Cancer - Interactions investigated in the context of cancer HDM2 was overexpressed in H1299 cells. Extracts were subjected to immunoprecipitation with anti-HDM2 antibody directly coupled with agarose. Proteins eluted from HDM2 immunoprecipitations were digested with trypsin and analysed by nanospray HPLC mass spectrometry. spdeb@vcu.edu 29-OCT-2007: Contacted by JYOTI. Only protein-protein interactions 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. anti bait coip anti bait coimmunoprecipitation anti bait coip ms participant Identification by mass spectrometry ms participant Frum R., Busby S.A., Ramamoorthy M., Deb S., Shabanowitz J., Hunt D.F., Deb S.P. Apoptosis - Interactions involving proteins with a function related to apoptosis HDM2 was overexpressed in H1299 cells. Extracts were subjected to immunoprecipitation with anti-HDM2 antibody directly coupled with agarose. Proteins eluted from HDM2 immunoprecipitations were digested with trypsin and analysed by nanospray HPLC mass spectrometry. spdeb@vcu.edu Accepted 2007-JUN-10 by JYOTI. J. Proteome Res. (1535-3893) 2007 Only protein-protein interactions imex curation HDM2-binding partners: interaction with translation elongation factor EF1alpha. HDM2-binding partners: interaction with translation elongation factor EF1alpha. J. Proteome Res. (1535-3893) 2007 imex curation Frum R., Busby S.A., Ramamoorthy M., Deb S., Shabanowitz J., Hunt D.F., Deb S.P. Apoptosis - Interactions involving proteins with a function related to apoptosis Cancer - Interactions investigated in the context of cancer HDM2 was overexpressed in H1299 cells. Extracts were subjected to immunoprecipitation with anti-HDM2 antibody directly coupled with agarose. Proteins eluted from HDM2 immunoprecipitations were digested with trypsin and analysed by nanospray HPLC mass spectrometry. spdeb@vcu.edu 29-OCT-2007: Contacted by JYOTI. Only protein-protein interactions human-sjsa_1 Homo sapiens SJSA-1 (formerly OsA-CL) osteosarcoma cells sjsa_1 SJSA-1 (formerly OsA-CL) osteosarcoma cells The SJSA-1 cell line (formerly OsA-CL) was established in 1982 from the primary tumor of a patient diagnosed with primitive multipotential sarcoma of the femur. There is amplification of the gene which encodes the p53 associated protein, MDM2. SJSA-1 cells also exhibit a 15 fold amplification of the gli proto-oncogene. anti bait coip anti bait coimmunoprecipitation anti bait coip weight autoradiogra molecular weight estimation by autoradiography weight autoradiogra Frum R., Busby S.A., Ramamoorthy M., Deb S., Shabanowitz J., Hunt D.F., Deb S.P. Apoptosis - Interactions involving proteins with a function related to apoptosis HDM2 was overexpressed in H1299 cells. Extracts were subjected to immunoprecipitation with anti-HDM2 antibody directly coupled with agarose. Proteins eluted from HDM2 immunoprecipitations were digested with trypsin and analysed by nanospray HPLC mass spectrometry. spdeb@vcu.edu Accepted 2007-JUN-10 by JYOTI. 29-OCT-2007: Contacted by JYOTI. J. Proteome Res. (1535-3893) 2007 Only protein-protein interactions imex curation HDM2-binding partners: interaction with translation elongation factor EF1alpha. HDM2-binding partners: interaction with translation elongation factor EF1alpha. J. Proteome Res. (1535-3893) 2007 imex curation Frum R., Busby S.A., Ramamoorthy M., Deb S., Shabanowitz J., Hunt D.F., Deb S.P. Apoptosis - Interactions involving proteins with a function related to apoptosis Cancer - Interactions investigated in the context of cancer HDM2 was overexpressed in H1299 cells. Extracts were subjected to immunoprecipitation with anti-HDM2 antibody directly coupled with agarose. Proteins eluted from HDM2 immunoprecipitations were digested with trypsin and analysed by nanospray HPLC mass spectrometry. spdeb@vcu.edu 29-OCT-2007: Contacted by JYOTI. Only protein-protein interactions human-wi_38 Homo sapiens Caucasian fetal lung fibroblast cell line wi_38 WI39 human Caucasian fetal lung fibroblast cell line From normal embryonic lung of female. Have very broad human virus spectra, especially useful for isolation of rhinoviruses. Cells form multilayered membrane when held for long periods at 37C with periodic pH adjustments. Used in preparation of human virus vaccines. 2n = 46, diploid except at high passage number confocal microscopy confocal microscopy antibody detection identification by antibody antibody detection Frum R., Busby S.A., Ramamoorthy M., Deb S., Shabanowitz J., Hunt D.F., Deb S.P. Apoptosis - Interactions involving proteins with a function related to apoptosis HDM2 was overexpressed in H1299 cells. Extracts were subjected to immunoprecipitation with anti-HDM2 antibody directly coupled with agarose. Proteins eluted from HDM2 immunoprecipitations were digested with trypsin and analysed by nanospray HPLC mass spectrometry. spdeb@vcu.edu Accepted 2007-JUN-10 by JYOTI. 29-OCT-2007: Contacted by JYOTI. J. Proteome Res. (1535-3893) 2007 Only protein-protein interactions imex curation HDM2-binding partners: interaction with translation elongation factor EF1alpha. HDM2-binding partners: interaction with translation elongation factor EF1alpha. J. Proteome Res. (1535-3893) 2007 imex curation Frum R., Busby S.A., Ramamoorthy M., Deb S., Shabanowitz J., Hunt D.F., Deb S.P. Apoptosis - Interactions involving proteins with a function related to apoptosis Cancer - Interactions investigated in the context of cancer HDM2 was overexpressed in H1299 cells. Extracts were subjected to immunoprecipitation with anti-HDM2 antibody directly coupled with agarose. Proteins eluted from HDM2 immunoprecipitations were digested with trypsin and analysed by nanospray HPLC mass spectrometry. spdeb@vcu.edu 29-OCT-2007: Contacted by JYOTI. Only protein-protein interactions 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. anti bait coip anti bait coimmunoprecipitation anti bait coip western blot western blot Immuno blot Immuno blot Frum R., Busby S.A., Ramamoorthy M., Deb S., Shabanowitz J., Hunt D.F., Deb S.P. Apoptosis - Interactions involving proteins with a function related to apoptosis HDM2 was overexpressed in H1299 cells. Extracts were subjected to immunoprecipitation with anti-HDM2 antibody directly coupled with agarose. Proteins eluted from HDM2 immunoprecipitations were digested with trypsin and analysed by nanospray HPLC mass spectrometry. spdeb@vcu.edu Accepted 2007-JUN-10 by JYOTI. 29-OCT-2007: Contacted by JYOTI. J. Proteome Res. (1535-3893) 2007 Only protein-protein interactions imex curation HDM2-binding partners: interaction with translation elongation factor EF1alpha. HDM2-binding partners: interaction with translation elongation factor EF1alpha. J. Proteome Res. (1535-3893) 2007 imex curation Frum R., Busby S.A., Ramamoorthy M., Deb S., Shabanowitz J., Hunt D.F., Deb S.P. Apoptosis - Interactions involving proteins with a function related to apoptosis Cancer - Interactions investigated in the context of cancer HDM2 was overexpressed in H1299 cells. Extracts were subjected to immunoprecipitation with anti-HDM2 antibody directly coupled with agarose. Proteins eluted from HDM2 immunoprecipitations were digested with trypsin and analysed by nanospray HPLC mass spectrometry. spdeb@vcu.edu 29-OCT-2007: Contacted by JYOTI. Only protein-protein interactions human-wi_38 Homo sapiens Caucasian fetal lung fibroblast cell line wi_38 WI39 human Caucasian fetal lung fibroblast cell line From normal embryonic lung of female. Have very broad human virus spectra, especially useful for isolation of rhinoviruses. Cells form multilayered membrane when held for long periods at 37C with periodic pH adjustments. Used in preparation of human virus vaccines. 2n = 46, diploid except at high passage number anti bait coip anti bait coimmunoprecipitation anti bait coip weight autoradiogra molecular weight estimation by autoradiography weight autoradiogra Frum R., Busby S.A., Ramamoorthy M., Deb S., Shabanowitz J., Hunt D.F., Deb S.P. Apoptosis - Interactions involving proteins with a function related to apoptosis HDM2 was overexpressed in H1299 cells. Extracts were subjected to immunoprecipitation with anti-HDM2 antibody directly coupled with agarose. Proteins eluted from HDM2 immunoprecipitations were digested with trypsin and analysed by nanospray HPLC mass spectrometry. spdeb@vcu.edu Accepted 2007-JUN-10 by JYOTI. 29-OCT-2007: Contacted by JYOTI. J. Proteome Res. (1535-3893) 2007 Only protein-protein interactions imex curation cdn2b_human Cyclin-dependent kinase 4 inhibitor B CDKN2B MTS2 p14-INK4b p15-INK4b Multiple tumor suppressor 2 protein protein human Homo sapiens Human MREENKGMPSGGGSDEGLASAAARGLVEKVRQLLEAGADPNGVNRFGRRAIQVMMMGSARVAELLLLHGAEPNCADPATLTRPVHDAAREGFLDTLVVLHRAGARLDVRDAWGRLPVDLAEERGHRDVAGYLRTATGD 0D6FFBDFA6FEAD21 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 DIP protein Q00987 original sequence version: 93 DIP protein Q00987-1 original sequence version: 192 DIP protein Q00987 original sequence version: 88 DIP protein Q00987 original sequence version: 180 DIP protein Q00987 original sequence version: 188 DIP protein Q00987 original sequence version: 189 DIP protein Q00987 original sequence version: 115 DIP protein Q00987-1 original sequence version: null DIP protein Q00987 original sequence version: 149 DIP protein Q00987 original sequence version: 146 DIP protein Q00987 original sequence version: 140 DIP protein Q00987 original sequence version: 183 DIP protein Q00987 original sequence version: 145 DIP protein Q00987 original sequence version: 182 DIP protein Q00987 original sequence version: 151 DIP protein Q00987 original sequence version: 156 DIP protein Q00987-1 original sequence version: 160 DIP protein Q00987 original sequence version: 160 DIP protein Q00987 original sequence version: 168 DIP protein Q00987 original sequence version: 186 DIP protein Q00987 original sequence version: 206 F37CE163876BC983 rl11_human 60S ribosomal protein L11 RPL11 CLL-associated antigen KW-12 Large ribosomal subunit protein uL5 protein protein human Homo sapiens Human MAQDQGEKENPMRELRIRKLCLNICVGESGDRLTRAAKVLEQLTGQTPVFSKARYTVRSFGIRRNEKIAVHCTVRGAKAEEILEKGLKVREYELRKNNFSDTGNFGFGIQEHIDLGIKYDPSIGIYGLDFYVVLGRPGFSIADKKRRTGCIGAKHRISKEEAMRWFQQKYDGIILPGK DIP protein P62913 original sequence version: null 26EC965C9239774E tba1c_human Tubulin alpha-1C chain TUBA1C TUBA6 Tubulin alpha-6 chain Alpha-tubulin 6 protein protein human Homo sapiens Human MRECISIHVGQAGVQIGNACWELYCLEHGIQPDGQMPSDKTIGGGDDSFNTFFSETGAGKHVPRAVFVDLEPTVIDEVRTGTYRQLFHPEQLITGKEDAANNYARGHYTIGKEIIDLVLDRIRKLADQCTGLQGFLVFHSFGGGTGSGFTSLLMERLSVDYGKKSKLEFSIYPAPQVSTAVVEPYNSILTTHTTLEHSDCAFMVDNEAIYDICRRNLDIERPTYTNLNRLISQIVSSITASLRFDGALNVDLTEFQTNLVPYPRIHFPLATYAPVISAEKAYHEQLTVAEITNACFEPANQMVKCDPRHGKYMACCLLYRGDVVPKDVNAAIATIKTKRTIQFVDWCPTGFKVGINYQPPTVVPGGDLAKVQRAVCMLSNTTAVAEAWARLDHKFDLMYAKRAFVHWYVGEGMEEGEFSEAREDMAALEKDYEEVGADSADGEDEGEEY 0CFE7212CB5E14F9 ef1a1_human Elongation factor 1-alpha 1 EEF1A1 EEF1A EF1A LENG7 Eukaryotic elongation factor 1 A-1 Elongation factor Tu Leukocyte receptor cluster member 7 protein protein human Homo sapiens Human MGKEKTHINIVVIGHVDSGKSTTTGHLIYKCGGIDKRTIEKFEKEAAEMGKGSFKYAWVLDKLKAERERGITIDISLWKFETSKYYVTIIDAPGHRDFIKNMITGTSQADCAVLIVAAGVGEFEAGISKNGQTREHALLAYTLGVKQLIVGVNKMDSTEPPYSQKRYEEIVKEVSTYIKKIGYNPDTVAFVPISGWNGDNMLEPSANMPWFKGWKVTRKDGNASGTTLLEALDCILPPTRPTDKPLRLPLQDVYKIGGIGTVPVGRVETGVLKPGMVVTFAPVNVTTEVKSVEMHHEALSEALPGDNVGFNVKNVSVKDVRRGNVAGDSKNDPPMEAAGFTAQVIILNHPGQISAGYAPVLDCHTAHIACKFAELKEKIDRRSGKKLEDGPKFLKSGDAAIVDMVPGKPMCVESFSDYPPLGRFAVRDMRQTVAVGVIKAVDKKAAGAGKVTKSAQKAQKAK DIP protein P68104 original sequence version: null DIP protein P68104 original sequence version: 134 D465615545AF686A rs20_human 40S ribosomal protein S20 RPS20 Small ribosomal subunit protein uS10 protein protein human Homo sapiens Human MAFKDTGKTPVEPEVAIHRIRITLTSRNVKSLEKVCADLIRGAKEKNLKVKGPVRMPTKTLRITTRKTPCGEGSKTWDRFQMRIHKRLIDLHSPSEIVKQITSISIEPGVEVEVTIADA DIP protein P60866 original sequence version: 112 8802DF7894ADDF94 k1c14_human Keratin, type I cytoskeletal 14 Cytokeratin-14 KRT14 Keratin-14 protein protein human Homo sapiens Human MTTCSRQFTSSSSMKGSCGIGGGIGGGSSRISSVLAGGSCRAPSTYGGGLSVSSSRFSSGGACGLGGGYGGGFSSSSSSFGSGFGGGYGGGLGAGLGGGFGGGFAGGDGLLVGSEKVTMQNLNDRLASYLDKVRALEEANADLEVKIRDWYQRQRPAEIKDYSPYFKTIEDLRNKILTATVDNANVLLQIDNARLAADDFRTKYETELNLRMSVEADINGLRRVLDELTLARADLEMQIESLKEELAYLKKNHEEEMNALRGQVGGDVNVEMDAAPGVDLSRILNEMRDQYEKMAEKNRKDAEEWFFTKTEELNREVATNSELVQSGKSEISELRRTMQNLEIELQSQLSMKASLENSLEETKGRYCMQLAQIQEMIGSVEEQLAQLRCEMEQQNQEYKILLDVKTRLEQEIATYRRLLEGEDAHLSSSQFSSGSQSSRDVTSSSRQIRTKVMDVHDGKVVSTHEQVLRTKN DIP protein P02533 original sequence version: 143 120BA30BA2F8E397 if90l_human_protein Chain L, Fab Fragment Of Monoclonal Antibody (Lnkb-2) Against Human Interleukin-2 In Complex With Antigenic Peptide protein protein mouse Mus musculus Mouse DVQMTQTPLTLSVTIGQPASISCESSQSLLYSNGKTYLNWLLQRPGQSPKRLIYLVSKLDSGVPDRFTGSGSGTDFTLRISRVEAEDLGVYYCVQGTHFPRTFGGGTKLEIKRADAAPTVSIFPPSSEQLTSGGASVVCFLNNFYPKDINVKWKIDGSERQNGVLNSWTDQDSKDSTYSMSSTLTLTKDEYERHNSYTCEATHKTSTSPIVKSFNRNEC An amino acid sequence which is not represented in UniProt 798BEFB420A5FBDB rl5_human 60S ribosomal protein L5 RPL5 Large ribosomal subunit protein uL18 MSTP030 protein protein human Homo sapiens Human MGFVKVVKNKAYFKRYQVKFRRRREGKTDYYARKRLVIQDKNKYNTPKYRMIVRVTNRDIICQIAYARIEGDMIVCAAYAHELPKYGVKVGLTNYAAAYCTGLLLARRLLNRFGMDKIYEGQVEVTGDEYNVESIDGQPGAFTCYLDAGLARTTTGNKVFGALKGAVDGGLSIPHSTKRFPGYDSESKEFNAEVHRKHIMGQNVADYMRYLMEEDEDAYKKQFSQYIKNSVTPDMMEEMYKKAHAAIRENPVYEKKPKKEVKKKRWNRPKMSLAQKKDRVAQKKASFLRAQERAAES DIP protein P46777 original sequence version: 147 DIP protein P46777 original sequence version: 109 07CA9FBA842BD2A3 dcd_human Dermcidin AIDD DSEP Preproteolysin DCD protein protein human Homo sapiens Human MRFMTLLFLTALAGALVCAYDPEAASAPGSGNPCHEASAAQKENAGEDPGLARQAPKPRKQRSSLLEKGLDGAKKAVGGLGKLGKDAVEDLESVGKGAVHDVKDVLDSVL 23666FBD20019465 tbb1_human Tubulin beta-1 chain TUBB1 protein protein human Homo sapiens Human MREIVHIQIGQCGNQIGAKFWEMIGEEHGIDLAGSDRGASALQLERISVYYNEAYGRKYVPRAVLVDLEPGTMDSIRSSKLGALFQPDSFVHGNSGAGNNWAKGHYTEGAELIENVLEVVRHESESCDCLQGFQIVHSLGGGTGSGMGTLLMNKIREEYPDRIMNSFSVMPSPKVSDTVVEPYNAVLSIHQLIENADACFCIDNEALYDICFRTLKLTTPTYGDLNHLVSLTMSGITTSLRFPGQLNADLRKLAVNMVPFPRLHFFMPGFAPLTAQGSQQYRALSVAELTQQMFDARNTMAACDLRRGRYLTVACIFRGKMSTKEVDQQLLSVQTRNSSCFVEWIPNNVKVAVCDIPPRGLSMAATFIGNNTAIQEIFNRVSEHFSAMFKRKAFVHWYTSEGMDINEFGEAENNIHDLVSEYQQFQDAKAVLEEDEEVTEEAEMEPEDKGH DIP protein Q9H4B7 original sequence version: 109 6A3E5208C1C89AE4 g3pt_human Glyceraldehyde-3-phosphate dehydrogenase, testis-specific GAPDHS GAPD2 GAPDH2 GAPDS HSD35 HSD-35 Spermatogenic glyceraldehyde-3-phosphate dehydrogenase Spermatogenic cell-specific glyceraldehyde 3-phosphate dehydrogenase 2 protein protein human Homo sapiens Human MSKRDIVLTNVTVVQLLRQPCPVTRAPPPPEPKAEVEPQPQPEPTPVREEIKPPPPPLPPHPATPPPKMVSVARELTVGINGFGRIGRLVLRACMEKGVKVVAVNDPFIDPEYMVYMFKYDSTHGRYKGSVEFRNGQLVVDNHEISVYQCKEPKQIPWRAVGSPYVVESTGVYLSIQAASDHISAGAQRVVISAPSPDAPMFVMGVNENDYNPGSMNIVSNASCTTNCLAPLAKVIHERFGIVEGLMTTVHSYTATQKTVDGPSRKAWRDGRGAHQNIIPASTGAAKAVTKVIPELKGKLTGMAFRVPTPDVSVVDLTCRLAQPAPYSAIKEAVKAAAKGPMAGILAYTEDEVVSTDFLGDTHSSIFDAKAGIALNDNFVKLISWYDNEYGYSHRVVDLLRYMFSRDK 301F71C768CD95D8 mdm2-cdkn2b 1 16 unspecified role unspecified role prey prey 11 unspecified role unspecified role prey prey 15 unspecified role unspecified role prey prey 13 unspecified role unspecified role prey prey 9 unspecified role unspecified role prey prey 8 unspecified role unspecified role prey prey 6 unspecified role unspecified role prey prey 14 unspecified role unspecified role prey prey 17 unspecified role unspecified role prey prey 10 unspecified role unspecified role prey prey 7 unspecified role unspecified role bait bait association association Table 1, 1 B Table 1A number of proteins were conclusively identified in the HDM2 immunoprecipitation fractions in addition to the bait protein HDM2 itself. The protein identified with the most number of unique peptide hits was the eukaryotic elongation factor EF1R, and this result was confirmed by manual examination of spectra identified as EF1R Q00987 transfection Q00987 over-expressed BAIT protein gi266516; PREY protein gi4503571 The author identifier for the PREY protein does not correspond to eukaryotic elongation factor 1. mdm2-if90l_human_pro 1 13 unspecified role unspecified role prey prey 12 unspecified role unspecified role prey prey 7 unspecified role unspecified role bait bait association association Table 1 Q00987 transfection Q00987 over-expressed BAIT protein gi266516; PREY protein gi1752236 eef1a1-mdm2 4 10 unspecified role unspecified role bait bait 7 unspecified role unspecified role prey prey physical association physical association 2 A Q00987 transfection BAIT protein gi266516; PREY protein gi4503571 The author identifier for the PREY protein does not correspond to eukaryotic elongation factor 1. The results (Figure 2A) show that anti-EF1R antibody coimmunoprecipitates HDM2 from HDM2 expression plasmid transfected cell extracts mdm2-eef1a1-1 2 10 unspecified role unspecified role prey prey region 35s radiolabel 35s radiolabel 35S s35 radiolabelled S35 s35 radiolabelled 35S S35 undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined 7 unspecified role unspecified role bait bait region 35s radiolabel 35s radiolabel 35S s35 radiolabelled S35 s35 radiolabelled 35S S35 undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined physical association physical association 2 B The data show that anti-HDM2 rabbit antiserum coimmunoprecipitates EF1R (lane 2), whereas the preimmune serum cannot (lane 1). Q00987 over-expression mdm2-eef1a1-2 5 10 unspecified role unspecified role prey prey region 35s radiolabel 35s radiolabel 35S s35 radiolabelled S35 s35 radiolabelled 35S S35 undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined region required to bind necessary binding region required to bind deletion analysis deletion analysis certain certain sequence position certain certain certain sequence position certain 7 unspecified role unspecified role bait bait region 35s radiolabel 35s radiolabel 35S s35 radiolabelled S35 s35 radiolabelled 35S S35 undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined n-terminal required to bind necessary binding region required to bind deletion analysis deletion analysis certain certain sequence position certain certain certain sequence position certain physical association physical association 3 A, B, C Analysis of the immunoprecipitates by SDS polyacrylamide gel electrophoresis and autoradiography (Figure 3A-C) showed that anti-HDM2 antibody coimmunoprecipitates EF1R in the presence of HDM2 WI38 cells are not transformed. Q00987 transfection eef1a1-mdm2-1 5 7 unspecified role unspecified role prey prey region 35s radiolabel 35s radiolabel 35S s35 radiolabelled S35 s35 radiolabelled 35S S35 undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined 10 unspecified role unspecified role bait bait region 35s radiolabel 35s radiolabel 35S s35 radiolabelled S35 s35 radiolabelled 35S S35 undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined physical association physical association 3 B Analysis of the immunoprecipitates by SDS polyacrylamide gel electrophoresis and autoradiography (Figure 3A-C) showed that anti-HDM2 antibody coimmunoprecipitates EF1R in the presence of HDM2 WI38 cells are not transformed. Q00987 transfection eef1a1-mdm2-2 3 10 unspecified role unspecified role neutral component neutral component 7 unspecified role unspecified role neutral component neutral component physical association physical association WI38 cells are not transformed. Q00987 transfection Results of this experiment (Figure 4) show that although the HDM2 (detected by green fluorescence) is predominantly localized in the nucleus and EF1R (red fluorescence) in the cytoplasm, co-localization of HDM2 and EF1R can be detected (yellow fluorescence) in the cytoplasm.