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/ The tetramer structure of the Nervy homology two domain, NHR2, is critical for AML1/ETO's activity. The tetramer structure of the Nervy homology two domain, NHR2, is critical for AML1/ETO's activity. Cancer Cell (1535-6108) 2006 imex curation Liu Y., Cheney MD., Gaudet JJ., Chruszcz M., Lukasik SM., Sugiyama D., Lary J., Cole J., Dauter Z., Minor W., Speck NA., Bushweller JH. Cancer - Interactions investigated in the context of cancer nancy.speck@dartmouth.edu 13-Nov-2007: Contacted by JYOTI. Only protein-protein interactions 3 in vitro In vitro x-ray diffraction x-ray crystallography X-ray X-ray x-ray diffraction predetermined predetermined participant predetermined Liu Y., Cheney MD., Gaudet JJ., Chruszcz M., Lukasik SM., Sugiyama D., Lary J., Cole J., Dauter Z., Minor W., Speck NA., Bushweller JH. Cancer - Interactions investigated in the context of cancer nancy.speck@dartmouth.edu Accepted 2006-OCT-23 by DAVET. Cancer Cell (1535-6108) 2006 Only protein-protein interactions 3 imex curation The tetramer structure of the Nervy homology two domain, NHR2, is critical for AML1/ETO's activity. The tetramer structure of the Nervy homology two domain, NHR2, is critical for AML1/ETO's activity. Cancer Cell (1535-6108) 2006 imex curation Liu Y., Cheney MD., Gaudet JJ., Chruszcz M., Lukasik SM., Sugiyama D., Lary J., Cole J., Dauter Z., Minor W., Speck NA., Bushweller JH. Cancer - Interactions investigated in the context of cancer nancy.speck@dartmouth.edu 13-Nov-2007: Contacted by JYOTI. Only protein-protein interactions 3 in vitro In vitro cosedimentation cosedimentation predetermined predetermined participant predetermined Liu Y., Cheney MD., Gaudet JJ., Chruszcz M., Lukasik SM., Sugiyama D., Lary J., Cole J., Dauter Z., Minor W., Speck NA., Bushweller JH. Cancer - Interactions investigated in the context of cancer Absorbance data 280 nm (NHR2, m4), 230 nm (m7, m2) were collected at a rotor speed of 50, 000 rpm at 20 C in aluminum-filled double sector cells at a scan interval of approximately 260 s. nancy.speck@dartmouth.edu Accepted 2006-OCT-23 by DAVET. 13-Nov-2007: Contacted by JYOTI. Cancer Cell (1535-6108) 2006 Only protein-protein interactions 3 imex curation The tetramer structure of the Nervy homology two domain, NHR2, is critical for AML1/ETO's activity. The tetramer structure of the Nervy homology two domain, NHR2, is critical for AML1/ETO's activity. Cancer Cell (1535-6108) 2006 imex curation Liu Y., Cheney MD., Gaudet JJ., Chruszcz M., Lukasik SM., Sugiyama D., Lary J., Cole J., Dauter Z., Minor W., Speck NA., Bushweller JH. Cancer - Interactions investigated in the context of cancer nancy.speck@dartmouth.edu 13-Nov-2007: Contacted by JYOTI. Only protein-protein interactions 3 in vitro In vitro molecular sieving molecular sieving Gel Filtration Size Exclusion Chromatography Size Exclusion Chromatography Gel Filtration Sizing column predetermined predetermined participant predetermined Liu Y., Cheney MD., Gaudet JJ., Chruszcz M., Lukasik SM., Sugiyama D., Lary J., Cole J., Dauter Z., Minor W., Speck NA., Bushweller JH. Cancer - Interactions investigated in the context of cancer The coimmunoprecipitations in Fig 5 could not be curated as the species of origin of all proteins is not given in the paper and the author did not reply to a request for information. nancy.speck@dartmouth.edu Accepted 2006-OCT-23 by DAVET. 13-Nov-2007: Contacted by JYOTI. Cancer Cell (1535-6108) 2006 Only protein-protein interactions 3 imex curation aml1_eto_chimera AML1-ETO chimera, human RUNX1(1-177)-N-MTG8(31-604) protein protein human Homo sapiens Human MRIPVDASTSRRFTPPSTALSPGKMSEALPLGAPDAGAALAGKLRSGDRSMVEVLADHPGELVRTDSPNFLCSVLPTHWRCNKTLPIAFKVVALGDVPDGTLVTVMAGNDENYSAELRNATAAMKNQVARFNDLRFVGRSGRGKSFTLTITVFTNPPQVATYHRAIKITVDGPREPRNRTEKHSTMPDSPVDVKTQSRLTPPTMPPPPTTQGAPRTSSFTPTTLTNGTSHSPTALNGAPSPPNGFSNGPSSSSSSSLANQQLPPACGARQLSKLKRFLTTLQQFGNDISPEIGERVRTLVLGLVNSTLTIEEFHSKLQEATNFPLRPFVIPFLKANLPLLQRELLHCARLAKQNPAQYLAQHEQLLLDASTTSPVDSSELLLDVNENGKRRTPDRTKENGFDREPLHSEHPSKRPCTISPGQRYSPNNGLSYQPNGLPHPTPPPPQHYRLDDMAIAHHYRDSYRHPSHRDLRDRNRPMGLHGTRQEEMIDHRLTDREWAEEWKHLDHLLNCIMDMVEKTRRSLTVLRRCQEADREELNYWIRRYSDAEDLKKGGGSSSSHSRQQSPVNPDPVALDAHREFLHRPASGYVPEEIWKKAEEAVNEVKRQAMTELQKAVSEAERKAHDMITTERAKMERTVAEAKRQAAEDALAVINQQEDSSESCWNCGRKASETCSGCNTARYCGSFCQHKDWEKHHHICGQTLQAQQQGDTPAVSSSVTPNSGAGSPMDTPPAATPRSTTPGTPSTIETTPR mtg8_human Protein CBFA2T1 Protein MTG8 Protein ETO Eight twenty one protein Cyclin-D-related protein Zinc finger MYND domain-containing protein 2 RUNX1T1 AML1T1 CBFA2T1 CDR ETO MTG8 ZMYND2 protein protein human Homo sapiens Human MISVKRNTWRALSLVIGDCRKKGNFEYCQDRTEKHSTMPDSPVDVKTQSRLTPPTMPPPPTTQGAPRTSSFTPTTLTNGTSHSPTALNGAPSPPNGFSNGPSSSSSSSLANQQLPPACGARQLSKLKRFLTTLQQFGNDISPEIGERVRTLVLGLVNSTLTIEEFHSKLQEATNFPLRPFVIPFLKANLPLLQRELLHCARLAKQNPAQYLAQHEQLLLDASTTSPVDSSELLLDVNENGKRRTPDRTKENGFDREPLHSEHPSKRPCTISPGQRYSPNNGLSYQPNGLPHPTPPPPQHYRLDDMAIAHHYRDSYRHPSHRDLRDRNRPMGLHGTRQEEMIDHRLTDREWAEEWKHLDHLLNCIMDMVEKTRRSLTVLRRCQEADREELNYWIRRYSDAEDLKKGGGSSSSHSRQQSPVNPDPVALDAHREFLHRPASGYVPEEIWKKAEEAVNEVKRQAMTELQKAVSEAERKAHDMITTERAKMERTVAEAKRQAAEDALAVINQQEDSSESCWNCGRKASETCSGCNTARYCGSFCQHKDWEKHHHICGQTLQAQQQGDTPAVSSSVTPNSGAGSPMDTPPAATPRSTTPGTPSTIETTPR DIP protein Q06455 original sequence version: 75 DIP protein Q06455 original sequence version: 146 C3D2452F96E65679 runx11 1 5 unspecified role unspecified role neutral component neutral component region SeMet substituted se(s)met L-selenomethionine Delta:S(-1)Se(1) Se(S)Met Selenium replaces sulphur semetm undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined region NHR2 domain sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain ecoli Escherichia coli (strain K12) direct interaction direct interaction Note the pdb entry refers to the Uniprot entry Q06455 (resides 335-403). However, the authors refer to the numbering for AML1_ETO (480-551). The protein crystallised has an extra ala-met-ala at the N-terminus. 1, Table 1 The homotetrameric model is established by rotating the asymmetric unit around the 2-fold axis of the P21212 space group 2.0 A R(%) 21.6; Rfree(%) 27.0 runx11-1 2 4 unspecified role unspecified role neutral component neutral component region NHR2 domain sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain aml1_eto_chimera:p.[Leu493Arg;Leu505Glu;Leu508Glu;Leu509Arg;Leu523Arg;Leu526Glu;Leu537Arg] m7 mutation disrupting mutation disrupting interaction mutation disrupting mutation analysis mutation analysis certain certain sequence position certain certain certain sequence position certain L R certain certain sequence position certain certain certain sequence position certain L E certain certain sequence position certain certain certain sequence position certain L E certain certain sequence position certain certain certain sequence position certain L R certain certain sequence position certain certain certain sequence position certain L R certain certain sequence position certain certain certain sequence position certain L E certain certain sequence position certain certain certain sequence position certain L R Fri Jul 22 10:52:54 BST 2016 This feature has been corrected as a result of our quality control procedures. The original label was 'leu493glu,leu505arg,leu508arg,leu509glu,leu523glu,leu526arg,leu537arg' Wed May 02 20:23:47 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'leu493arg,leu505glu,leu508glu,leu509arg,leu523arg,leu526glu,leu537arg' Tue Jun 30 20:10:29 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.[Leu493Arg;Leu505Glu;Leu508Glu;Leu509Arg;Leu523Arg;Leu526Glu;Leu537Arg]' ecoli Escherichia coli (strain K12) direct interaction direct interaction The Uniprot entry Q06455 (resides 335-403)contains the NHR2 domain as does the chimeric protein AML1_ETO. 3 C, Table 2 the continuous sedimentation coefficient distribution for NHR2 contains a major peak near s = 2.7 S, corresponding to 91% of the material. The molecular mass of this predominant species is 36.1 kDa (Table 2) which corresponds closely to the predicted tetramer mass of 35.8 kDa. Thus, the NHR2 domain is clearly a tetramer in solution. Hydrodynamic modeling using the crystal structure predicts a sedimentation coefficient of 2.87 S, which agrees well with the experimental value of 2.68 S. The sedimentation coefficient of m7 was w1.1 S and the mass 7.6 kDa, indicating that m7 is a soluble monomer The m4 (leu345gln-leu375glu-leu378lys-leu389gln) mutant was tetrameric at 20�C by analytical ultracentrifugation (Table 2), but CD spectroscopy revealed that it was only 41% helical, suggesting that the m4 tetramer is significantly destabilized. runx1-human-protein 3 4 unspecified role unspecified role neutral component neutral component aml1_eto_chimera:p.[Leu493Glu;Leu505Arg;Leu508Arg;Leu509Glu;Leu523Glu;Leu526Arg;Leu537Arg] m7 mutation decreasing mutation decreasing interaction hotspot hotspot mutation decreasing mutation analysis mutation analysis certain certain sequence position certain certain certain sequence position certain L E certain certain sequence position certain certain certain sequence position certain L R certain certain sequence position certain certain certain sequence position certain L R certain certain sequence position certain certain certain sequence position certain L E certain certain sequence position certain certain certain sequence position certain L E certain certain sequence position certain certain certain sequence position certain L R certain certain sequence position certain certain certain sequence position certain L R Wed May 02 17:36:05 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'leu493glu,leu505arg,leu508arg,leu509glu,leu523glu,leu526arg,leu537arg' Tue Jun 30 18:43:05 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.[Leu493Glu;Leu505Arg;Leu508Arg;Leu509Glu;Leu523Glu;Leu526Arg;Leu537Arg]' aml1_eto_chimera:p.[Leu493Gln;Leu523Glu;Leu526Lys;Leu537Gln] m4 mutation decreasing mutation decreasing interaction hotspot hotspot mutation decreasing mutation analysis mutation analysis certain certain sequence position certain certain certain sequence position certain L Q certain certain sequence position certain certain certain sequence position certain L E certain certain sequence position certain certain certain sequence position certain L K certain certain sequence position certain certain certain sequence position certain L Q Wed May 02 16:30:53 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'leu493gln,leu523glu,leu526lys,leu537gln' Tue Jun 30 18:43:39 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.[Leu493Gln;Leu523Glu;Leu526Lys;Leu537Gln]' region deletion of the NHR2 domain required to bind necessary binding region required to bind deletion analysis deletion analysis certain certain sequence position certain certain certain sequence position certain in vitro In vitro direct interaction direct interaction 3 A, E full-length AML1/ETO (84 kDa) migrates with an apparent molecular mass greater than 400 kDa. Deletion of the NHR2 domain (Zhang et al., 2001), the m7 mutation, and the m4 mutation shifted the migration of AML1/ETO to that of lower-molecular weight species (Figure 3E). Consistent with the sedimentation velocity and CD measurements, the m4 version of AML1-ETO shows an elution volume intermediate between m7 and AML1-ETO. It is not clear whether the deletion of the NHR2 domain completely abolishes oligomerisation of AML1_ETO. The elution volume of the m4 mutant is in between that for AML1_ETO and the m7 mutant.