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/ Early mitotic degradation of Nek2A depends on Cdc20-independent interaction with the APC/C. Early mitotic degradation of Nek2A depends on Cdc20-independent interaction with the APC/C. Nat. Cell Biol. (1465-7392) 2006 imex curation Hayes MJ., Kimata Y., Wattam SL., Lindon C., Mao G., Yamano H., Fry AM. BioCreative - Critical Assessment of Information Extraction systems in Biology amf5@le.ac.uk 24-JAN-2007: Contacted by JYOTI. Only protein-protein interactions in vitro In vitro anti bait coip anti bait coimmunoprecipitation anti bait coip predetermined predetermined participant predetermined Hayes MJ., Kimata Y., Wattam SL., Lindon C., Mao G., Yamano H., Fry AM. BioCreative - Critical Assessment of Information Extraction systems in Biology Nek2A and Nek2A constructs were 35S-radiolabelled and detected by autoradiography. Other proteins were detected by western blotting. amf5@le.ac.uk Accepted 2006-SEP-13 by ORCHARD. 24-JAN-2007: Contacted by JYOTI. Nat. Cell Biol. (1465-7392) 2006 Only protein-protein interactions imex curation Early mitotic degradation of Nek2A depends on Cdc20-independent interaction with the APC/C. Early mitotic degradation of Nek2A depends on Cdc20-independent interaction with the APC/C. Nat. Cell Biol. (1465-7392) 2006 imex curation Hayes MJ., Kimata Y., Wattam SL., Lindon C., Mao G., Yamano H., Fry AM. BioCreative - Critical Assessment of Information Extraction systems in Biology amf5@le.ac.uk 24-JAN-2007: Contacted by JYOTI. Only protein-protein interactions in vitro In vitro pull down pull down western blot western blot Immuno blot Immuno blot Hayes MJ., Kimata Y., Wattam SL., Lindon C., Mao G., Yamano H., Fry AM. BioCreative - Critical Assessment of Information Extraction systems in Biology Yes have changed the bait and prey around. Also I have corrected the figure legend. Peptides representing the C-terminal 25 residues of Nek2A with (WT) or without (ΔMR) the methionine�arginine (MR) dipeptide were coupled to streptavidin beads and incubated with untreated (CSF), mock-, Cdc20 (Fizzy)- or APC/Cdepleted CSF extracts. Input extracts and bound proteins were analysed by western blotting using antibodies against Apc3, Cdc20 (Fizzy) or PSTAIRE. amf5@le.ac.uk Accepted 2006-SEP-13 by ORCHARD. 24-JAN-2007: Contacted by JYOTI. Nat. Cell Biol. (1465-7392) 2006 Only protein-protein interactions imex curation Early mitotic degradation of Nek2A depends on Cdc20-independent interaction with the APC/C. Early mitotic degradation of Nek2A depends on Cdc20-independent interaction with the APC/C. Nat. Cell Biol. (1465-7392) 2006 imex curation Hayes MJ., Kimata Y., Wattam SL., Lindon C., Mao G., Yamano H., Fry AM. BioCreative - Critical Assessment of Information Extraction systems in Biology amf5@le.ac.uk 24-JAN-2007: Contacted by JYOTI. Only protein-protein interactions human-hela Homo sapiens epitheloid cervix carcinoma cells Human hela Human epitheloid cervix carcinoma HeLa cells derivation: cervix; epithelial; adenocarcinoma ATCC no:CCL-2 anti bait coip anti bait coimmunoprecipitation anti bait coip western blot western blot Immuno blot Immuno blot Hayes MJ., Kimata Y., Wattam SL., Lindon C., Mao G., Yamano H., Fry AM. BioCreative - Critical Assessment of Information Extraction systems in Biology amf5@le.ac.uk Accepted 2006-SEP-13 by ORCHARD. Nat. Cell Biol. (1465-7392) 2006 Only protein-protein interactions imex curation cdc23_human Cell division cycle protein 23 homolog CDC23 ANAPC8 Anaphase-promoting complex subunit 8 Cyclosome subunit 8 protein protein human Homo sapiens Human MAASTSMVPVAVTAAVAPVLSINSDFSDLREIKKQLLLIAGLTRERGLLHSSKWSAELAFSLPALPLAELQPPPPITEEDAQDMDAYTLAKAYFDVKEYDRAAHFLHGCNSKKAYFLYMYSRYLSGEKKKDDETVDSLGPLEKGQVKNEALRELRVELSKKHQARELDGFGLYLYGVVLRKLDLVKEAIDVFVEATHVLPLHWGAWLELCNLITDKEMLKFLSLPDTWMKEFFLAHIYTELQLIEEALQKYQNLIDVGFSKSSYIVSQIAVAYHNIRDIDKALSIFNELRKQDPYRIENMDTFSNLLYVRSMKSELSYLAHNLCEIDKYRVETCCVIGNYYSLRSQHEKAALYFQRALKLNPRYLGAWTLMGHEYMEMKNTSAAIQAYRHAIEVNKRDYRAWYGLGQTYEILKMPFYCLYYYRRAHQLRPNDSRMLVALGECYEKLNQLVEAKKCYWRAYAVGDVEKMALVKLAKLHEQLTESEQAAQCYIKYIQDIYSCGEIVEHLEESTAFRYLAQYYFKCKLWDEASTCAQKCCAFNDTREEGKALLRQILQLRNQGETPTTEVPAPFFLPASLSANNTPTRRVSPLNLSSVTP DIP protein Q9UJX2 original sequence version: 103 DIP protein Q9UJX2 original sequence version: 131 DIP protein Q9UJX2 original sequence version: 135 DIP protein Q9UJX2 original sequence version: 145 DIP protein Q9UJX2 original sequence version: 154 358F2B8745DB9D32 nek2_human Serine/threonine-protein kinase Nek2 NEK2A NimA-like protein kinase 1 HSPK 21 NEK2 NLK1 Never in mitosis A-related kinase 2 protein protein human Homo sapiens Human MPSRAEDYEVLYTIGTGSYGRCQKIRRKSDGKILVWKELDYGSMTEAEKQMLVSEVNLLRELKHPNIVRYYDRIIDRTNTTLYIVMEYCEGGDLASVITKGTKERQYLDEEFVLRVMTQLTLALKECHRRSDGGHTVLHRDLKPANVFLDGKQNVKLGDFGLARILNHDTSFAKTFVGTPYYMSPEQMNRMSYNEKSDIWSLGCLLYELCALMPPFTAFSQKELAGKIREGKFRRIPYRYSDELNEIITRMLNLKDYHRPSVEEILENPLIADLVADEQRRNLERRGRQLGEPEKSQDSSPVLSELKLKEIQLQERERALKAREERLEQKEQELCVRERLAEDKLARAENLLKNYSLLKERKFLSLASNPELLNLPSSVIKKKVHFSGESKENIMRSENSESQLTSKSKCKDLKKRLHAAQLRAQALSDIEKNYQLKSRQILGMR D33A37778ABB6D9E q6gq04_xenla cdc27.S MGC80529 cdc27 protein protein xenla Xenopus laevis (African clawed frog) African clawed frog MTVLQEPVQAAIWQALNHYAYRDAVFLAERLYAEVHSEEALFLLATCYYRSGKAYKAYRLLKAHSCTTPQCKYLLAKCCVDLGKLAEGEQILSGGVLNKQKSQEEIVTEFGDSACFTLSLLGHVYCKTDRLAKGAECYQKSLSLNPFLWSPFESLCEIGEKPDAEQTFKLTSLQSFSNCVPNTCTMTVNTHSVVQHRQPDTVLMETPQDTIELNRLNLESSNIKYPLNSDSSISYIDPGVISPDSLPLGTGNAILSKQVQNKPKTGRSLLGGPAALSPLTPSFGILPLETPSPGDGSYLQSYNSSCVLDVPSSGAPTKKSVARISQAGTKSVFSQSGNSRDVTPVLLVQTQGSGPQTSTTPQVLSPTIAAPPNALPRRSSRLFTSDSSTTKENSKKLKMKFPPKMPNRKTKSKTSKGGGTQTNLNDSLEISKLDSSIISEGKISSVAPQIQALTIQKAAAEGLMSLLRDMGKGYVALCSYNCKEAINILSHLPSHHYNTGWVLCQIGRAYFELAEYLQAERVFSEVRRIESYRVEGMEIYSTTLWHLQKDVALSVLSKDLTDMDKNSPEAWCAAGNCFSLQREHDIAIKFFQRAIQVDPGYAYAFTLLGHEFVLTEELDKALACFRNSIRVNSRHYNAWYGLGMIYYKQEKFSLAEMHFQKALDINPQSSVLLCHIGVVQHALKKSDKALDTLNKAISIDPKNPLCKFHRASILFANEKYKLALQELEELKQIVPKESLVYFLIGKVYKKLGQTHLALMNFSWAMDLDPKGANNQIKEAIDKRYLPDDEEAVTQEEQISECYPYESVATDESQESSMTDADDTQLHAAESDEF DIP protein Q6GQ04 original sequence version: 70 F3E4E06D8CB53604 cdc27_human Cell division cycle protein 27 homolog CDC27 D0S1430E D17S978E CDC27 homolog H-NUC ANAPC3 Anaphase-promoting complex subunit 3 protein protein human Homo sapiens Human MTVLQEPVQAAIWQALNHYAYRDAVFLAERLYAEVHSEEALFLLATCYYRSGKAYKAYRLLKGHSCTTPQCKYLLAKCCVDLSKLAEGEQILSGGVFNKQKSHDDIVTEFGDSACFTLSLLGHVYCKTDRLAKGSECYQKSLSLNPFLWSPFESLCEIGEKPDPDQTFKFTSLQNFSNCLPNSCTTQVPNHSLSHRQPETVLTETPQDTIELNRLNLESSNSKYSLNTDSSVSYIDSAVISPDTVPLGTGTSILSKQVQNKPKTGRSLLGGPAALSPLTPSFGILPLETPSPGDGSYLQNYTNTPPVIDVPSTGAPSKKSVARIGQTGTKSVFSQSGNSREVTPILAQTQSSGPQTSTTPQVLSPTITSPPNALPRRSSRLFTSDSSTTKENSKKLKMKFPPKIPNRKTKSKTNKGGITQPNINDSLEITKLDSSIISEGKISTITPQIQAFNLQKAAAEGLMSLLREMGKGYLALCSYNCKEAINILSHLPSHHYNTGWVLCQIGRAYFELSEYMQAERIFSEVRRIENYRVEGMEIYSTTLWHLQKDVALSVLSKDLTDMDKNSPEAWCAAGNCFSLQREHDIAIKFFQRAIQVDPNYAYAYTLLGHEFVLTEELDKALACFRNAIRVNPRHYNAWYGLGMIYYKQEKFSLAEMHFQKALDINPQSSVLLCHIGVVQHALKKSEKALDTLNKAIVIDPKNPLCKFHRASVLFANEKYKSALQELEELKQIVPKESLVYFLIGKVYKKLGQTHLALMNFSWAMDLDPKGANNQIKEAIDKRYLPDDEEPITQEEQIMGTDESQESSMTDADDTQLHAAESDEF DIP protein P30260 original sequence version: 141 DIP protein P30260 original sequence version: 111 DIP protein P30260 original sequence version: 117 DIP protein P30260 original sequence version: 135 DIP protein P30260 original sequence version: 148 DIP protein P30260 original sequence version: 153 DIP protein P30260 original sequence version: 157 DIP protein P30260 original sequence version: 162 DIP protein P30260 original sequence version: 171 DIP protein P30260 original sequence version: 118 E6C8F59C1EF1DCBA cdc16_human Cell division cycle protein 16 homolog ANAPC6 CDC16 homolog Anaphase-promoting complex subunit 6 Cyclosome subunit 6 CDC16 protein protein human Homo sapiens Human MNLERLRKRVRQYLDQQQYQSALFWADKVASLSREEPQDIYWLAQCLYLTAQYHRAAHALRSRKLDKLYEACRYLAARCHYAAKEHQQALDVLDMEEPINKRLFEKYLKDESGFKDPSSDWEMSQSSIKSSICLLRGKIYDALDNRTLATYSYKEALKLDVYCFEAFDLLTSHHMLTAQEEKELLESLPLSKLCNEEQELLRFLFENKLKKYNKPSETVIPESVDGLQENLDVVVSLAERHYYNCDFKMCYKLTSVVMEKDPFHASCLPVHIGTLVELNKANELFYLSHKLVDLYPSNPVSWFAVGCYYLMVGHKNEHARRYLSKATTLEKTYGPAWIAYGHSFAVESEHDQAMAAYFTAAQLMKGCHLPMLYIGLEYGLTNNSKLAERFFSQALSIAPEDPFVMHEVGVVAFQNGEWKTAEKWFLDALEKIKAIGNEVTVDKWEPLLNNLGHVCRKLKKYAEALDYHRQALVLIPQNASTYSAIGYIHSLMGNFENAVDYFHTALGLRRDDTFSVTMLGHCIEMYIGDSEAYIGADIKDKLKCYDFDVHTMKTLKNIISPPWDFREFEVEKQTAEETGLTPLETSRKTPDSRPSLEETFEIEMNESDMMLETSMSDHST A9C4F24615DF17EB anc2_human Anaphase-promoting complex subunit 2 ANAPC2 APC2 KIAA1406 Cyclosome subunit 2 protein protein human Homo sapiens Human MAAAVVVAEGDSDSRPGQELLVAWNTVSTGLVPPAALGLVSSRTSGAVPPKEEELRAAVEVLRGHGLHSVLEEWFVEVLQNDLQANISPEFWNAISQCENSADEPQCLLLLLDAFGLLESRLDPYLRSLELLEKWTRLGLLMGTGAQGLREEVHTMLRGVLFFSTPRTFQEMIQRLYGCFLRVYMQSKRKGEGGTDPELEGELDSRYARRRYYRLLQSPLCAGCSSDKQQCWCRQALEQFHQLSQVLHRLSLLERVSAEAVTTTLHQVTRERMEDRCRGEYERSFLREFHKWIERVVGWLGKVFLQDGPARPASPEAGNTLRRWRCHVQRFFYRIYASLRIEELFSIVRDFPDSRPAIEDLKYCLERTDQRQQLLVSLKAALETRLLHPGVNTCDIITLYISAIKALRVLDPSMVILEVACEPIRRYLRTREDTVRQIVAGLTGDSDGTGDLAVELSKTDPASLETGQDSEDDSGEPEDWVPDPVDADPGKSSSKRRSSDIISLLVSIYGSKDLFINEYRSLLADRLLHQFSFSPEREIRNVELLKLRFGEAPMHFCEVMLKDMADSRRINANIREEDEKRPAEEQPPFGVYAVILSSEFWPPFKDEKLEVPEDIRAALEAYCKKYEQLKAMRTLSWKHTLGLVTMDVELADRTLSVAVTPVQAVILLYFQDQASWTLEELSKAVKMPVALLRRRMSVWLQQGVLREEPPGTFSVIEEERPQDRDNMVLIDSDDESDSGMASQADQKEEELLLFWTYIQAMLTNLESLSLDRIYNMLRMFVVTGPALAEIDLQELQGYLQKKVRDQQLVYSAGVYRLPKNCS DIP protein Q9UJX6 original sequence version: 116 DIP protein Q9UJX6 original sequence version: 91 DIP protein Q9UJX6 original sequence version: 123 DIP protein Q9UJX6 original sequence version: 128 DIP protein Q9UJX6 original sequence version: 138 DIP protein Q9UJX6 original sequence version: 148 94A2B1D015805573 cdc27-nek2 3 9 unspecified role unspecified role prey prey 5 unspecified role unspecified role prey prey 4 unspecified role unspecified role prey prey 8 unspecified role unspecified role prey prey 7 unspecified role unspecified role bait bait association association The APC/C is a large multi-subunit complex and thus it is not proven that these proteins interact directly. To determine whether Nek2A interacts directly with the APC/C, a human APC/C preparation that was immunopurified using an anti-Apc3 antibody was first western blotted with antibodies against APC/C subunits and Nek2. Nek2A was clearly detected in the purified APC/C preparation, but not in the control immunoprecipitation sample 5 A cdc27-nek2-1 1 7 unspecified role unspecified role bait bait human-hela Homo sapiens epitheloid cervix carcinoma cells Human hela Human epitheloid cervix carcinoma HeLa cells derivation: cervix; epithelial; adenocarcinoma ATCC no:CCL-2 5 unspecified role unspecified role prey prey n-terminal sufficient to bind sufficient binding region sufficient to bind deletion analysis deletion analysis certain certain sequence position certain certain certain sequence position certain region 35s radiolabel 35s radiolabel 35S s35 radiolabelled S35 s35 radiolabelled 35S S35 undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined P51955:p.[Lys391Ala;Glu392Ala;Asn393Ala;Asn399Ala] mutation decreasing mutation decreasing interaction hotspot hotspot mutation decreasing mutation analysis mutation analysis certain certain sequence position certain certain certain sequence position certain K A certain certain sequence position certain certain certain sequence position certain E A certain certain sequence position certain certain certain sequence position certain N A certain certain sequence position certain certain certain sequence position certain N A Wed May 02 15:27:12 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'lys391ala,glu392ala,asn393ala,asn399ala' Tue Jun 30 19:11:58 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.[Lys391Ala;Glu392Ala;Asn393Ala;Asn399Ala]' in vitro In vitro physical association physical association The APC/C is a large multi-subunit complex and thus it is not proven that these proteins interact directly. To verify that interaction between the APC/C and Nek2A is dependent on its destruction motifs, radiolabelled Nek2A, Nek2AΔK−ΔD and both full-length (Cdc13) and nondegradable (Cdc13Δ67) fission yeast cyclin B were incubated with HeLa nuclear extracts before APC/C immunopurification. Nek2A co-purified with the APC/C, whereas only trace amounts of Nek2AΔK−ΔD, Cdc13 or Cdc13Δ67 were co-purified and Nek2. Nek2A was clearly detected in the purified APC/C preparation, but not in the control immunoprecipitation sample 5 B Nek2A strongly interacts with the APC/C in a manner that is dependent on its C-terminal destruction motifs. cdc27-nek2-2 1 6 unspecified role unspecified role bait bait xenla-oocyte Xenopus laevis oocyte Oocyte 5 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 P51955:p.[Lys391Ala;Glu392Ala;Asn393Ala;Asn399Ala] mutation mutation mutation analysis mutation analysis certain certain sequence position certain certain certain sequence position certain K A certain certain sequence position certain certain certain sequence position certain E A certain certain sequence position certain certain certain sequence position certain N A certain certain sequence position certain certain certain sequence position certain N A Wed May 02 17:57:45 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'lys391ala,glu392ala,asn393ala,asn399ala' Tue Jun 30 17:15:28 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.[Lys391Ala;Glu392Ala;Asn393Ala;Asn399Ala]' undefined mutation mutation disrupting mutation disrupting interaction mutation disrupting certain certain sequence position certain certain certain sequence position certain Wed Jul 06 17:45:48 BST 2016 Sequence change details about this feature cannot be ascertained or do not fit with the current version of the referenced protein, so they have been deleted as a result of our quality control procedures. The original label was 'n-terminal' in vitro In vitro physical association physical association The APC/C is a large multi-subunit complex and thus it is not proven that these proteins interact directly. 5 C Nek2A strongly interacts with the APC/C in a manner that is dependent on its C-terminal destruction motifs. We added 35Slabelled Nek2AΔK and Nek2AΔK−ΔMR to CSF extracts and immunopurified the APC/C to assess their ability to bind the Xenopus APC/C. Nek2A-ΔK, but not Nek2AΔK−ΔMR, bound to the APC/C in mock-treated CSF extractsand, equally well, to the APC/C in CSF extracts from which Cdc20 (Fizzy) had been depleted This interaction is not dependent on the presence of Cdc20. Nek2A does not bind Cdc20. cdc27-nek2-3 2 5 unspecified role unspecified role bait bait region biotin tag biotin tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined c-terminal c-terminal 25 residues with terminal MR binding region binding-associated region binding region deletion analysis deletion analysis certain certain sequence position certain certain certain sequence position certain in vitro In vitro 6 unspecified role unspecified role prey prey xenla-oocyte Xenopus laevis oocyte Oocyte physical association physical association The APC/C is a large multi-subunit complex and thus it is not proven that these proteins interact directly. 5 E Nek2A clearly binds directly to the APC/C independently of Cdc20 in a manner that is entirely dependent on the terminal MR dipeptide. peptides representing the C-terminal 25 amino acids of Nek2A with or without the terminal MR dipeptide were used to bind the APC/C in CSF extracts. Again, the full-length peptide, but not the ΔMR peptide, could bind Xenopus APC/C in untreated and Cdc20 (Fizzy)-depleted extracts equally well