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/ Characterization of the interaction between the human DNA topoisomerase IIbeta-binding protein 1 (TopBP1) and the cell division cycle 45 (Cdc45) protein. Characterization of the interaction between the human DNA topoisomerase IIbeta-binding protein 1 (TopBP1) and the cell division cycle 45 (Cdc45) protein. Biochem. J. (0264-6021) 2008 imex curation Schmidt U., Wollmann Y., Franke C., Grosse F., Saluz HP., Hanel F. Cancer - Interactions investigated in the context of cancer frank.haenel@hki-jena.de 17-April-2008: Contacted by JYOTI. Only protein-protein interactions in vitro In vitro pull down pull down weight autoradiogra molecular weight estimation by autoradiography weight autoradiogra Schmidt U., Wollmann Y., Franke C., Grosse F., Saluz HP., Hanel F. Cancer - Interactions investigated in the context of cancer frank.haenel@hki-jena.de Accepted 2008-FEB-10 by JYOTI. 17-April-2008: Contacted by JYOTI. Biochem. J. (0264-6021) 2008 Only protein-protein interactions imex curation Characterization of the interaction between the human DNA topoisomerase IIbeta-binding protein 1 (TopBP1) and the cell division cycle 45 (Cdc45) protein. Characterization of the interaction between the human DNA topoisomerase IIbeta-binding protein 1 (TopBP1) and the cell division cycle 45 (Cdc45) protein. Biochem. J. (0264-6021) 2008 imex curation Schmidt U., Wollmann Y., Franke C., Grosse F., Saluz HP., Hanel F. Cancer - Interactions investigated in the context of cancer frank.haenel@hki-jena.de 17-April-2008: Contacted by JYOTI. Only protein-protein interactions human-hela_s3 Homo sapiens HeLa-S3 cervix carcinoma cell line hela_s3 HeLa-S3 cervix carcinoma cell line Human cervix carcinoma cell line is a subclone of its parent HELA (DSM ACC 57) derived in 1955 confirmed as human with IEF of AST, MDH anti bait coip anti bait coimmunoprecipitation anti bait coip western blot western blot Immuno blot Immuno blot Schmidt U., Wollmann Y., Franke C., Grosse F., Saluz HP., Hanel F. Cancer - Interactions investigated in the context of cancer frank.haenel@hki-jena.de Accepted 2008-FEB-10 by JYOTI. 17-April-2008: Contacted by JYOTI. The stage of the cell cycle Biochem. J. (0264-6021) 2008 Only protein-protein interactions imex curation cdc45_human Cell division control protein 45 homolog CDC45 CDC45L2 UNQ374/PRO710 PORC-PI-1 CDC45L protein protein human Homo sapiens Human MFVSDFRKEFYEVVQSQRVLLFVASDVDALCACKILQALFQCDHVQYTLVPVSGWQELETAFLEHKEQFHYFILINCGANVDLLDILQPDEDTIFFVCDTHRPVNVVNVYNDTQIKLLIKQDDDLEVPAYEDIFRDEEEDEEHSGNDSDGSEPSEKRTRLEEEIVEQTMRRRQRREWEARRRDILFDYEQYEYHGTSSAMVMFELAWMLSKDLNDMLWWAIVGLTDQWVQDKITQMKYVTDVGVLQRHVSRHNHRNEDEENTLSVDCTRISFEYDLRLVLYQHWSLHDSLCNTSYTAARFKLWSVHGQKRLQEFLADMGLPLKQVKQKFQAMDISLKENLREMIEESANKFGMKDMRVQTFSIHFGFKHKFLASDVVFATMSLMESPEKDGSGTDHFIQALDSLSRSNLDKLYHGLELAKKQLRATQQTIASCLCTNLVISQGPFLYCSLMEGTPDVMLFSRPASLSLLSKHLLKSFVCSTKNRRCKLLPLVMAAPLSMEHGTVTVVGIPPETDSSDRKNFFGRAFEKAAESTSSRMLHNHFDLSVIELKAEDRSKFLDALISLLS DIP protein O75419 original sequence version: 88 DIP protein O75419 original sequence version: 96 DIP protein O75419 original sequence version: 135 AE1BE2C8C8E5F867 topb1_human DNA topoisomerase 2-binding protein 1 KIAA0259 TOPBP1 DNA topoisomerase II-binding protein 1 DNA topoisomerase II-beta-binding protein 1 protein protein human Homo sapiens Human MSRNDKEPFFVKFLKSSDNSKCFFKALESIKEFQSEEYLQIITEEEALKIKENDRSLYICDPFSGVVFDHLKKLGCRIVGPQVVIFCMHHQRCVPRAEHPVYNMVMSDVTISCTSLEKEKREEVHKYVQMMGGRVYRDLNVSVTHLIAGEVGSKKYLVAANLKKPILLPSWIKTLWEKSQEKKITRYTDINMEDFKCPIFLGCIICVTGLCGLDRKEVQQLTVKHGGQYMGQLKMNECTHLIVQEPKGQKYECAKRWNVHCVTTQWFFDSIEKGFCQDESIYKTEPRPEAKTMPNSSTPTSQINTIDSRTLSDVSNISNINASCVSESICNSLNSKLEPTLENLENLDVSAFQAPEDLLDGCRIYLCGFSGRKLDKLRRLINSGGGVRFNQLNEDVTHVIVGDYDDELKQFWNKSAHRPHVVGAKWLLECFSKGYMLSEEPYIHANYQPVEIPVSHKPESKAALLKKKNSSFSKKDFAPSEKHEQADEDLLSQYENGSSTVVEAKTSEARPFNDSTHAEPLNDSTHISLQEENQSSVSHCVPDVSTITEEGLFSQKSFLVLGFSNENESNIANIIKENAGKIMSLLSRTVADYAVVPLLGCEVEATVGEVVTNTWLVTCIDYQTLFDPKSNPLFTPVPVMTGMTPLEDCVISFSQCAGAEKESLTFLANLLGASVQEYFVRKSNAKKGMFASTHLILKERGGSKYEAAKKWNLPAVTIAWLLETARTGKRADESHFLIENSTKEERSLETEITNGINLNSDTAEHPGTRLQTHRKTVVTPLDMNRFQSKAFRAVVSQHARQVAASPAVGQPLQKEPSLHLDTPSKFLSKDKLFKPSFDVKDALAALETPGRPSQQKRKPSTPLSEVIVKNLQLALANSSRNAVALSASPQLKEAQSEKEEAPKPLHKVVVCVSKKLSKKQSELNGIAASLGADYRWSFDETVTHFIYQGRPNDTNREYKSVKERGVHIVSEHWLLDCAQECKHLPESLYPHTYNPKMSLDISAVQDGRLCNSRLLSAVSSTKDDEPDPLILEENDVDNMATNNKESAPSNGSGKNDSKGVLTQTLEMRENFQKQLQEIMSATSIVKPQGQRTSLSRSGCNSASSTPDSTRSARSGRSRVLEALRQSRQTVPDVNTEPSQNEQIIWDDPTAREERARLASNLQWPSCPTQYSELQVDIQNLEDSPFQKPLHDSEIAKQAVCDPGNIRVTEAPKHPISEELETPIKDSHLIPTPQAPSIAFPLANPPVAPHPREKIITIEETHEELKKQYIFQLSSLNPQERIDYCHLIEKLGGLVIEKQCFDPTCTHIVVGHPLRNEKYLASVAAGKWVLHRSYLEACRTAGHFVQEEDYEWGSSSILDVLTGINVQQRRLALAAMRWRKKIQQRQESGIVEGAFSGWKVILHVDQSREAGFKRLLQSGGAKVLPGHSVPLFKEATHLFSDLNKLKPDDSGVNIAEAAAQNVYCLRTEYIADYLMQESPPHVENYCLPEAISFIQNNKELGTGLSQKRKAPTEKNKIKRPRVH 02C25880EDCD6AC7 cdc45l-topbp1 2 3 unspecified role unspecified role bait bait 4 unspecified role unspecified role prey prey physical association physical association Fig. 1 A This interaction varies in strength with cell cycle phase. TopBP1 could be co-precipitated with Cdc45 at the G1/S boundary (0 h) and 1 h after release from the mimosine-block. At later time points (3 and 6 h) when more than half of the cell population was in the S-phase co-precipitation, efficiency dramatically declined. Only weak co-precipitation could be obtained with nocodazole-treated cells at the G2/M boundary. TopBP1 could be co-precipitated with Cdc45 G1/S boundary (0h and 1 h after release from the mimosine-block). cdc45l-topbp1-1 1 4 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 in vitro In vitro 3 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 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 ecobb Escherichia coli (strain B / BL21) direct interaction direct interaction Fig. 2 full-length TopBP1 bound specifically to the GST-Cdc45 matrix topbp1-cdc45l 2 4 unspecified role unspecified role prey prey 3 unspecified role unspecified role bait bait physical association physical association Fig. 1 E Cdc45 could be co-precipitated together with endogenous TopBP1 using anti-TopBP1 antibody bound to Sepharose beads in cell extracts from synchronized HeLa-S3 cells in early S-phase 3 h after release from the double TdR-block This interaction occurred in lysates from synchronized HeLa-S3 cells in early S-phase 3 h after release from the double TdR-block. Early S phase (3 h after release from the double TdR-block). topbp1-cdc45l-1 1 4 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 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 sufficient to bind sufficient binding region sufficient to bind deletion analysis deletion analysis certain certain sequence position certain certain certain sequence position certain ecobb Escherichia coli (strain B / BL21) 3 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 in vitro In vitro direct interaction direct interaction Fig. 3 Cdc45 did not bind to the third, the fourth and fifth and the seventh and eighth BRCT domains of TopBP1, neither to GST alone, whereas the first and second as well as the sixth BRCT domain were sufficient for Cdc45 binding. cdc45l-topbp1-2 2 3 unspecified role unspecified role bait bait 4 unspecified role unspecified role prey prey physical association physical association Fig. 1 A This interaction varies in strength with cell cycle phase. TopBP1 could be co-precipitated with Cdc45 at the G1/S boundary (0 h) and 1 h after release from the mimosine-block. At later time points (3 and 6 h) when more than half of the cell population was in the S-phase co-precipitation, efficiency dramatically declined. Only weak co-precipitation could be obtained with nocodazole-treated cells at the G2/M boundary. TopBP1 could be co-precipitated with Cdc45 S phase (3 h and 6 h after release from the mimosine-block). cdc45l-topbp1-3 2 3 unspecified role unspecified role bait bait 4 unspecified role unspecified role prey prey physical association physical association Fig. 1 C This interaction varies in strength with cell cycle phase. TopBP1 could be co-precipitated with Cdc45 at the G1/S boundary (0 h) and 1 h after release from the mimosine-block. At later time points (3 and 6 h) when more than half of the cell population was in the S-phase co-precipitation, efficiency dramatically declined. Only weak co-precipitation could be obtained with nocodazole-treated cells at the G2/M boundary. TopBP1 could be co-precipitated with Cdc45 G2/M boundary (nocodazole-treated cells).