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/ Topology and regulation of the human eIF4A/4G/4H helicase complex in translation initiation. Topology and regulation of the human eIF4A/4G/4H helicase complex in translation initiation. Cell (0092-8674) 2009 imex curation Marintchev A., Edmonds KA., Marintcheva B., Hendrickson E., Oberer M., Suzuki C., Herdy B., Sonenberg N., Wagner G. RNA binding interactions have not been annotated as I could not trace sequence of RNA, even on manufacturers website. amarint@bu.edu 09-April-2009: Contacted by JYOTI. davet Only protein-protein interactions 6 in vitro In vitro spr surface plasmon resonance Optical biosensor BIAcore(r) BIAcore(r) Optical biosensor spr predetermined predetermined participant predetermined Marintchev A., Edmonds KA., Marintcheva B., Hendrickson E., Oberer M., Suzuki C., Herdy B., Sonenberg N., Wagner G. RNA binding interactions have not been annotated as I could not trace sequence of RNA, even on manufacturers website. amarint@bu.edu Accepted 2009-APR-25 by DAVET. 09-April-2009: Contacted by JYOTI. Cell (0092-8674) 2009 davet Only protein-protein interactions 6 imex curation Topology and regulation of the human eIF4A/4G/4H helicase complex in translation initiation. Topology and regulation of the human eIF4A/4G/4H helicase complex in translation initiation. Cell (0092-8674) 2009 imex curation Marintchev A., Edmonds KA., Marintcheva B., Hendrickson E., Oberer M., Suzuki C., Herdy B., Sonenberg N., Wagner G. RNA binding interactions have not been annotated as I could not trace sequence of RNA, even on manufacturers website. amarint@bu.edu 09-April-2009: Contacted by JYOTI. davet Only protein-protein interactions 6 in vitro In vitro nmr nuclear magnetic resonance NMR NMR nmr predetermined predetermined participant predetermined Marintchev A., Edmonds KA., Marintcheva B., Hendrickson E., Oberer M., Suzuki C., Herdy B., Sonenberg N., Wagner G. RNA binding interactions have not been annotated as I could not trace sequence of RNA, even on manufacturers website. amarint@bu.edu Accepted 2009-APR-25 by DAVET. 09-April-2009: Contacted by JYOTI. Cell (0092-8674) 2009 davet Only protein-protein interactions 6 imex curation Topology and regulation of the human eIF4A/4G/4H helicase complex in translation initiation. Topology and regulation of the human eIF4A/4G/4H helicase complex in translation initiation. Cell (0092-8674) 2009 imex curation Marintchev A., Edmonds KA., Marintcheva B., Hendrickson E., Oberer M., Suzuki C., Herdy B., Sonenberg N., Wagner G. RNA binding interactions have not been annotated as I could not trace sequence of RNA, even on manufacturers website. amarint@bu.edu 09-April-2009: Contacted by JYOTI. davet Only protein-protein interactions 6 in vitro In vitro molecular sieving molecular sieving Gel Filtration Size Exclusion Chromatography Size Exclusion Chromatography Gel Filtration Sizing column weight by comassie molecular weight estimation by coomasie staining weight by comassie Marintchev A., Edmonds KA., Marintcheva B., Hendrickson E., Oberer M., Suzuki C., Herdy B., Sonenberg N., Wagner G. RNA binding interactions have not been annotated as I could not trace sequence of RNA, even on manufacturers website. amarint@bu.edu Accepted 2009-APR-25 by DAVET. 09-April-2009: Contacted by JYOTI. Cell (0092-8674) 2009 davet Only protein-protein interactions 6 imex curation if4a1_human Eukaryotic initiation factor 4A-I EIF4A1 EIF4A DDX2A ATP-dependent RNA helicase eIF4A-1 protein protein human Homo sapiens Human MSASQDSRSRDNGPDGMEPEGVIESNWNEIVDSFDDMNLSESLLRGIYAYGFEKPSAIQQRAILPCIKGYDVIAQAQSGTGKTATFAISILQQIELDLKATQALVLAPTRELAQQIQKVVMALGDYMGASCHACIGGTNVRAEVQKLQMEAPHIIVGTPGRVFDMLNRRYLSPKYIKMFVLDEADEMLSRGFKDQIYDIFQKLNSNTQVVLLSATMPSDVLEVTKKFMRDPIRILVKKEELTLEGIRQFYINVEREEWKLDTLCDLYETLTITQAVIFINTRRKVDWLTEKMHARDFTVSAMHGDMDQKERDVIMREFRSGSSRVLITTDLLARGIDVQQVSLVINYDLPTNRENYIHRIGRGGRFGRKGVAINMVTEEDKRTLRDIETFYNTSIEEMPLNVADLI DIP protein P60842 original sequence version: 125 DIP protein P60842 original sequence version: 59 DIP protein P60842 original sequence version: 63 DIP protein P60842 original sequence version: 85 DIP protein P60842 original sequence version: 86 DIP protein P60842 original sequence version: 92 DIP protein P60842-1 original sequence version: 114 DIP protein P60842 original sequence version: 106 6EF89939F3045420 if4h_human Eukaryotic translation initiation factor 4H Williams-Beuren syndrome chromosomal region 1 protein EIF4H WBSCR1 KIAA0038 WSCR1 protein protein human Homo sapiens Human MADFDTYDDRAYSSFGGGRGSRGSAGGHGSRSQKELPTEPPYTAYVGNLPFNTVQGDIDAIFKDLSIRSVRLVRDKDTDKFKGFCYVEFDEVDSLKEALTYDGALLGDRSLRVDIAEGRKQDKGGFGFRKGGPDDRGMGSSRESRGGWDSRDDFNSGFRDDFLGGRGGSRPGDRRTGPPMGSRFRDGPPLRGSNMDFREPTEEERAQRPRLQLKPRTVATPLNQVANPNSAIFGGARPREEVVQKEQE D3098B7270A9CF38 if4g1_human Eukaryotic translation initiation factor 4 gamma 1 p220 EIF4G1 EIF4G EIF4GI EIF4F protein protein human Homo sapiens Human MNKAPQSTGPPPAPSPGLPQPAFPPGQTAPVVFSTPQATQMNTPSQPRQHFYPSRAQPPSSAASRVQSAAPARPGPAAHVYPAGSQVMMIPSQISYPASQGAYYIPGQGRSTYVVPTQQYPVQPGAPGFYPGASPTEFGTYAGAYYPAQGVQQFPTGVAPTPVLMNQPPQIAPKRERKTIRIRDPNQGGKDITEEIMSGARTASTPTPPQTGGGLEPQANGETPQVAVIVRPDDRSQGAIIADRPGLPGPEHSPSESQPSSPSPTPSPSPVLEPGSEPNLAVLSIPGDTMTTIQMSVEESTPISRETGEPYRLSPEPTPLAEPILEVEVTLSKPVPESEFSSSPLQAPTPLASHTVEIHEPNGMVPSEDLEPEVESSPELAPPPACPSESPVPIAPTAQPEELLNGAPSPPAVDLSPVSEPEEQAKEVTASMAPPTIPSATPATAPSATSPAQEEEMEEEEEEEEGEAGEAGEAESEKGGEELLPPESTPIPANLSQNLEAAAATQVAVSVPKRRRKIKELNKKEAVGDLLDAFKEANPAVPEVENQPPAGSNPGPESEGSGVPPRPEEADETWDSKEDKIHNAENIQPGEQKYEYKSDQWKPLNLEEKKRYDREFLLGFQFIFASMQKPEGLPHISDVVLDKANKTPLRPLDPTRLQGINCGPDFTPSFANLGRTTLSTRGPPRGGPGGELPRGPAGLGPRRSQQGPRKEPRKIIATVLMTEDIKLNKAEKAWKPSSKRTAADKDRGEEDADGSKTQDLFRRVRSILNKLTPQMFQQLMKQVTQLAIDTEERLKGVIDLIFEKAISEPNFSVAYANMCRCLMALKVPTTEKPTVTVNFRKLLLNRCQKEFEKDKDDDEVFEKKQKEMDEAATAEERGRLKEELEEARDIARRRSLGNIKFIGELFKLKMLTEAIMHDCVVKLLKNHDEESLECLCRLLTTIGKDLDFEKAKPRMDQYFNQMEKIIKEKKTSSRIRFMLQDVLDLRGSNWVPRRGDQGPKTIDQIHKEAEMEEHREHIKVQQLMAKGSDKRRGGPPGPPISRGLPLVDDGGWNTVPISKGSRPIDTSRLTKITKPGSIDSNNQLFAPGGRLSWGKGSSGGSGAKPSDAASEAARPATSTLNRFSALQQAVPTESTDNRRVVQRSSLSRERGEKAGDRGDRLERSERGGDRGDRLDRARTPATKRSFSKEVEERSRERPSQPEGLRKAASLTEDRDRGRDAVKREAALPPVSPLKAALSEEELEKKSKAIIEEYLHLNDMKEAVQCVQELASPSLLFIFVRHGVESTLERSAIAREHMGQLLHQLLCAGHLSTAQYYQGLYEILELAEDMEIDIPHVWLYLAELVTPILQEGGVPMGELFREITKPLRPLGKAASLLLEILGLLCKSMGPKKVGTLWREAGLSWKEFLPEGQDIGAFVAEQKVEYTLGEESEAPGQRALPSEELNRQLEKLLKEGSSNQRVFDWIEANLSEQQIVSNTLVRALMTAVCYSAIIFETPLRVDVAVLKARAKLLQKYLCDEQKELQALYALQALVVTLEQPPNLLRMFFDALYDEDVVKEDAFYSWESSKDPAEQQGKGVALKSVTAFFKWLREAEEESDHN DIP protein Q04637 original sequence version: 68 DIP protein Q04637 original sequence version: 166 DIP protein Q04637 original sequence version: 160 DIP protein Q04637-1 original sequence version: 94 DIP protein Q04637 original sequence version: 115 DIP protein Q04637 original sequence version: 119 DIP protein Q04637 original sequence version: 146 DIP protein Q04637-1 original sequence version: 148 DIP protein Q04637 original sequence version: 150 324088B60863DA34 eif4g1-eif4a1-4 1 6 unspecified role unspecified role bait bait heat1 region sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain Q04637:p.Phe977Ala mutation decreasing mutation decreasing interaction hotspot hotspot mutation decreasing mutation analysis mutation analysis certain certain sequence position certain certain certain sequence position certain F A Wed May 02 19:19:20 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'phe977ala' Tue Jun 30 16:32:36 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Phe977Ala' ecoli Escherichia coli (strain K12) 4 unspecified role unspecified role prey prey n-terminal region sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain ecoli Escherichia coli (strain K12) 9 12 direct interaction direct interaction KD for binding of 747-993 EIF4G1 to full length EIF4A1 = 1.2 E-5M. KD for binding of EIF4G1 to 237-406 EIF4A1 = 2.5E-4M. Fig 1 Fig. 5A To test whether the C-terminal portion of eIF4G HEAT-1 contacts eIF4A-NTD, we studied eIF4A binding to WT eIF4G HEAT-1 and the F978A mutant using Surface Plasmon Resonance ATP, ADP eIF4A (1-236) is not required for binding to eIF4G1 HEAT-1, but it stablizes the binding. eif4g1-eif4a1-5 2 4 unspecified role unspecified role neutral component neutral component n-terminal region sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain c-terminal region sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain ecoli Escherichia coli (strain K12) 6 unspecified role unspecified role neutral component neutral component heat2 region sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain Q04637:p.Phe977Ala mutation decreasing mutation decreasing interaction hotspot hotspot mutation decreasing mutation analysis mutation analysis certain certain sequence position certain certain certain sequence position certain F A Wed May 02 16:23:33 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'phe977ala' Tue Jun 30 20:09:21 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Phe977Ala' region 15n label 15n label 15N N15 15N N15 undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined ecoli Escherichia coli (strain K12) direct interaction direct interaction Figs. 3A and 3B, Supp. Fig.1A Using 15N-labeled HEAT-2 and unlabeled individual eIF4A domains, we found that HEAT-2 binds to both eIF4A-NTD (Figure 3A) and eIF4A-CTD (Figure 3B). An asp1259asn mutation affected binding of eIF4G HEAT-2 to full-length eIF4A but had no effect on binding to eIF4A N-terminal domain eif4h-eif4a1-1 2 4 unspecified role unspecified role neutral component neutral component c-terminal region sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain ecoli Escherichia coli (strain K12) 5 unspecified role unspecified role neutral component neutral component region 15n label 15n label 15N N15 15N N15 undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined c-terminal region sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain ecoli Escherichia coli (strain K12) 26 23 direct interaction direct interaction Supp. Figs. 2, 3B Using NMR chemical shift perturbation, we observed binding of eIF4H to eIF4A-CTD (Figures 4B and S2B), but not to eIF4A-NTD (data not shown). The C-terminal 72-residue region of eIF4H was sufficient for the interaction with eIF4A C-terminal domain, although eIF4A C-terminal domain appeared to contact the N-terminal portion of eIF4H as well eif4g1-eif4a1-6 1 4 unspecified role unspecified role prey prey n-terminal region required to bind necessary binding region required to bind certain certain sequence position certain certain certain sequence position certain ecoli Escherichia coli (strain K12) 6 unspecified role unspecified role bait bait heat2 region sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain Q04637:p.Phe977Ala mutation decreasing mutation decreasing interaction hotspot hotspot mutation decreasing mutation analysis mutation analysis certain certain sequence position certain certain certain sequence position certain F A Wed May 02 18:55:23 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'phe977ala' Tue Jun 30 18:20:57 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Phe977Ala' ecoli Escherichia coli (strain K12) direct interaction direct interaction Fig. 5B, Supp. Figs. 1B, 1C SPR graphs of eIF4A binding to immobilized eIF4G HEAT-1 (A) or eIF4G HEAT-2 (B) ATP, ADP An asp1259asn mutation affected binding of eIF4G HEAT-2 to full-length eIF4A but had no effect on binding to eIF4A N-terminal domain. eif4g1-eif4a1-7 3 4 unspecified role unspecified role neutral component neutral component ecoli Escherichia coli (strain K12) 6 unspecified role unspecified role neutral component neutral component se region 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 Supp. Fig 4C When the three proteins are analyzed together, but ATP is omitted from the running buffer, we observe a complex of eIF4A and eIF4G-Se, eluting together beginning in fraction 9 Complex forms in the absence of ATP eif4h-eif4a1-2 3 4 unspecified role unspecified role neutral component neutral component ecoli Escherichia coli (strain K12) 5 unspecified role unspecified role neutral component neutral component ecoli Escherichia coli (strain K12) physical association physical association Supp. Fig 4A, 4B We first confirmed our observation of a binary complex between eIF4A and eIF4H in the presence of ATP (Figure 5C and pink line in Figure S4A), Complex forms in the presence of ATP