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/ http://www.ebi.ac.uk/ czabotar-2013-4 Bax crystal structures reveal how BH3 domains activate Bax and nucleate its oligomerization to induce apoptosis. in vitro In vitro spr surface plasmon resonance Optical biosensor BIAcore(r) BIAcore(r) Optical biosensor spr predetermined predetermined participant predetermined Czabotar P.E., Westphal D., Dewson G., Ma S., Hockings C., Fairlie W.D., Lee E.F., Yao S., Robin A.Y., Smith B.J., Huang D.C., Kluck R.M., Adams J.M., Colman P.M. Apoptosis - Interactions involving proteins with a function related to apoptosis Bax was incubated with increasing concentrations of peptides, samples were injected onto a CM7 chip bearing immobilized BimBH3. czabotar@wehi.edu.au, pcolman@wehi.edu.au Cell (0092-8674) 2013 Only protein-protein interactions imex curation czabotar-2013-3 Bax crystal structures reveal how BH3 domains activate Bax and nucleate its oligomerization to induce apoptosis. in vitro In vitro x-ray diffraction x-ray crystallography X-ray X-ray x-ray diffraction predetermined predetermined participant predetermined Czabotar P.E., Westphal D., Dewson G., Ma S., Hockings C., Fairlie W.D., Lee E.F., Yao S., Robin A.Y., Smith B.J., Huang D.C., Kluck R.M., Adams J.M., Colman P.M. Apoptosis - Interactions involving proteins with a function related to apoptosis A BAX-GFP fusion protein also crystallised (4BDU) but has not been entered as this fusion is no biological existence. czabotar@wehi.edu.au, pcolman@wehi.edu.au Cell (0092-8674) 2013 Only protein-protein interactions imex curation czabotar-2013-1 Bax crystal structures reveal how BH3 domains activate Bax and nucleate its oligomerization to induce apoptosis. in vitro In vitro molecular sieving molecular sieving Gel Filtration Size Exclusion Chromatography Size Exclusion Chromatography Gel Filtration Sizing column predetermined predetermined participant predetermined Czabotar P.E., Westphal D., Dewson G., Ma S., Hockings C., Fairlie W.D., Lee E.F., Yao S., Robin A.Y., Smith B.J., Huang D.C., Kluck R.M., Adams J.M., Colman P.M. Apoptosis - Interactions involving proteins with a function related to apoptosis czabotar@wehi.edu.au, pcolman@wehi.edu.au Cell (0092-8674) 2013 Only protein-protein interactions imex curation czabotar-2013-2 Bax crystal structures reveal how BH3 domains activate Bax and nucleate its oligomerization to induce apoptosis. in vitro In vitro bn-page blue native page bn-page weight by comassie molecular weight estimation by coomasie staining weight by comassie Czabotar P.E., Westphal D., Dewson G., Ma S., Hockings C., Fairlie W.D., Lee E.F., Yao S., Robin A.Y., Smith B.J., Huang D.C., Kluck R.M., Adams J.M., Colman P.M. Apoptosis - Interactions involving proteins with a function related to apoptosis czabotar@wehi.edu.au, pcolman@wehi.edu.au Cell (0092-8674) 2013 Only protein-protein interactions imex curation czabotar-2013-5 Bax crystal structures reveal how BH3 domains activate Bax and nucleate its oligomerization to induce apoptosis. mouse-embryonic_fibroblast Mus musculus embryonic fibroblasts embryonic_fibroblast Embryonic fibroblast Fibroblasts taken from an embryonic as opposed to a foetal or adult source. Foetus is defined as the stage at which tissue diferentiation is initiated. Tissue taken from an embryonic as opposed to a foetal or adult source. Embryo is defined as the development stage prior to organ formation. crosslink cross-linking study crosslink western blot western blot Immuno blot Immuno blot Czabotar P.E., Westphal D., Dewson G., Ma S., Hockings C., Fairlie W.D., Lee E.F., Yao S., Robin A.Y., Smith B.J., Huang D.C., Kluck R.M., Adams J.M., Colman P.M. Apoptosis - Interactions involving proteins with a function related to apoptosis Cross-linking agent is Bismaleimidoethane (BMOE) - a short-arm, maleimide crosslinker for sulfhydryl conjugation. czabotar@wehi.edu.au, pcolman@wehi.edu.au Cell (0092-8674) 2013 Only protein-protein interactions imex curation b2l11_human Bcl-2-like protein 11 BCL2L11 BIM Bcl2-interacting mediator of cell death protein protein human Homo sapiens Human MAKQPSDVSSECDREGRQLQPAERPPQLRPGAPTSLQTEPQGNPEGNHGGEGDSCPHGSPQGPLAPPASPGPFATRSPLFIFMRRSSLLSRSSSGYFSFDTDRSPAPMSCDKSTQTPSPPCQAFNHYLSAMASMRQAEPADMRPEIWIAQELRRIGDEFNAYYARRVFLNNYQAAEDHPRMVILRLLRYIVRLVWRMH bax_human Apoptosis regulator BAX BAX BCL2L4 Bcl-2-like protein 4 protein protein human Homo sapiens Human MDGSGEQPRGGGPTSSEQIMKTGALLLQGFIQDRAGRMGGEAPELALDPVPQDASTKKLSECLKRIGDELDSNMELQRMIAAVDTDSPREVFFRVAADMFSDGNFNWGRVVALFYFASKLVLKALCTKVPELIRTIMGWTLDFLRERLLGWIQDQGGWDGLLSYFGTPTWQTVTIFVAGVLTASLTIWKKMG apr_human Phorbol-12-myristate-13-acetate-induced protein 1 NOXA Immediate-early-response protein APR PMAIP1 Protein Noxa protein protein human Homo sapiens Human MPGKKARKNAQPSPARAPAELEVECATQLRRFGDKLNFRQKLLNLISKLFCSGT bid_human BH3-interacting domain death agonist p22 BID BID protein protein human Homo sapiens Human MDCEVNNGSSLRDECITNLLVFGFLQSCSDNSFRRELDALGHELPVLAPQWEGYDELQTDGNRSSHSRLGRIEADSESQEDIIRNIARHLAQVGDSMDRSIPPGLVNGLALQLRNTSRSEEDRNRDLATALEQLLQAYPRDMEKEKTMLVLALLLAKKVASHTPSLLRDVFHTTVNFINQNLRTYVRSLARNGMD bak_human Bcl-2 homologous antagonist/killer Apoptosis regulator BAK Bcl-2-like protein 7 BAK1 BAK BCL2L7 CDN1 protein protein human Homo sapiens Human MASGQGPGPPRQECGEPALPSASEEQVAQDTEEVFRSYVFYRHQQEQEAEGVAAPADPEMVTLPLQPSSTMGQVGRQLAIIGDDINRRYDSEFQTMLQHLQPTAENAYEYFTKIATSLFESGINWGRVVALLGFGYRLALHVYQHGLTGFLGQVTRFVVDFMLHHCIARWIAQRGGWVAALNLGNGPILNVLVVLGVVLLGQFVVRRFFKS bcl2l11-bax-8 1419365 n/a 1419370 unspecified role unspecified role bait bait bh3 region sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain false n/a 1419371 unspecified role unspecified role prey prey region sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain false ecoli Escherichia coli (strain K12) n/a 1419371 unspecified role unspecified role prey prey region sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain false chemical synthesis Chemical synthesis (Chemical synthesis) 1419381 1419379 physical association physical association false false false Supp. Fig. 3A, 3B bcl2l11-bax-9 1419365 n/a 1419370 unspecified role unspecified role bait bait bh3 region sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain false n/a 1419372 unspecified role unspecified role prey prey bh3 region sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain false chemical synthesis Chemical synthesis (Chemical synthesis) n/a 1419371 unspecified role unspecified role prey prey region sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain false ecoli Escherichia coli (strain K12) 1419388 1419386 physical association physical association false false false Supp. Fig. 3A, 3C Mutation Cys25Ile of the BCL2L11 protein stimulated the blocking of binding of Bax to BIM by this protein. bcl2l11-bax-7 1419365 n/a 1419370 unspecified role unspecified role bait bait bh3 region sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain false n/a 1419371 unspecified role unspecified role prey prey region sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain false ecoli Escherichia coli (strain K12) n/a 1419370 unspecified role unspecified role prey prey region sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain false chemical synthesis Chemical synthesis (Chemical synthesis) 1419395 1419393 physical association physical association false false false Fig. 5G Supp. Fig. 3A, 3B, 3D Mutations Leu152Ala of the BIM protein inhibit the blocking of binding of Bax to immobilised BIM by this protein. bcl2l11-bax-10 1419365 n/a 1419370 unspecified role unspecified role bait bait bh3 region sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain false n/a 1419373 unspecified role unspecified role prey prey bh3 region sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain false P55957:p.Ile82_Ile83delinsAlaAla mutation disrupting strength mutation disrupting interaction strength mutation disrupting strength certain certain sequence position certain certain certain sequence position certain false P55957:p.Met97Ala mutation decreasing strength mutation decreasing interaction strength mutation decreasing strength certain certain sequence position certain certain certain sequence position certain false chemical synthesis Chemical synthesis (Chemical synthesis) n/a 1419371 unspecified role unspecified role prey prey region sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain false ecoli Escherichia coli (strain K12) 1419404 1419400 physical association physical association false false false Supp. Fig. 3A, 3B Mutations Ile82_83Ala and also Met97Ala of the Bid protein inhibit the blocking of binding of Bax to BIM by this protein. bcl2l11-bak1 1419365 n/a 1419370 unspecified role unspecified role bait bait bh3 region sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain false n/a 1419371 unspecified role unspecified role prey prey region sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain false ecoli Escherichia coli (strain K12) n/a 1419374 unspecified role unspecified role prey prey region sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain false chemical synthesis Chemical synthesis (Chemical synthesis) 1419411 1419409 physical association physical association false false false Supp. Fig. 3A, 3B bax-bid-4 1419366 n/a 1419371 unspecified role unspecified role neutral component neutral component region sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain false ecoli Escherichia coli (strain K12) n/a 1419373 unspecified role unspecified role neutral component neutral component bh3 region sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain false chemical synthesis Chemical synthesis (Chemical synthesis) 1419416 1419414 direct interaction direct interaction false false false Rworking = 0.18 Rfree = 0.23 2.2A Fig. 1A, 5H, Supp Table 1, Supp Fig. 4D bax-bax-5 1419366 n/a 1419371 unspecified role unspecified role neutral component neutral component region sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain false chemical synthesis Chemical synthesis (Chemical synthesis) n/a 1419371 unspecified role unspecified role neutral component neutral component region sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain false ecoli Escherichia coli (strain K12) 1419421 1419419 direct interaction direct interaction false false false Supp Table 1 Rworking = 0.25 Rfree = 0.29 2.2A bax-bax-6 1419366 n/a 1419371 unspecified role unspecified role neutral component neutral component region sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain false ecoli Escherichia coli (strain K12) n/a 1419371 unspecified role unspecified role neutral component neutral component region sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain false ecoli Escherichia coli (strain K12) 1419426 1419424 direct interaction direct interaction false false false Supp Table 1 Rworking = 0.2 Rfree = 0.27 (4BD7), Rworking = 0.2 Rfree = 0.26 (4BD8) 2.2A (4BD7/4BD8) bax-1 1419367 n/a 1419371 unspecified role unspecified role neutral component neutral component region sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain false Q07812:p.[Val121Cys;Ile136Cys] mutation disrupting strength mutation disrupting interaction strength mutation disrupting strength certain certain sequence position certain certain certain sequence position certain false certain certain sequence position certain certain certain sequence position certain false ecoli Escherichia coli (strain K12) Stoichiometry: 2.0 direct interaction direct interaction false false false Octylmaltoside nonionic detergent BIM BH3 peptide + CHAPS BID BH3 peptide + CHAPS BAX BH3 peptide + CHAPS BAK BH3 peptide + CHAPS Mutation of the BimBH3 phe159ala and met97ala, respectively) reduces their ability to promote dimerization of Bax wild-type and particularly of Bax ile66ala. DTT Fig. 1A, 5H, Supp Table 1, Supp Fig. 4D bax-bax-7 1419367 n/a 1419371 unspecified role unspecified role neutral component neutral component Q07812:p.Arg109Asp mutation decreasing strength mutation decreasing interaction strength mutation decreasing strength certain certain sequence position certain certain certain sequence position certain false region sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain false ecoli Escherichia coli (strain K12) n/a 1419371 unspecified role unspecified role neutral component neutral component region sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain false Q07812:p.Asp68Arg mutation decreasing strength mutation decreasing interaction strength mutation decreasing strength certain certain sequence position certain certain certain sequence position certain false chemical synthesis Chemical synthesis (Chemical synthesis) 1419436 1419434 direct interaction direct interaction false false false Fig. 3E bax-1 1419368 n/a 1419371 unspecified role unspecified role neutral component neutral component region sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain false ecoli Escherichia coli (strain K12) Stoichiometry: 2.0 direct interaction direct interaction false false false Fig. 1B, 1C, 3B Octylmaltoside nonionic detergent BIM BH3 peptide + CHAPS - mutations leu152ala and pro144_glu145ala inhibit this. BID BH3 peptide + CHAPS - mutations ile182-183ala inhibit this. BK BH3 peptide + CHAPS BAX BH3 peptide + CHAPS NOXA and BAD BH3 peptides + CHAPS did not induce dimer formation bax-1 1419369 n/a 1419371 unspecified role unspecified role neutral component neutral component Q07812:p.Glu69Cys mutation increasing strength mutation increasing interaction strength mutation increasing strength certain certain sequence position certain certain certain sequence position certain false Q07812:p.Ser184Leu mutation mutation certain certain sequence position certain certain certain sequence position certain false Stoichiometry: 2.0 physical association physical association false false false Fig. 6C BID BH3 peptide + CHAPS Full-length Bax glu69cys residues exposed to an apoptotic stimulus (tBid) could be crosslinked with the 8A linker BMOE, whereas residues farther apart (arg65cys) could not