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/ Crystal structure of the Caenorhabditis elegans apoptosome reveals an octameric assembly of CED-4. Crystal structure of the Caenorhabditis elegans apoptosome reveals an octameric assembly of CED-4. Cell (0092-8674) 2010 imex curation Qi S., Pang Y., Hu Q., Liu Q., Li H., Zhou Y., He T., Liang Q., Liu Y., Yuan X., Luo G., Li H., Wang J., Yan N., Shi Y. Apoptosis - Interactions involving proteins with a function related to apoptosis nyan@tsinghua.edu.cn, shi-lab@tsinghua.edu.cn 13-Sept-2010: Contacted by JYOTI. jyoti Only protein-protein interactions in vitro In vitro x-ray diffraction x-ray crystallography X-ray X-ray x-ray diffraction predetermined predetermined participant predetermined Qi S., Pang Y., Hu Q., Liu Q., Li H., Zhou Y., He T., Liang Q., Liu Y., Yuan X., Luo G., Li H., Wang J., Yan N., Shi Y. Apoptosis - Interactions involving proteins with a function related to apoptosis nyan@tsinghua.edu.cn, shi-lab@tsinghua.edu.cn Accepted 2010-JUL-30 by JYOTI. Cell (0092-8674) 2010 jyoti Only protein-protein interactions imex curation Crystal structure of the Caenorhabditis elegans apoptosome reveals an octameric assembly of CED-4. Crystal structure of the Caenorhabditis elegans apoptosome reveals an octameric assembly of CED-4. Cell (0092-8674) 2010 imex curation Qi S., Pang Y., Hu Q., Liu Q., Li H., Zhou Y., He T., Liang Q., Liu Y., Yuan X., Luo G., Li H., Wang J., Yan N., Shi Y. Apoptosis - Interactions involving proteins with a function related to apoptosis nyan@tsinghua.edu.cn, shi-lab@tsinghua.edu.cn 13-Sept-2010: Contacted by JYOTI. jyoti Only protein-protein interactions in vitro In vitro molecular sieving molecular sieving Gel Filtration Size Exclusion Chromatography Size Exclusion Chromatography Gel Filtration Sizing column anti tag western anti tag western blot anti tag western Qi S., Pang Y., Hu Q., Liu Q., Li H., Zhou Y., He T., Liang Q., Liu Y., Yuan X., Luo G., Li H., Wang J., Yan N., Shi Y. Apoptosis - Interactions involving proteins with a function related to apoptosis nyan@tsinghua.edu.cn, shi-lab@tsinghua.edu.cn Accepted 2010-JUL-30 by JYOTI. 13-Sept-2010: Contacted by JYOTI. Cell (0092-8674) 2010 jyoti Only protein-protein interactions imex curation Crystal structure of the Caenorhabditis elegans apoptosome reveals an octameric assembly of CED-4. Crystal structure of the Caenorhabditis elegans apoptosome reveals an octameric assembly of CED-4. Cell (0092-8674) 2010 imex curation Qi S., Pang Y., Hu Q., Liu Q., Li H., Zhou Y., He T., Liang Q., Liu Y., Yuan X., Luo G., Li H., Wang J., Yan N., Shi Y. Apoptosis - Interactions involving proteins with a function related to apoptosis nyan@tsinghua.edu.cn, shi-lab@tsinghua.edu.cn 13-Sept-2010: Contacted by JYOTI. jyoti Only protein-protein interactions in vitro In vitro biochemical biochemical predetermined predetermined participant predetermined Qi S., Pang Y., Hu Q., Liu Q., Li H., Zhou Y., He T., Liang Q., Liu Y., Yuan X., Luo G., Li H., Wang J., Yan N., Shi Y. Apoptosis - Interactions involving proteins with a function related to apoptosis Capillary electrophoresis nyan@tsinghua.edu.cn, shi-lab@tsinghua.edu.cn Accepted 2010-JUL-30 by JYOTI. 13-Sept-2010: Contacted by JYOTI. Cell (0092-8674) 2010 jyoti Only protein-protein interactions imex curation Crystal structure of the Caenorhabditis elegans apoptosome reveals an octameric assembly of CED-4. Crystal structure of the Caenorhabditis elegans apoptosome reveals an octameric assembly of CED-4. Cell (0092-8674) 2010 imex curation Qi S., Pang Y., Hu Q., Liu Q., Li H., Zhou Y., He T., Liang Q., Liu Y., Yuan X., Luo G., Li H., Wang J., Yan N., Shi Y. Apoptosis - Interactions involving proteins with a function related to apoptosis nyan@tsinghua.edu.cn, shi-lab@tsinghua.edu.cn 13-Sept-2010: Contacted by JYOTI. jyoti Only protein-protein interactions in vitro In vitro cosedimentation cosedimentation predetermined predetermined participant predetermined Qi S., Pang Y., Hu Q., Liu Q., Li H., Zhou Y., He T., Liang Q., Liu Y., Yuan X., Luo G., Li H., Wang J., Yan N., Shi Y. Apoptosis - Interactions involving proteins with a function related to apoptosis Sedimentation equilibrium analysis nyan@tsinghua.edu.cn, shi-lab@tsinghua.edu.cn Accepted 2010-JUL-30 by JYOTI. 13-Sept-2010: Contacted by JYOTI. Cell (0092-8674) 2010 jyoti Only protein-protein interactions imex curation Crystal structure of the Caenorhabditis elegans apoptosome reveals an octameric assembly of CED-4. Crystal structure of the Caenorhabditis elegans apoptosome reveals an octameric assembly of CED-4. Cell (0092-8674) 2010 imex curation Qi S., Pang Y., Hu Q., Liu Q., Li H., Zhou Y., He T., Liang Q., Liu Y., Yuan X., Luo G., Li H., Wang J., Yan N., Shi Y. Apoptosis - Interactions involving proteins with a function related to apoptosis nyan@tsinghua.edu.cn, shi-lab@tsinghua.edu.cn 13-Sept-2010: Contacted by JYOTI. jyoti Only protein-protein interactions in vitro In vitro molecular sieving molecular sieving Gel Filtration Size Exclusion Chromatography Size Exclusion Chromatography Gel Filtration Sizing column predetermined predetermined participant predetermined Qi S., Pang Y., Hu Q., Liu Q., Li H., Zhou Y., He T., Liang Q., Liu Y., Yuan X., Luo G., Li H., Wang J., Yan N., Shi Y. Apoptosis - Interactions involving proteins with a function related to apoptosis nyan@tsinghua.edu.cn, shi-lab@tsinghua.edu.cn Accepted 2010-JUL-30 by JYOTI. 13-Sept-2010: Contacted by JYOTI. Cell (0092-8674) 2010 jyoti Only protein-protein interactions imex curation Crystal structure of the Caenorhabditis elegans apoptosome reveals an octameric assembly of CED-4. Crystal structure of the Caenorhabditis elegans apoptosome reveals an octameric assembly of CED-4. Cell (0092-8674) 2010 imex curation Qi S., Pang Y., Hu Q., Liu Q., Li H., Zhou Y., He T., Liang Q., Liu Y., Yuan X., Luo G., Li H., Wang J., Yan N., Shi Y. Apoptosis - Interactions involving proteins with a function related to apoptosis nyan@tsinghua.edu.cn, shi-lab@tsinghua.edu.cn 13-Sept-2010: Contacted by JYOTI. jyoti Only protein-protein interactions in vitro In vitro comigration in sds comigration in sds page comigration in sds weight by comassie molecular weight estimation by coomasie staining weight by comassie Qi S., Pang Y., Hu Q., Liu Q., Li H., Zhou Y., He T., Liang Q., Liu Y., Yuan X., Luo G., Li H., Wang J., Yan N., Shi Y. Apoptosis - Interactions involving proteins with a function related to apoptosis Crystals were harvested, washed, and applied to SDS-PAGE followed by coomassie blue staining nyan@tsinghua.edu.cn, shi-lab@tsinghua.edu.cn Accepted 2010-JUL-30 by JYOTI. 13-Sept-2010: Contacted by JYOTI. Cell (0092-8674) 2010 jyoti Only protein-protein interactions imex curation Crystal structure of the Caenorhabditis elegans apoptosome reveals an octameric assembly of CED-4. Crystal structure of the Caenorhabditis elegans apoptosome reveals an octameric assembly of CED-4. Cell (0092-8674) 2010 imex curation Qi S., Pang Y., Hu Q., Liu Q., Li H., Zhou Y., He T., Liang Q., Liu Y., Yuan X., Luo G., Li H., Wang J., Yan N., Shi Y. Apoptosis - Interactions involving proteins with a function related to apoptosis nyan@tsinghua.edu.cn, shi-lab@tsinghua.edu.cn 13-Sept-2010: Contacted by JYOTI. jyoti Only protein-protein interactions in vitro In vitro itc isothermal titration calorimetry ITC ITC itc predetermined predetermined participant predetermined Qi S., Pang Y., Hu Q., Liu Q., Li H., Zhou Y., He T., Liang Q., Liu Y., Yuan X., Luo G., Li H., Wang J., Yan N., Shi Y. Apoptosis - Interactions involving proteins with a function related to apoptosis Sedimentation equilibrium analysis nyan@tsinghua.edu.cn, shi-lab@tsinghua.edu.cn Accepted 2010-JUL-30 by JYOTI. 13-Sept-2010: Contacted by JYOTI. Cell (0092-8674) 2010 jyoti Only protein-protein interactions imex curation q9xyf4-pro_0000004671 Caspase Nc subunit 1 Dronc NEDD2-like caspase Nc CG8091 protein protein drome Drosophila melanogaster (Fruit fly) Fruit fly DTPSPEASEGPCVSKLRNEPLGALTPYVGVVDGPEVKKSKKIHGGDSAILGTYKMQSRFNRGVLLMVNIMDYPDQNRRRIGAEKDSKSLIHLFQELNFTIFPYGNVNQDQFFKLLTMVTSSSYVQNTECFVMVLMTHGNSVEGKEKVEFCDGSVVDMQKIKDHFQTAKCPYLVNKPKVLMFPFCRG 135 320 B48D5E1D0D85930A p30429-2 Cell death protein 4 Short C35D10.9 ced-4 protein protein caeel Caenorhabditis elegans MLCEIECRALSTAHTRLIHDFEPRDALTYLEGKNIFTEDHSELISKMSTRLERIANFLRIYRRQASELGPLIDFFNYNNQSHLADFLEDYIDFAINEPDLLRPVVIAPQFSRQMLDRKLLLGNVPKQMTCYIREYHVDRVIKKLDEMCDLDSFFLFLHGRAGSGKSVIASQALSKSDQLIGINYDSIVWLKDSGTAPKSTFDLFTDILLMLKSEDDLLNFPSVEHVTSVVLKRMICNALIDRPNTLFVFDDVVQEETIRWAQELRLRCLVTTRDVEISNAASQTCEFIEVTSLEIDECYDFLEAYGMPMPVGEKEEDVLNKTIELSSGNPATLMMFFKSCEPKTFEKMAQLNNKLESRGLVGVECITPYSYKSLAMALQRCVEVLSDEDRSALAFAVVMPPGVDIPVKLWSCVIPVDICSNEEEQLDDEVADRLKRLSKRGALLSGKRMPVLTFKIDHIIHMFLKHVVDAQTIANGISILEQRLLEIGNNNVSVPERHIPSHFQKFRRSSASEMYPKTTEETVIRPEDFPKFMQLHQKFYDSLKNFACC Major transcript. DB2A7969BDA50AF8 ac-devd-cho Ac-Asp-Glu-Val-Asp-CHO small molecule small molecule chemical synthesis Chemical synthesis (Chemical synthesis) ced3_caeel Cell death protein 3 ced-3 Caspase ced-3 C48D1.2 protein protein caeel Caenorhabditis elegans MMRQDRRSLLERNIMMFSSHLKVDEILEVLIAKQVLNSDNGDMINSCGTVREKRREIVKAVQRRGDVAFDAFYDALRSTGHEGLAEVLEPLARSVDSNAVEFECPMSPASHRRSRALSPAGYTSPTRVHRDSVSSVSSFTSYQDIYSRARSRSRSRALHSSDRHNYSSPPVNAFPSQPSSANSSFTGCSSLGYSSSRNRSFSKASGPTQYIFHEEDMNFVDAPTISRVFDEKTMYRNFSSPRGMCLIINNEHFEQMPTRNGTKADKDNLTNLFRCMGYTVICKDNLTGRGMLLTIRDFAKHESHGDSAILVILSHGEENVIIGVDDIPISTHEIYDLLNAANAPRLANKPKIVFVQACRGERRDNGFPVLDSVDGVPAFLRRGWDNRDGPLFNFLGCVRPQVQQVWRKKPSQADILIAYATTAQYVSWRNSARGSWFIQAVCEVFSTHAKDMDVVELLTEVNKKVACGFQTSQGSNILKQMPEMTSRLLKKFYFWPEARNSAV DIP protein P42573 original sequence version: 122 DIP protein P42573 original sequence version: 131 DIP protein P42573 original sequence version: 85 722D5831F94DAA69 q9xyf4-pro_0000004672 Dronc NEDD2-like caspase Nc CG8091 protein protein drome Drosophila melanogaster (Fruit fly) Fruit fly DEYD 321 324 6ABB5B1AC0000000 ced4 1 9 unspecified role unspecified role neutral component neutral component ecoli Escherichia coli (strain K12) direct interaction direct interaction 3.55A Rworking 27.5%, Rfree 31.1% We succeeded in the generation of diffraction-quality crystals for the proteolysed CED-4 apoptosome and determined its X-ray structure at 3.55 A resolution (Table S1). Supp. Table 1 The assembly contains eight molecules of CED-4, arranged as a tetramer of an asymmetric dimer. Each CED-4 contains an ATP molecule and a Mg2+ ion, both buried in the assembly and inaccessible to water molecules. nc-1 5 12 unspecified role unspecified role neutral component neutral component 8 unspecified role unspecified role neutral component neutral component direct interaction direct interaction Supp. Fig. 5C The two-chain Dronc, known to exist as a homodimer, served as a positive control for gel filtration. Expression system of Nc not given. The two chains of mature Nc form a homodimer. ced3 5 11 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 ecoli Escherichia coli (strain K12) 11 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 P42573:p.Cys358Ala mutation mutation certain certain sequence position certain certain certain sequence position certain C A Wed May 02 16:40:04 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'cys358ala' Tue Jun 30 16:58:08 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Cys358Ala' ecoli Escherichia coli (strain K12) 22 20 direct interaction direct interaction The two-chain CED-3 (residues 198-374, 389-503, C358A) exhibited the same apparent molecular mass of approximately 35 kD as the unprocessed CED-3 (residues 198-503, C358A). Supp. Fig. 5C ced3-2 4 11 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 P42573:p.Cys358Ala mutation mutation certain certain sequence position certain certain certain sequence position certain C A Wed May 02 19:16:09 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'cys358ala' Tue Jun 30 19:54:32 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Cys358Ala' ecoli Escherichia coli (strain K12) 11 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 ecoli Escherichia coli (strain K12) direct interaction direct interaction Analytical ultracentrifugation analysis revealed that the two-chain CED-3 (residues 198�374, 389�503, C358A) had a molecular mass of 35.25 kDa, consistent with the calculated molecular weight of a monomer. Supp. Fig. 5D ced3-ced4 5 9 unspecified role unspecified role neutral component neutral component ecoli Escherichia coli (strain K12) 11 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 ecoli Escherichia coli (strain K12) 11 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 P42573:p.Cys358Ala mutation mutation certain certain sequence position certain certain certain sequence position certain C A Wed May 02 16:27:50 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'cys358ala' Tue Jun 30 17:46:37 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Cys358Ala' ecoli Escherichia coli (strain K12) direct interaction direct interaction We generated a two-chain CED-3 variant (residues 198�374 and 389�503, C358S) and examined its potential interaction with CED-4 using gel filtration. Supp. Fig. 5D ced3-ced4-2 3 11 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 P42573:p.Cys358Ala mutation mutation certain certain sequence position certain certain certain sequence position certain C A Wed May 02 16:48:10 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'cys358ala' Tue Jun 30 17:23:40 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Cys358Ala' ecoli Escherichia coli (strain K12) 11 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 ecoli Escherichia coli (strain K12) 9 unspecified role unspecified role neutral component neutral component ecoli Escherichia coli (strain K12) physical association physical association The peak fraction of the CED-4:CED-3 complex from gel filtration was subjected to capillary electrophoresis under denaturing conditions. Supp. Fig. 5E ced3-ced4-3 7 11 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 P42573:p.Cys358Ala mutation mutation certain certain sequence position certain certain certain sequence position certain C A Wed May 02 19:55:07 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'cys358ala' Tue Jun 30 18:47:06 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Cys358Ala' ecoli Escherichia coli (strain K12) 11 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 ecoli Escherichia coli (strain K12) 9 unspecified role unspecified role neutral component neutral component ecoli Escherichia coli (strain K12) physical association physical association Binding of uncleaved CED-3 (residues 198�503, C358A) or two-chain CED-3 (residues 198�374, 389�503, C358A) by CED-4 was examined by isothermal titration calorimetry (ITC). Supp. Fig. 5F ced3-ced4-4 6 11 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 ecoli Escherichia coli (strain K12) 9 unspecified role unspecified role neutral component neutral component ecoli Escherichia coli (strain K12) 11 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 P42573:p.Cys358Ala mutation mutation certain certain sequence position certain certain certain sequence position certain C A Wed May 02 18:57:08 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'cys358ala' Tue Jun 30 16:46:17 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Cys358Ala' ecoli Escherichia coli (strain K12) physical association physical association SDS-PAGE of crystals of the CED-4:CED-3 complex. Supp. Fig. 6A Although ced-3 was not visible in a ced-3:ced-4 crystal, subsequent analysis of the crystals showed both proteins to be present. ced3-devd 2 11 unspecified role unspecified role neutral component neutral component c-terminal his tag his tag Hexa-His-tag 6-His-tag Histidine-tag Hexa-His-tag Histidine-tag 6-His-tag c-term range C-terminal range c-term range c-term range C-terminal range c-term range 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) 10 unspecified role unspecified role neutral component neutral component direct interaction direct interaction The processed, active CED-3, derived from residues 198-503 with a C-terminal His6 tag, was incubated with the covalent inhibitor DEVD-CHO and then loaded into gel filtration. Supp. Fig. 6A