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/ Molecular basis of the death-associated protein kinase-calcium/calmodulin regulator complex. Molecular basis of the death-associated protein kinase-calcium/calmodulin regulator complex. Sci Signal 2010 imex curation de Diego I., Kuper J., Bakalova N., Kursula P., Wilmanns M. IBD - Inflammatory bowel disease Ulcerative colitis - Interactions of proteins identified as having a link to ulcerative colitis wilmanns@embl-hamburg.de 05-Jan-2015: Contacted by IntAct-Help. 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 de Diego I., Kuper J., Bakalova N., Kursula P., Wilmanns M. IBD - Inflammatory bowel disease IBD - Inflammatory bowel disease wilmanns@embl-hamburg.de Accepted 2014-NOV-17 by MGT Sci Signal 2010 Only protein-protein interactions imex curation Molecular basis of the death-associated protein kinase-calcium/calmodulin regulator complex. Molecular basis of the death-associated protein kinase-calcium/calmodulin regulator complex. Sci Signal 2010 imex curation de Diego I., Kuper J., Bakalova N., Kursula P., Wilmanns M. IBD - Inflammatory bowel disease Ulcerative colitis - Interactions of proteins identified as having a link to ulcerative colitis wilmanns@embl-hamburg.de 05-Jan-2015: Contacted by IntAct-Help. Only protein-protein interactions in vitro In vitro protein kinase assay protein kinase assay predetermined predetermined participant predetermined de Diego I., Kuper J., Bakalova N., Kursula P., Wilmanns M. IBD - Inflammatory bowel disease The synthetic DAPK consensus substrate peptide is described in PMID:11483604. The protein kinase assay is described in more detail in PMID:6295440. wilmanns@embl-hamburg.de Accepted 2014-NOV-17 by MGT Sci Signal 2010 Only protein-protein interactions imex curation Molecular basis of the death-associated protein kinase-calcium/calmodulin regulator complex. Molecular basis of the death-associated protein kinase-calcium/calmodulin regulator complex. Sci Signal 2010 imex curation de Diego I., Kuper J., Bakalova N., Kursula P., Wilmanns M. IBD - Inflammatory bowel disease Ulcerative colitis - Interactions of proteins identified as having a link to ulcerative colitis wilmanns@embl-hamburg.de 05-Jan-2015: Contacted by IntAct-Help. Only protein-protein interactions in vitro In vitro protein kinase assay protein kinase assay western blot western blot Immuno blot Immuno blot de Diego I., Kuper J., Bakalova N., Kursula P., Wilmanns M. IBD - Inflammatory bowel disease wilmanns@embl-hamburg.de Accepted 2014-NOV-17 by MGT Sci Signal 2010 Only protein-protein interactions imex curation Molecular basis of the death-associated protein kinase-calcium/calmodulin regulator complex. Molecular basis of the death-associated protein kinase-calcium/calmodulin regulator complex. Sci Signal 2010 imex curation de Diego I., Kuper J., Bakalova N., Kursula P., Wilmanns M. IBD - Inflammatory bowel disease Ulcerative colitis - Interactions of proteins identified as having a link to ulcerative colitis wilmanns@embl-hamburg.de 05-Jan-2015: Contacted by IntAct-Help. Only protein-protein interactions ecobd Escherichia coli (strain B / BL21-DE3) molecular sieving molecular sieving Gel Filtration Size Exclusion Chromatography Size Exclusion Chromatography Gel Filtration Sizing column weight by staining molecular weight estimation by staining weight by staining de Diego I., Kuper J., Bakalova N., Kursula P., Wilmanns M. IBD - Inflammatory bowel disease IBD - Inflammatory bowel disease wilmanns@embl-hamburg.de Accepted 2014-NOV-17 by MGT Sci Signal 2010 Only protein-protein interactions imex curation Molecular basis of the death-associated protein kinase-calcium/calmodulin regulator complex. Molecular basis of the death-associated protein kinase-calcium/calmodulin regulator complex. Sci Signal 2010 imex curation de Diego I., Kuper J., Bakalova N., Kursula P., Wilmanns M. IBD - Inflammatory bowel disease Ulcerative colitis - Interactions of proteins identified as having a link to ulcerative colitis wilmanns@embl-hamburg.de 05-Jan-2015: Contacted by IntAct-Help. Only protein-protein interactions in vitro In vitro itc isothermal titration calorimetry ITC ITC itc predetermined predetermined participant predetermined de Diego I., Kuper J., Bakalova N., Kursula P., Wilmanns M. IBD - Inflammatory bowel disease IBD - Inflammatory bowel disease wilmanns@embl-hamburg.de Accepted 2014-NOV-17 by MGT Sci Signal 2010 Only protein-protein interactions imex curation dap_kinase_substrate_peptide Death-associated protein kinase (DAPK) consensus substrate peptide peptide peptide polypeptide oligopeptide oligopeptide polypeptide chemical synthesis Chemical synthesis (Chemical synthesis) KKRPQRRYSNVF calm_human Calmodulin CALM1 CALM CAM CAM1 CALM2 CAMB CALM3 CAM3 CAMC CAMIII CAM2 CALML2 protein protein human Homo sapiens Human MADQLTEEQIAEFKEAFSLFDKDGDGTITTKELGTVMRSLGQNPTEAELQDMINEVDADGNGTIDFPEFLTMMARKMKDTDSEEEIREAFRVFDKDGNGYISAAELRHVMTNLGEKLTDEEVDEMIREADIDGDGQVNYEEFVQMMTAK DIP protein P62158 original sequence version: 97 DIP protein P62158 original sequence version: 144 DIP protein P62158 original sequence version: 146 DIP protein P62158 original sequence version: 73 DIP protein P62158 original sequence version: 102 DIP protein P62158 original sequence version: 98 DIP protein P62158 original sequence version: 96 DIP protein P62158 original sequence version: 111 DIP protein P62158 original sequence version: 131 DIP protein P62158 original sequence version: 136 DIP protein P62158 original sequence version: 145 DIP protein P62158 original sequence version: 154 DIP protein P62158 original sequence version: 148 DIP protein P62158 original sequence version: 116 DIP protein P62158 original sequence version: 120 DIP protein P62158 original sequence version: 130 6B4BC3FCDE10727B dapk1_human Death-associated protein kinase 1 DAPK1 DAPK protein protein human Homo sapiens Human MTVFRQENVDDYYDTGEELGSGQFAVVKKCREKSTGLQYAAKFIKKRRTKSSRRGVSREDIEREVSILKEIQHPNVITLHEVYENKTDVILILELVAGGELFDFLAEKESLTEEEATEFLKQILNGVYYLHSLQIAHFDLKPENIMLLDRNVPKPRIKIIDFGLAHKIDFGNEFKNIFGTPEFVAPEIVNYEPLGLEADMWSIGVITYILLSGASPFLGDTKQETLANVSAVNYEFEDEYFSNTSALAKDFIRRLLVKDPKKRMTIQDSLQHPWIKPKDTQQALSRKASAVNMEKFKKFAARKKWKQSVRLISLCQRLSRSFLSRSNMSVARSDDTLDEEDSFVMKAIIHAINDDNVPGLQHLLGSLSNYDVNQPNKHGTPPLLIAAGCGNIQILQLLIKRGSRIDVQDKGGSNAVYWAARHGHVDTLKFLSENKCPLDVKDKSGEMALHVAARYGHADVAQLLCSFGSNPNIQDKEEETPLHCAAWHGYYSVAKALCEAGCNVNIKNREGETPLLTASARGYHDIVECLAEHGADLNACDKDGHIALHLAVRRCQMEVIKTLLSQGCFVDYQDRHGNTPLHVACKDGNMPIVVALCEANCNLDISNKYGRTPLHLAANNGILDVVRYLCLMGASVEALTTDGKTAEDLARSEQHEHVAGLLARLRKDTHRGLFIQQLRPTQNLQPRIKLKLFGHSGSGKTTLVESLKCGLLRSFFRRRRPRLSSTNSSRFPPSPLASKPTVSVSINNLYPGCENVSVRSRSMMFEPGLTKGMLEVFVAPTHHPHCSADDQSTKAIDIQNAYLNGVGDFSVWEFSGNPVYFCCYDYFAANDPTSIHVVVFSLEEPYEIQLNQVIFWLSFLKSLVPVEEPIAFGGKLKNPLQVVLVATHADIMNVPRPAGGEFGYDKDTSLLKEIRNRFGNDLHISNKLFVLDAGASGSKDMKVLRNHLQEIRSQIVSVCPPMTHLCEKIISTLPSWRKLNGPNQLMSLQQFVYDVQDQLNPLASEEDLRRIAQQLHSTGEINIMQSETVQDVLLLDPRWLCTNVLGKLLSVETPRALHHYRGRYTVEDIQRLVPDSDVEELLQILDAMDICARDLSSGTMVDVPALIKTDNLHRSWADEEDEVMVYGGVRIVPVEHLTPFPCGIFHKVQVNLCRWIHQQSTEGDADIRLWVNGCKLANRGAELLVLLVNHGQGIEVQVRGLETEKIKCCLLLDSVCSTIENVMATTLPGLLTVKHYLSPQQLREHHEPVMIYQPRDFFRAQTLKETSLTNTMGGYKESFSSIMCFGCHDVYSQASLGMDIHASDLNLLTRRKLSRLLDPPDPLGKDWCLLAMNLGLPDLVAKYNTSNGAPKDFLPSPLHALLREWTTYPESTVGTLMSKLRELGRRDAADFLLKASSVFKINLDGNGQEAYASSCNSGTSYNSISSVVSR E2C4246E7C78A6D2 calm1-dapk1 1 7 unspecified role unspecified role neutral component neutral component ef-hand-dom_pair sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain ecobb Escherichia coli (strain B / BL21) 8 unspecified role unspecified role neutral component neutral component ser/thr_dual-sp_kinase sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain ecobb Escherichia coli (strain B / BL21) direct interaction direct interaction Table 1 2.20A working 20.1%, free 26.7% P62158 increases activity of P53355 calm1-dapk1-1 1 8 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 chemical synthesis Chemical synthesis (Chemical synthesis) 7 unspecified role unspecified role neutral component neutral component ef-hand-dom_pair sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain ecobd Escherichia coli (strain B / BL21-DE3) direct interaction direct interaction Table 1 1.70A working 20.3%, free 25.7% calm1-dapk1-2 5 7 unspecified role unspecified role neutral component neutral component ecobd Escherichia coli (strain B / BL21-DE3) 8 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 chemical synthesis Chemical synthesis (Chemical synthesis) direct interaction direct interaction Fig. S3, Table S1 Temp = 25C, pH = 7.4 In this experiment, mutation of the trp305 residue in DAPK has little effect on CaM binding: the trp305tyr mutant binds with Kd = 1.1 +/- 0.2 x 10E-8 molar, while the trp305asp mutant binds with Kd = 9.9 +/- 0.9 x 10E-9 molar. calm1-dapk1-3 4 7 unspecified role unspecified role neutral component neutral component 8 unspecified role unspecified role neutral component neutral component P53355:p.Trp305Asp mutation decreasing mutation decreasing interaction hotspot hotspot mutation decreasing mutation analysis mutation analysis certain certain sequence position certain certain certain sequence position certain W D Wed May 02 17:02:20 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'trp305asp' Tue Jun 30 18:06:39 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Trp305Asp' P53355:p.Trp305Tyr mutation decreasing mutation decreasing interaction hotspot hotspot mutation decreasing mutation analysis mutation analysis certain certain sequence position certain certain certain sequence position certain W Y Wed May 02 15:58:20 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'trp305tyr' Tue Jun 30 16:40:41 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Trp305Tyr' ser/thr_dual-sp_kinase sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain physical association physical association Fig. 5a, 5d dap-dapk1 2 6 enzyme target enzyme target substrate substrate neutral component neutral component chemical synthesis Chemical synthesis (Chemical synthesis) 8 enzyme enzyme neutral component neutral component ser/thr_dual-sp_kinase sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain ecobd Escherichia coli (strain B / BL21-DE3) phosphorylation phosphorylation reaction phosphorylation Fig. 5b, Table 3 Temp = 30C, pH = 7.2 Peptide substrate was phosphorylated by constitutively active DAPK (residues 1-277), while the apo version of DAPK (residues 1-334) showed little catalytic activity (kcat = 2.0 +/- 0.2 x 10E-1 s-1). calm1-dap-4 2 6 enzyme target enzyme target substrate substrate neutral component neutral component chemical synthesis Chemical synthesis (Chemical synthesis) 7 stimulator stimulator neutral component neutral component ecobd Escherichia coli (strain B / BL21-DE3) 8 enzyme enzyme neutral component neutral component P53355:p.Trp305Tyr mutation decreasing rate mutation decreasing interaction rate mutation decreasing rate mutation analysis mutation analysis certain certain sequence position certain certain certain sequence position certain W Y Wed May 02 18:18:09 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'trp305tyr' Tue Jun 30 16:21:43 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Trp305Tyr' P53355:p.Trp305Asp mutation decreasing rate mutation decreasing interaction rate mutation decreasing rate mutation analysis mutation analysis certain certain sequence position certain certain certain sequence position certain W D Wed May 02 15:36:18 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'trp305asp' Tue Jun 30 16:54:40 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Trp305Asp' ser/thr_dual-sp_kinase sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain ecobd Escherichia coli (strain B / BL21-DE3) phosphorylation phosphorylation reaction phosphorylation Fig. 5b, Table 3 Temp = 30C, pH = 7.2 Mutations of the trp305 residue in DAPK decreased the catalytic activity of DAPK towards the peptide substrate: for the trp305tyr mutant kcat = 9.8 +/- 0.3 x 10E-1 s-1; for the trp305asp mutant kcat = 2.6 +/- 0.1 x 10E-1 s-1. calm1-dapk1-4 3 8 putative self putative self putative self putative self ser/thr_dual-sp_kinase sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain P53355:p.Trp305Asp mutation increasing rate mutation increasing interaction rate mutation increasing rate mutation analysis mutation analysis certain certain sequence position certain certain certain sequence position certain W D Wed May 02 14:39:38 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'trp305asp' Tue Jun 30 16:08:59 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Trp305Asp' P53355:p.Ser308Ala mutation disrupting rate mutation disrupting interaction rate mutation disrupting rate mutation analysis mutation analysis certain certain sequence position certain certain certain sequence position certain S A Wed May 02 19:22:40 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'ser308ala' Tue Jun 30 18:25:39 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Ser308Ala' ecobd Escherichia coli (strain B / BL21-DE3) 7 inhibitor inhibitor neutral component neutral component ecobd Escherichia coli (strain B / BL21-DE3) phosphorylation phosphorylation reaction phosphorylation Fig. 5c P53355 ser-308 O-phospho-L-serine dapk1-4 3 8 putative self putative self putative self putative self ser/thr_dual-sp_kinase sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain P53355:p.Ser308Ala mutation disrupting rate mutation disrupting interaction rate mutation disrupting rate mutation analysis mutation analysis certain certain sequence position certain certain certain sequence position certain S A Wed May 02 16:23:10 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'ser308ala' Tue Jun 30 17:43:42 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Ser308Ala' ecobd Escherichia coli (strain B / BL21-DE3) phosphorylation phosphorylation reaction phosphorylation true Fig. 5c P53355 ser-308 O-phospho-L-serine