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/ The coiled-coil domain of Stat3 is essential for its SH2 domain-mediated receptor binding and subsequent activation induced by epidermal growth factor and interleukin-6. The coiled-coil domain of Stat3 is essential for its SH2 domain-mediated receptor binding and subsequent activation induced by epidermal growth factor and interleukin-6. Mol. Cell. Biol. (0270-7306) 2000 Zhang T., Kee WH., Seow KT., Fung W., Cao X. IBD - Inflammatory bowel disease Crohn's disease - Interactions of proteins identified as having a link to Crohn's disease Cancer - Interactions investigated in the context of cancer COS-1 cells were transfected with the Stat3 plasmids and lysed after 48 h. The biotinylated peptides derived from gp130 were incubated with streptavidin-Sepharose. The beads were washed and then incubated with aliquots of lysates. The complexes were washed, fractionated on SDS-PAGE, and immunoblotted with anti-Stat3 antibody. The blot was then stripped and reprobed with anti-SHP-2 antibody. mcbcaoxm@imcb.nus.edu.sg 27-NOV-2006: Contacted by JYOTI. in vitro In vitro enzymatic study enzymatic study predetermined predetermined participant predetermined Zhang T., Kee WH., Seow KT., Fung W., Cao X. IBD - Inflammatory bowel disease Crohn's disease - Interactions of proteins identified as having a link to Crohn's disease Cancer - Interactions investigated in the context of cancer COS-1 cells were transfected with the Stat3 plasmids and lysed after 48 h. The biotinylated peptides derived from gp130 were incubated with streptavidin-Sepharose. The beads were washed and then incubated with aliquots of lysates. The complexes were washed, fractionated on SDS-PAGE, and immunoblotted with anti-Stat3 antibody. The blot was then stripped and reprobed with anti-SHP-2 antibody. mcbcaoxm@imcb.nus.edu.sg Accepted 2006-NOV-13 by DAVET. 27-NOV-2006: Contacted by JYOTI. Mol. Cell. Biol. (0270-7306) 2000 The coiled-coil domain of Stat3 is essential for its SH2 domain-mediated receptor binding and subsequent activation induced by epidermal growth factor and interleukin-6. The coiled-coil domain of Stat3 is essential for its SH2 domain-mediated receptor binding and subsequent activation induced by epidermal growth factor and interleukin-6. Mol. Cell. Biol. (0270-7306) 2000 Zhang T., Kee WH., Seow KT., Fung W., Cao X. IBD - Inflammatory bowel disease Crohn's disease - Interactions of proteins identified as having a link to Crohn's disease Cancer - Interactions investigated in the context of cancer COS-1 cells were transfected with the Stat3 plasmids and lysed after 48 h. The biotinylated peptides derived from gp130 were incubated with streptavidin-Sepharose. The beads were washed and then incubated with aliquots of lysates. The complexes were washed, fractionated on SDS-PAGE, and immunoblotted with anti-Stat3 antibody. The blot was then stripped and reprobed with anti-SHP-2 antibody. mcbcaoxm@imcb.nus.edu.sg 27-NOV-2006: Contacted by JYOTI. in vitro In vitro pull down pull down antibody detection identification by antibody antibody detection Zhang T., Kee WH., Seow KT., Fung W., Cao X. IBD - Inflammatory bowel disease Crohn's disease - Interactions of proteins identified as having a link to Crohn's disease Cancer - Interactions investigated in the context of cancer COS-1 cells were transfected with the Stat3 plasmids and lysed after 48 h. The biotinylated peptides derived from gp130 were incubated with streptavidin-Sepharose. The beads were washed and then incubated with aliquots of lysates. The complexes were washed, fractionated on SDS-PAGE, and immunoblotted with anti-Stat3 antibody. The blot was then stripped and reprobed with anti-SHP-2 antibody. mcbcaoxm@imcb.nus.edu.sg Accepted 2006-NOV-13 by DAVET. 27-NOV-2006: Contacted by JYOTI. Mol. Cell. Biol. (0270-7306) 2000 The coiled-coil domain of Stat3 is essential for its SH2 domain-mediated receptor binding and subsequent activation induced by epidermal growth factor and interleukin-6. The coiled-coil domain of Stat3 is essential for its SH2 domain-mediated receptor binding and subsequent activation induced by epidermal growth factor and interleukin-6. Mol. Cell. Biol. (0270-7306) 2000 Zhang T., Kee WH., Seow KT., Fung W., Cao X. IBD - Inflammatory bowel disease Crohn's disease - Interactions of proteins identified as having a link to Crohn's disease Cancer - Interactions investigated in the context of cancer COS-1 cells were transfected with the Stat3 plasmids and lysed after 48 h. The biotinylated peptides derived from gp130 were incubated with streptavidin-Sepharose. The beads were washed and then incubated with aliquots of lysates. The complexes were washed, fractionated on SDS-PAGE, and immunoblotted with anti-Stat3 antibody. The blot was then stripped and reprobed with anti-SHP-2 antibody. mcbcaoxm@imcb.nus.edu.sg 27-NOV-2006: Contacted by JYOTI. in vitro In vitro pull down pull down western blot western blot Immuno blot Immuno blot Zhang T., Kee WH., Seow KT., Fung W., Cao X. IBD - Inflammatory bowel disease Crohn's disease - Interactions of proteins identified as having a link to Crohn's disease Cancer - Interactions investigated in the context of cancer COS-1 cells were transfected with the Stat3 plasmids and lysed after 48 h. The biotinylated peptides derived from gp130 were incubated with streptavidin-Sepharose. The beads were washed and then incubated with aliquots of lysates. The complexes were washed, fractionated on SDS-PAGE, and immunoblotted with anti-Stat3 antibody. The blot was then stripped and reprobed with anti-SHP-2 antibody. mcbcaoxm@imcb.nus.edu.sg Accepted 2006-NOV-13 by DAVET. 27-NOV-2006: Contacted by JYOTI. Mol. Cell. Biol. (0270-7306) 2000 il6rb_human Interleukin-6 receptor subunit beta IL6ST Interleukin-6 signal transducer Membrane glycoprotein 130 CDw130 Oncostatin-M receptor subunit alpha protein protein human Homo sapiens Human MLTLQTWLVQALFIFLTTESTGELLDPCGYISPESPVVQLHSNFTAVCVLKEKCMDYFHVNANYIVWKTNHFTIPKEQYTIINRTASSVTFTDIASLNIQLTCNILTFGQLEQNVYGITIISGLPPEKPKNLSCIVNEGKKMRCEWDGGRETHLETNFTLKSEWATHKFADCKAKRDTPTSCTVDYSTVYFVNIEVWVEAENALGKVTSDHINFDPVYKVKPNPPHNLSVINSEELSSILKLTWTNPSIKSVIILKYNIQYRTKDASTWSQIPPEDTASTRSSFTVQDLKPFTEYVFRIRCMKEDGKGYWSDWSEEASGITYEDRPSKAPSFWYKIDPSHTQGYRTVQLVWKTLPPFEANGKILDYEVTLTRWKSHLQNYTVNATKLTVNLTNDRYLATLTVRNLVGKSDAAVLTIPACDFQATHPVMDLKAFPKDNMLWVEWTTPRESVKKYILEWCVLSDKAPCITDWQQEDGTVHRTYLRGNLAESKCYLITVTPVYADGPGSPESIKAYLKQAPPSKGPTVRTKKVGKNEAVLEWDQLPVDVQNGFIRNYTIFYRTIIGNETAVNVDSSHTEYTLSSLTSDTLYMVRMAAYTDEGGKDGPEFTFTTPKFAQGEIEAIVVPVCLAFLLTTLLGVLFCFNKRDLIKKHIWPNVPDPSKSHIAQWSPHTPPRHNFNSKDQMYSDGNFTDVSVVEIEANDKKPFPEDLKSLDLFKKEKINTEGHSSGIGGSSCMSSSRPSISSSDENESSQNTSSTVQYSTVVHSGYRHQVPSVQVFSRSESTQPLLDSEERPEDLQLVDHVDGGDGILPRQQYFKQNCSQHESSPDISHFERSKQVSSVNEEDFVRLKQQISDHISQSCGSGQMKMFQEVSAADAFGPGTEGQVERFETVGMEAATDEGMPKSYLPQTVRQGGYMPQ 6510A4409FFCF08C src_human Proto-oncogene tyrosine-protein kinase Src pp60c-src SRC SRC1 Proto-oncogene c-Src protein protein human Homo sapiens Human MGSNKSKPKDASQRRRSLEPAENVHGAGGGAFPASQTPSKPASADGHRGPSAAFAPAAAEPKLFGGFNSSDTVTSPQRAGPLAGGVTTFVALYDYESRTETDLSFKKGERLQIVNNTEGDWWLAHSLSTGQTGYIPSNYVAPSDSIQAEEWYFGKITRRESERLLLNAENPRGTFLVRESETTKGAYCLSVSDFDNAKGLNVKHYKIRKLDSGGFYITSRTQFNSLQQLVAYYSKHADGLCHRLTTVCPTSKPQTQGLAKDAWEIPRESLRLEVKLGQGCFGEVWMGTWNGTTRVAIKTLKPGTMSPEAFLQEAQVMKKLRHEKLVQLYAVVSEEPIYIVTEYMSKGSLLDFLKGETGKYLRLPQLVDMAAQIASGMAYVERMNYVHRDLRAANILVGENLVCKVADFGLARLIEDNEYTARQGAKFPIKWTAPEAALYGRFTIKSDVWSFGILLTELTTKGRVPYPGMVNREVLDQVERGYRMPCPPECPESLHDLMCQCWRKEPEERPTFEYLQAFLEDYFTSTEPQYQPGENL DIP protein P12931 original sequence version: 188 DIP protein P12931 original sequence version: 189 DIP protein P12931 original sequence version: 190 DIP protein P12931 original sequence version: 191 DIP protein P12931 original sequence version: 115 DIP protein P12931 original sequence version: 116 DIP protein P12931 original sequence version: 164 DIP protein P12931 original sequence version: 142 DIP protein P12931 original sequence version: 143 DIP protein P12931 original sequence version: 150 DIP protein P12931 original sequence version: 160 DIP protein P12931 original sequence version: 184 DIP protein P12931 original sequence version: 155 DIP protein P12931-1 original sequence version: 155 DIP protein P12931 original sequence version: 202 C1908084683E5DE8 ptpn11-cerae-protein PTPN11 (SHP-2) African Green Monkey protein protein chlae Chlorocebus aethiops Cercopithecus aethiops Grivet Green monkey An amino acid sequence which is not represented in UniProt Similar to Q06124 stat3_mouse Signal transducer and activator of transcription 3 Acute-phase response factor Stat3 Aprf protein protein mouse Mus musculus Mouse MAQWNQLQQLDTRYLEQLHQLYSDSFPMELRQFLAPWIESQDWAYAASKESHATLVFHNLLGEIDQQYSRFLQESNVLYQHNLRRIKQFLQSRYLEKPMEIARIVARCLWEESRLLQTAATAAQQGGQANHPTAAVVTEKQQMLEQHLQDVRKRVQDLEQKMKVVENLQDDFDFNYKTLKSQGDMQDLNGNNQSVTRQKMQQLEQMLTALDQMRRSIVSELAGLLSAMEYVQKTLTDEELADWKRRQQIACIGGPPNICLDRLENWITSLAESQLQTRQQIKKLEELQQKVSYKGDPIVQHRPMLEERIVELFRNLMKSAFVVERQPCMPMHPDRPLVIKTGVQFTTKVRLLVKFPELNYQLKIKVCIDKDSGDVAALRGSRKFNILGTNTKVMNMEESNNGSLSAEFKHLTLREQRCGNGGRANCDASLIVTEELHLITFETEVYHQGLKIDLETHSLPVVVISNICQMPNAWASILWYNMLTNNPKNVNFFTKPPIGTWDQVAEVLSWQFSSTTKRGLSIEQLTTLAEKLLGPGVNYSGCQITWAKFCKENMAGKGFSFWVWLDNIIDLVKKYILALWNEGYIMGFISKERERAILSTKPPGTFLLRFSESSKEGGVTFTWVEKDISGKTQIQSVEPYTKQQLNNMSFAEIIMGYKIMDATNILVSPLVYLYPDIPKEEAFGKYCRPESQEHPEADPGSAAPYLKTKFICVTPTTCSNTIDLPMSPRTLDSLMQFGNNGEGAEPSAGGQFESLTFDMDLTSECATSPM DIP protein P42227 original sequence version: 169 DIP protein P42227 original sequence version: 162 DIP protein P42227 original sequence version: 164 DIP protein P42227 original sequence version: 141 6C00626711C8012D il6st-stat3 3 4 unspecified role unspecified role bait bait required phosphotyr-767 optyr O4'-phospho-L-tyrosine (2S)-2-amino-3-(4-phosphonooxyphenyl)propanoic acid 2-azanyl-3-(4-phosphonooxyphenyl)propanoic acid MOD_RES Phosphotyrosine O4-phosphotyrosine tyrosine phosphate 2-amino-3-(4-hydroxyphenyl)propanoic acid 4'-phosphate O4'-phospho-L-tyrosine O4'-phosphorylated L-tyrosine OPTyr Phospho Phospho Tyrosinyl Phosphorylation phosphorylationy certain certain sequence position certain certain certain sequence position certain region gp130 pY3 peptide sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain region biotin tag biotin tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined chemical synthesis Chemical synthesis (Chemical synthesis) 7 unspecified role unspecified role prey prey c-terminal ST3-DN1H required to bind necessary binding region required to bind deletion analysis deletion analysis certain certain sequence position certain certain certain sequence position certain P42227:p.Asp170Ala mutation decreasing mutation decreasing interaction hotspot hotspot mutation decreasing mutation analysis mutation analysis certain certain sequence position certain certain certain sequence position certain D A Wed May 02 15:07:45 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'asp170ala' Tue Jun 30 15:55:13 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Asp170Ala' P42227:p.Lys177Ala mutation decreasing mutation decreasing interaction hotspot hotspot mutation decreasing mutation analysis mutation analysis certain certain sequence position certain certain certain sequence position certain K A Wed May 02 20:00:44 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'lys177ala' Tue Jun 30 19:04:39 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Lys177Ala' chlae-cos_1 Chlorocebus aethiops COS-1 cells Grivet cos_1 CV-1 Simian cells (cercopithecus aethiops) transformed by SV40 physical association physical association Fig. 4b, Fig. 5b The phosphorylated peptide-containing sequences surrounding the third tyrosine (pY3), coprecipitated with full-length Stat3 as well as the N-domain mutant ST3-DN. Surprisingly, binding was diminished in ST3-DN1H and was totally abolished in ST3-DN2H, ST3-DN3H, and ST3-DN4H Phosphorylation of the tyrosine-767 in the gp130 peptide was required for the interaction. Binding of ST3-DN1H (lacking the first helix in the coiled-coil domain) was less than the full length protein and the deletion mutants ST3-DN2H (260-770), ST3-DN3H (296-770) and ST3-DN4H (320-770) did not bind at all. il6st-ptpn1 3 4 unspecified role unspecified role bait bait region gp130 pY2 peptide sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain region optyr O4'-phospho-L-tyrosine (2S)-2-amino-3-(4-phosphonooxyphenyl)propanoic acid 2-azanyl-3-(4-phosphonooxyphenyl)propanoic acid MOD_RES Phosphotyrosine O4-phosphotyrosine tyrosine phosphate 2-amino-3-(4-hydroxyphenyl)propanoic acid 4'-phosphate O4'-phospho-L-tyrosine O4'-phosphorylated L-tyrosine OPTyr Phospho Phospho Tyrosinyl Phosphorylation phosphorylationy certain certain sequence position certain certain certain sequence position certain region biotin tag biotin tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined chemical synthesis Chemical synthesis (Chemical synthesis) 6 unspecified role unspecified role prey prey chlae-cos_1 Chlorocebus aethiops COS-1 cells Grivet cos_1 CV-1 Simian cells (cercopithecus aethiops) transformed by SV40 physical association physical association Fig. 4c A strong binding of endogenous SHP-2 to pY2, but not to Y3 and pY3, was detected in COS-1 cells, Phosphorylation of the tyrosine-759 in the gp130 peptide was required for the interaction. src-stat3 1 5 enzyme enzyme unspecified role unspecified role spofr-sf_21 Spodoptera frugiperda insect cells sf_21 Insect cells derived from immature ovaries of fall armyworm Spodoptera frugiperda pupae. full name: IPLB-SF-21-AE cells are susceptible to Baculovirus infection. 7 enzyme target enzyme target substrate substrate unspecified role unspecified role c-terminal ST3-DN1H sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain spofr-sf_9 Spodoptera frugiperda insect cells Fall armyworm sf_9 insect cells Derived from pupal ovarian tissue of the fall armyworm Cloned from the parent cell line, IPLB-SF-21-AE (SF-21) derived from immature ovaries of fall armyworm Spodoptera frugiperda pupae cells are susceptible to Baculovirus infection enzymatic reaction enzymatic reaction Biochemical reaction Biochemical reaction Fig. 6a mutant Stat3 which lacks the N-domain and helix a1 (b-ST3-DN1H) and full-length Stat3 (b-ST3-FL) were expressed and purified in the baculovirus expression system. The recombinant proteins were phosphorylated with Src kinase in vitro, and tyrosine phosphorylation and DNA-binding activity were examined. As shown in Fig. 6, strong Tyr phosphorylation by Src was detected in both the wild type and the deletion mutant. Stat3 is phosphorylated by Src on Tyr-705. The authors do not specify the species of origin of Src. However, the website of Upstate Biotechnology from where the enzyme was purchased shows an active human Src in the catalogue. il6st-stat3-1 2 4 unspecified role unspecified role bait bait region gp130 pY3 peptide sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain region biotin tag biotin tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined required phosphotyr-767 optyr O4'-phospho-L-tyrosine (2S)-2-amino-3-(4-phosphonooxyphenyl)propanoic acid 2-azanyl-3-(4-phosphonooxyphenyl)propanoic acid MOD_RES Phosphotyrosine O4-phosphotyrosine tyrosine phosphate 2-amino-3-(4-hydroxyphenyl)propanoic acid 4'-phosphate O4'-phospho-L-tyrosine O4'-phosphorylated L-tyrosine OPTyr Phospho Phospho Tyrosinyl Phosphorylation phosphorylationy certain certain sequence position certain certain certain sequence position certain chemical synthesis Chemical synthesis (Chemical synthesis) 7 unspecified role unspecified role prey prey c-terminal ST3-DN1H required to bind necessary binding region required to bind deletion analysis deletion analysis certain certain sequence position certain certain certain sequence position certain spofr-sf_9 Spodoptera frugiperda insect cells Fall armyworm sf_9 insect cells Derived from pupal ovarian tissue of the fall armyworm Cloned from the parent cell line, IPLB-SF-21-AE (SF-21) derived from immature ovaries of fall armyworm Spodoptera frugiperda pupae cells are susceptible to Baculovirus infection direct interaction direct interaction Fig. 7 Phosphorylation of the tyrosine-767 in the gp130 peptide was required for the interaction. baculovirus-produced b-ST3- FL, b-ST3-DN1H, and b-ST3-DN4H (deleting four a-helices) were purified, and their receptor-binding abilities were analyzed by the peptide-binding assay. Similar to the results obtained in vivo (Fig. 4), we observed a strong interaction between wild-type Stat3 and the gp130-derived phosphopeptide pY3, but not Y3 (Fig. 7, lanes 1 and 2). The binding was significantly diminished for b-ST3-DN1H The binding of the b-ST3-DN1H (lacking the first helix in the coiled coil domain) protein was not as strong as the binding of the full length protein and ST3-DN4H (320-770) did not bind at all.