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/ Genetically Encoded Chemical Probes in Cells Reveal the Binding Path of Urocortin-I to CRF Class B GPCR. Genetically Encoded Chemical Probes in Cells Reveal the Binding Path of Urocortin-I to CRF Class B GPCR. Cell (0092-8674) 2013 imex curation Coin I., Katritch V., Sun T., Xiang Z., Siu F.Y., Beyermann M., Stevens R.C., Wang L. lwang@salk.edu 18-Feb-2014: Contacted by IntAct-Help. Only protein-protein interactions in vitro In vitro crosslink cross-linking study crosslink western blot western blot Immuno blot Immuno blot Coin I., Katritch V., Sun T., Xiang Z., Siu F.Y., Beyermann M., Stevens R.C., Wang L. Receptor mutants containing the photoactivatable unnatural amino acid p-azido-phenylalanine (Azi), are expressed in cells and incubated with the native ligand. Upon light activation, the ligand is covalently captured by the receptor only if Azi lies in its proximity. Cells expressing the Azi-CRF1R were incubated with agonist and irradiated at the binding equilibrium with UV light (365 nm). Crosslinked samples were resolved with SDS-PAGE and immunoblotted. lwang@salk.edu Accepted 2014-JAN-29 by PPORRAS Cell (0092-8674) 2013 Only protein-protein interactions imex curation Genetically Encoded Chemical Probes in Cells Reveal the Binding Path of Urocortin-I to CRF Class B GPCR. Genetically Encoded Chemical Probes in Cells Reveal the Binding Path of Urocortin-I to CRF Class B GPCR. Cell (0092-8674) 2013 imex curation Coin I., Katritch V., Sun T., Xiang Z., Siu F.Y., Beyermann M., Stevens R.C., Wang L. lwang@salk.edu 18-Feb-2014: Contacted by IntAct-Help. Only protein-protein interactions in vitro In vitro crosslink cross-linking study crosslink western blot western blot Immuno blot Immuno blot Coin I., Katritch V., Sun T., Xiang Z., Siu F.Y., Beyermann M., Stevens R.C., Wang L. Crosslinking agent: tris(2-carboxyethyl)phosphine lwang@salk.edu Accepted 2014-JAN-29 by PPORRAS Cell (0092-8674) 2013 Only protein-protein interactions imex curation astressin Astressin small molecule small molecule chemical synthesis Chemical synthesis (Chemical synthesis) crfr1_rat Corticotropin-releasing factor receptor 1 Crhr1 Crhr Corticotropin-releasing hormone receptor 1 protein protein rat Rattus norvegicus (Rat) Rat MGRRPQLRLVKALLLLGLNPVSTSLQDQRCENLSLTSNVSGLQCNASVDLIGTCWPRSPAGQLVVRPCPAFFYGVRYNTTNNGYRECLANGSWAARVNYSECQEILNEEKKSKVHYHVAVIINYLGHCISLVALLVAFVLFLRLRSIRCLRNIIHWNLISAFILRNATWFVVQLTVSPEVHQSNVAWCRLVTAAYNYFHVTNFFWMFGEGCYLHTAIVLTYSTDRLRKWMFVCIGWGVPFPIIVAWAIGKLHYDNEKCWFGKRPGVYTDYIYQGPMILVLLINFIFLFNIVRILMTKLRASTTSETIQYRKAVKATLVLLPLLGITYMLFFVNPGEDEVSRVVFIYFNSFLESFQGFFVSVFYCFLNSEVRSAIRKRWRRWQDKHSIRARVARAMSIPTSPTRVSFHSIKQSTAV 48D6704B31D4C013 ucn1_rat Urocortin Ucn Corticotensin protein protein rat Rattus norvegicus (Rat) Rat MRQRGRATLLVALLLLVQLRPESSQWSPAAAAANVVQDPNLRWNPGVRNQGGGVRALLLLLAERFPRRAGSEPAGERQRRDDPPLSIDLTFHLLRTLLELARTQSQRERAEQNRIIFDSVGK 9F0AF834CBFFCE74 crhr1-ucn 1 5 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) 4 anti tag western anti tag western blot anti tag western western blot western blot Immuno blot Immuno blot western blot western blot Immuno blot Immuno blot unspecified role unspecified role neutral component neutral component c-terminal flag tag flag tag FLAG-tagged FLAG DYKDDDDKV epitope tag DYKDDDDKV epitope tag FLAG FLAG-tagged c-term range C-terminal range c-term range c-term range C-terminal range c-term range human-293t Homo sapiens 293 cells transformed with SV40 large T antigen 293t 293 cells expressing SV40 large T antigen. The original designation of this cell line was 293tsA1609neo. It is not in Cabri,ATCC or HyperCLDB. physical association physical association Fig. 1E, 2, 3A, Supp. Fig. 2, 3A Azi was systematically incorporated throughout all positions of the juxta-membrane region (J region) of CRF1R, including the three extracellular loops and the extracellular halves of the trans-membrane helices, as well as the hinge region joining helix I to the extra-cellular domain. Only 6 mutants (Asp-49, Asn-166, Ala-186, Arg-189, Thr-201, and Gly-356) failed to express. 35 mutated residues intensely photo-crosslinking, a further 104 positions showed lower levels of crosslinking. The addition of Dithiothreitol was necessary to achieve complete separation of non-crosslinked Ucn1 from the reaction mixture astressin-crhr1 1 5 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) 4 anti tag western anti tag western blot anti tag western western blot western blot Immuno blot Immuno blot western blot western blot Immuno blot Immuno blot unspecified role unspecified role neutral component neutral component c-terminal flag tag flag tag FLAG-tagged FLAG DYKDDDDKV epitope tag DYKDDDDKV epitope tag FLAG FLAG-tagged c-term range C-terminal range c-term range c-term range C-terminal range c-term range P35353:p.Asn123Xaa mutation mutation certain certain sequence position certain certain certain sequence position certain N X Asparagine 123 was mutated to unconventional amino acid p-azido-phenylalanine (Azi). Fri Jul 22 11:07:32 BST 2016 This feature has been corrected as a result of our quality control procedures. The original label was 'asn123unc' Wed May 02 16:55:47 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'asn123xaa' Tue Jun 30 19:52:33 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Asn123Xaa' human-293t Homo sapiens 293 cells transformed with SV40 large T antigen 293t 293 cells expressing SV40 large T antigen. The original designation of this cell line was 293tsA1609neo. It is not in Cabri,ATCC or HyperCLDB. 3 inhibitor inhibitor unspecified role unspecified role physical association physical association Fig. 1E crhr1-ucn-1 2 5 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) 4 anti tag western anti tag western blot anti tag western western blot western blot Immuno blot Immuno blot western blot western blot Immuno blot Immuno blot unspecified role unspecified role neutral component neutral component c-terminal flag tag flag tag FLAG-tagged FLAG DYKDDDDKV epitope tag DYKDDDDKV epitope tag FLAG FLAG-tagged c-term range C-terminal range c-term range c-term range C-terminal range c-term range human-293t Homo sapiens 293 cells transformed with SV40 large T antigen 293t 293 cells expressing SV40 large T antigen. The original designation of this cell line was 293tsA1609neo. It is not in Cabri,ATCC or HyperCLDB. physical association physical association Fig. 4C Cys86-, Cys88-, Cys90-, Cys92-, and Cys94-Ucn1 mutants were crosslinked to CRHR1 with the unnatural amino acid p-2′-fluoroacetyl-phenylalanine substituted at 23 different sites. Each Cys-Ucn1 analog reacted with a subset of Ffact-CRF1R mutants. No intense signals were detected with Cys6- or Cys10-Ucn1. Cys88-Ucn1 reacted with Gln-273-CRF1R Cys92-Ucn1 reacted with Leu-329-CRF1R Cys88-Ucn1 and Cys92-Ucn1 reacted with Phe330-CRF1R Cys92-Ucn1 reacted with Asn-333-CRF1R Cys92-Ucn1 reacted with Ile-345-CRF1R Cys92-Ucn1 reacted with Asn-348-CRF1R Cys92-Ucn1 and Cys94-Ucn1 reacted with Ser349-CRF1R crhr1-ucn-2 1 5 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 P55090:p.His92Cys mutation mutation certain certain sequence position certain certain certain sequence position certain H C Wed May 02 15:15:22 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'his92cys' Tue Jun 30 17:44:16 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.His92Cys' P55090:p.Leu94Cys mutation mutation certain certain sequence position certain certain certain sequence position certain L C Wed May 02 20:12:14 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'leu94cys' Tue Jun 30 17:33:53 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Leu94Cys' P55090:p.Ser86Cys mutation mutation certain certain sequence position certain certain certain sequence position certain S C Wed May 02 14:40:56 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'ser86cys' Tue Jun 30 18:20:52 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Ser86Cys' P55090:p.Asp88Cys mutation mutation certain certain sequence position certain certain certain sequence position certain D C Wed May 02 19:42:10 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'asp88cys' Tue Jun 30 16:19:46 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Asp88Cys' P55090:p.Thr90Cys mutation mutation certain certain sequence position certain certain certain sequence position certain T C Wed May 02 20:17:39 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'thr90cys' Tue Jun 30 17:20:25 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Thr90Cys' chemical synthesis Chemical synthesis (Chemical synthesis) 4 anti tag western anti tag western blot anti tag western western blot western blot Immuno blot Immuno blot western blot western blot Immuno blot Immuno blot unspecified role unspecified role neutral component neutral component c-terminal flag tag flag tag FLAG-tagged FLAG DYKDDDDKV epitope tag DYKDDDDKV epitope tag FLAG FLAG-tagged c-term range C-terminal range c-term range c-term range C-terminal range c-term range P35353:p.Tyr116Xaa mutation mutation certain certain sequence position certain certain certain sequence position certain Y X Tyrosine 116 was mutated to unconventional amino acid p-azido-phenylalanine (Azi). Fri Jul 22 11:06:23 BST 2016 This feature has been corrected as a result of our quality control procedures. The original label was 'tyr116unc' Wed May 02 16:35:00 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'tyr116xaa' Tue Jun 30 16:32:22 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Tyr116Xaa' P35353:p.Tyr73Xaa mutation mutation certain certain sequence position certain certain certain sequence position certain Y X Tyrosine 73 was mutated to unconventional amino acid p-azido-phenylalanine (Azi). Fri Jul 22 10:59:47 BST 2016 This feature has been corrected as a result of our quality control procedures. The original label was 'tyr73unc' Wed May 02 18:45:58 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'tyr73xaa' Tue Jun 30 19:34:58 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Tyr73Xaa' P35353:p.Lys262Xaa mutation mutation certain certain sequence position certain certain certain sequence position certain K X Lysine 262 was mutated to unconventional amino acid p-azido-phenylalanine (Azi). Fri Jul 22 11:13:40 BST 2016 This feature has been corrected as a result of our quality control procedures. The original label was 'lys262unc' Wed May 02 20:15:26 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'lys262xaa' Tue Jun 30 19:46:01 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Lys262Xaa' human-293t Homo sapiens 293 cells transformed with SV40 large T antigen 293t 293 cells expressing SV40 large T antigen. The original designation of this cell line was 293tsA1609neo. It is not in Cabri,ATCC or HyperCLDB. physical association physical association Supp. Fig. 3B