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 architecture and functional model of the complete yeast ESCRT-I heterotetramer. Molecular architecture and functional model of the complete yeast ESCRT-I heterotetramer. Cell (0092-8674) 2007 imex curation Kostelansky MS., Schluter C., Tam YY., Lee S., Ghirlando R., Beach B., Conibear E., Hurley JH. hurley@helix.nih.gov 07-Aug-2007: Contacted by JYOTI. orchard Only protein-protein interactions 7 in vitro In vitro molecular sieving molecular sieving Gel Filtration Size Exclusion Chromatography Size Exclusion Chromatography Gel Filtration Sizing column predetermined predetermined participant predetermined Kostelansky MS., Schluter C., Tam YY., Lee S., Ghirlando R., Beach B., Conibear E., Hurley JH. Superdex 200 used for molecular seiving. hurley@helix.nih.gov Accepted 2007-AUG-4 by ORCHARD. 07-Aug-2007: Contacted by JYOTI. Cell (0092-8674) 2007 orchard Only protein-protein interactions 7 imex curation Molecular architecture and functional model of the complete yeast ESCRT-I heterotetramer. Molecular architecture and functional model of the complete yeast ESCRT-I heterotetramer. Cell (0092-8674) 2007 imex curation Kostelansky MS., Schluter C., Tam YY., Lee S., Ghirlando R., Beach B., Conibear E., Hurley JH. hurley@helix.nih.gov 07-Aug-2007: Contacted by JYOTI. orchard Only protein-protein interactions 7 yeasx Saccharomyces cerevisiae (Baker's yeast) Baker's yeast anti bait coip anti bait coimmunoprecipitation anti bait coip anti tag western anti tag western blot anti tag western Kostelansky MS., Schluter C., Tam YY., Lee S., Ghirlando R., Beach B., Conibear E., Hurley JH. hurley@helix.nih.gov Accepted 2007-AUG-4 by ORCHARD. 07-Aug-2007: Contacted by JYOTI. Cell (0092-8674) 2007 orchard Only protein-protein interactions 7 imex curation Molecular architecture and functional model of the complete yeast ESCRT-I heterotetramer. Molecular architecture and functional model of the complete yeast ESCRT-I heterotetramer. Cell (0092-8674) 2007 imex curation Kostelansky MS., Schluter C., Tam YY., Lee S., Ghirlando R., Beach B., Conibear E., Hurley JH. hurley@helix.nih.gov 07-Aug-2007: Contacted by JYOTI. orchard Only protein-protein interactions 7 in vitro In vitro x-ray diffraction x-ray crystallography X-ray X-ray x-ray diffraction predetermined predetermined participant predetermined Kostelansky MS., Schluter C., Tam YY., Lee S., Ghirlando R., Beach B., Conibear E., Hurley JH. hurley@helix.nih.gov Accepted 2007-AUG-4 by ORCHARD. Cell (0092-8674) 2007 orchard Only protein-protein interactions 7 imex curation Molecular architecture and functional model of the complete yeast ESCRT-I heterotetramer. Molecular architecture and functional model of the complete yeast ESCRT-I heterotetramer. Cell (0092-8674) 2007 imex curation Kostelansky MS., Schluter C., Tam YY., Lee S., Ghirlando R., Beach B., Conibear E., Hurley JH. hurley@helix.nih.gov 07-Aug-2007: Contacted by JYOTI. orchard Only protein-protein interactions 7 in vitro In vitro solution sedimentati cosedimentation in solution solution sedimentati predetermined predetermined participant predetermined Kostelansky MS., Schluter C., Tam YY., Lee S., Ghirlando R., Beach B., Conibear E., Hurley JH. hurley@helix.nih.gov Accepted 2007-AUG-4 by ORCHARD. 07-Aug-2007: Contacted by JYOTI. Cell (0092-8674) 2007 orchard Only protein-protein interactions 7 imex curation stp22_yeast Suppressor protein STP22 of temperature-sensitive alpha-factor receptor and arginine permease STP22 VPS23 YCL008C YCL8C Vacuolar protein sorting-associated protein 23 ESCRT-I complex subunit VPS23 protein protein yeast Saccharomyces cerevisiae Baker's yeast MSANGKISVPEAVVNWLFKVIQPIYNDGRTTFHDSLALLDNFHSLRPRTRVFTHSDGTPQLLLSIYGTISTGEDGSSPHSIPVIMWVPSMYPVKPPFISINLENFDMNTISSSLPIQEYIDSNGWIALPILHCWDPAAMNLIMVVQELMSLLHEPPQDQAPSLPPKPNTQLQQEQNTPPLPPKPKSPHLKPPLPPPPPPQPASNALDLMDMDNTDISPTNHHEMLQNLQTVVNELYREDVDYVADKILTRQTVMQESIARFHEIIAIDKNHLRAVEQAIEQTMHSLNAQIDVLTANRAKVQQFSSTSHVDDEDVNSIAVAKTDGLNQLYNLVAQDYALTDTIECLSRMLHRGTIPLDTFVKQGRELARQQFLVRWHIQRITSPLS FEDE3BE79F063BCE FEDE3BE79F063BCE vps28_yeast Vacuolar protein sorting-associated protein 28 VPS28 VPT28 YPL065W LPE5W ESCRT-I complex subunit VPS28 protein protein yeast Saccharomyces cerevisiae Baker's yeast MQKHNIKLNQNQDISQLFHDEVPLFDNSITSKDKEVIETLSEIYSIVITLDHVEKAYLKDSIDDTQYTNTVDKLLKQFKVYLNSQNKEEINKHFQSIEAFCDTYNITASNAITRLERGIPITAEHAISTTTSAPSGDNKQSSSSDKKFNAKYVAEATGNFITVMDALKLNYNAKDQLHPLLAELLISINRVTRDDFENRSKLIDWIVRINKLSIGDTLTETQIRELLFDLELAYKSFYALLD DE091D3F60C9D132 DE091D3F60C9D132 mvb12_yeast Multivesicular body sorting factor 12 YGR206W G7740 12 kDa multivesicular body sorting factor ESCRT-I complex subunit MVB12 MVB12 protein protein yeast Saccharomyces cerevisiae Baker's yeast MNNNVEELLRRIPLYNKYGKDFPQETVTRFQMPEFKLPALQPTRDLLCPWYEECDNITKVCQLHDSSNKKFDQWYKEQYLSKKPPGIVGNTLLSPSRKDNS E48B085E0FACBAC8 E48B085E0FACBAC8 srn2_yeast Protein SRN2 SRN2 ESCRT-I complex subunit VPS37 SRN10 Vacuolar protein sorting-associated protein 37 VPS37 L2958 L9233.5 YLR119W protein protein yeast Saccharomyces cerevisiae Baker's yeast MKVKATKLRIKQRRKNKGLNISRLDIIRAEMDVVPSPGLPEKVNEKSKNIPLPEGINLLSSKEIIDLIQTHRHQLELYVTKFNPLTDFAGKIHAFRDQFKQLEENFEDLHEQKDKVQALLENCRILESKYVASWQDYHSEFSKKYGDIALKKKLEQNTKKLDEESSQLETTTRSIDSADDLDQFIKNYLDIRTQYHLRREKLATWDKQGNLKY 4E99806CED03E345 4E99806CED03E345 stp22-srn2-1 3 6 unspecified role unspecified role neutral component neutral component ecoli Escherichia coli (strain K12) 8 unspecified role unspecified role neutral component neutral component ecoli Escherichia coli (strain K12) 5 unspecified role unspecified role neutral component neutral component ecoli Escherichia coli (strain K12) 7 unspecified role unspecified role neutral component neutral component ecoli Escherichia coli (strain K12) physical association physical association working 23.3 percent, free 31.5 percent 2.70 A 2, 3 Table S1 The structure of a heterotetrameric core complex of ESCRT-I was determined at a nominal resolution of 2.7 Figure 2A Table S1. mvb12 vps23 2 7 unspecified role unspecified role prey prey region gfp tag green fluorescent protein tag GFP green fluorescent protein GFP green fluorescent protein gfp tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined P42939:p.[Leu47Asp;Ile57Asp] mutation decreasing mutation decreasing interaction hotspot hotspot mutation decreasing certain certain sequence position certain certain certain sequence position certain I D certain certain sequence position certain certain certain sequence position certain L D Fri Jul 22 11:09:20 BST 2016 This feature has been corrected as a result of our quality control procedures. The original label was 'ile57asp,leu47asp' Wed May 02 16:50:04 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'leu47asp,ile57asp' Tue Jun 30 18:29:22 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.[Leu47Asp;Ile57Asp]' 5 unspecified role unspecified role bait bait region ha tag ha tag YPYDVPDYA epitope tag YPYDVPDYA epitope tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined P25604:p.Met254Asp mutation decreasing mutation decreasing interaction hotspot hotspot mutation decreasing certain certain sequence position certain certain certain sequence position certain M D Wed May 02 20:31:34 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'met254asp' Tue Jun 30 19:24:31 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Met254Asp' physical association physical association 1E, 5D, S5D The stoichiometry was determined as percent of the protein in the cell lysate. The copurification of Mvb12 and Vps23 required Vps37. Mvb12 and Vps23 copurified efficiently, with immunoprecipitation of 80 percent of the cellular Vps23 resulting in the coprecipitation of a nearly equal proportion 70 percent of cellular Mvb12 stp22-srn2-2 1 8 unspecified role unspecified role neutral component neutral component ecoli Escherichia coli (strain K12) 6 unspecified role unspecified role neutral component neutral component ecoli Escherichia coli (strain K12) 5 unspecified role unspecified role neutral component neutral component ecoli Escherichia coli (strain K12) 7 unspecified role unspecified role neutral component neutral component ecoli Escherichia coli (strain K12) physical association physical association 4B The full-length heterotetrameric ESCRT-I complex consisting of intact Vps23, Vps28, Vps37, and Mvb12 runs on gel filtration chromatography with an apparent molecular weight of 340 kDa ESCRT-I complex consisting of intact Vps23, Vps28, Vps37, and Mvb12 runs on gel filtration chromatography with an apparent molecular weight of 340 kDa stp22-srn2-3 4 8 unspecified role unspecified role neutral component neutral component ecoli Escherichia coli (strain K12) 5 unspecified role unspecified role neutral component neutral component ecoli Escherichia coli (strain K12) 7 unspecified role unspecified role neutral component neutral component ecoli Escherichia coli (strain K12) 6 unspecified role unspecified role neutral component neutral component ecoli Escherichia coli (strain K12) physical association physical association 4B Sedimentation equilibrium centrifugation analysis revealed that full-length ESCRT-I sample has an experimental molecular mass of 117 kDa ESCRT-I complex has a molecular mass of 117 Kda. vps37-vps23-1 2 8 unspecified role unspecified role prey prey region myc tag myc tag EUKLISEED epitope tag EUKLISEED epitope tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined 5 unspecified role unspecified role bait bait P25604:p.Met254Asp mutation decreasing mutation decreasing interaction hotspot hotspot mutation decreasing certain certain sequence position certain certain certain sequence position certain M D Wed May 02 15:13:56 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'met254asp' Tue Jun 30 16:45:59 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Met254Asp' region ha tag ha tag YPYDVPDYA epitope tag YPYDVPDYA epitope tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined physical association physical association 5D Consistent with the in vivo localization data, Vps23M254D and Mvb12L47D/I57D mutations had the greatest effect on the incorporation of Mvb12 into the complex. Vps23M254D blocked the Vps23-Mvb12 interaction and reduced, but did not prevent, Vps23 binding to Vps37, whereas Mvb12L47D/I57D showed reduced binding to both Vps23 and Vps37. Vps37L67D-containing complexes contained wild-type levels of Mvb12 and Vps23, in keeping with the MVB localization of both proteins. vps37-mvb12 2 7 unspecified role unspecified role prey prey region yfp tag yellow fluorescent protein tag YFP yellow fluorescent protein yellow fluorescent protein YFP yfp tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined P42939:p.[Leu47Asp;Ile57Asp] mutation decreasing mutation decreasing interaction hotspot hotspot mutation decreasing certain certain sequence position certain certain certain sequence position certain I D certain certain sequence position certain certain certain sequence position certain L D Fri Jul 22 10:58:17 BST 2016 This feature has been corrected as a result of our quality control procedures. The original label was 'ile57asp,leu47asp' Wed May 02 14:31:21 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'leu47asp,ile57asp' Tue Jun 30 16:32:35 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.[Leu47Asp;Ile57Asp]' 8 unspecified role unspecified role bait bait region myc tag myc tag EUKLISEED epitope tag EUKLISEED epitope tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined physical association physical association 5E Consistent with the in vivo localization data, Vps23M254D and Mvb12L47D/I57D mutations had the greatest effect on the incorporation of Mvb12 into the complex. Vps23M254D blocked the Vps23-Mvb12 interaction and reduced, but did not prevent, Vps23 binding to Vps37, whereas Mvb12L47D/I57D showed reduced binding to both Vps23 and Vps37. Vps37L67D-containing complexes contained wild-type levels of Mvb12 and Vps23, in keeping with the MVB localization of both proteins. mvb12 vps23-1 2 5 unspecified role unspecified role bait bait region ha tag ha tag YPYDVPDYA epitope tag YPYDVPDYA epitope tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined 7 unspecified role unspecified role prey prey region yfp tag yellow fluorescent protein tag YFP yellow fluorescent protein yellow fluorescent protein YFP yfp tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined P42939:p.[Leu47Asp;Ile57Asp] mutation decreasing mutation decreasing interaction hotspot hotspot mutation decreasing certain certain sequence position certain certain certain sequence position certain I D certain certain sequence position certain certain certain sequence position certain L D Fri Jul 22 10:55:18 BST 2016 This feature has been corrected as a result of our quality control procedures. The original label was 'ile57asp,leu47asp' Wed May 02 16:14:58 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'leu47asp,ile57asp' Tue Jun 30 15:40:12 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.[Leu47Asp;Ile57Asp]' physical association physical association 5E Consistent with the in vivo localization data, Vps23M254D and Mvb12L47D/I57D mutations had the greatest effect on the incorporation of Mvb12 into the complex. Vps23M254D blocked the Vps23-Mvb12 interaction and reduced, but did not prevent, Vps23 binding to Vps37, whereas Mvb12L47D/I57D showed reduced binding to both Vps23 and Vps37