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/ http://www.ebi.ac.uk/ saravanan-2009-1 Targeting of the oxysterol-binding protein ORP3a to the endoplasmic reticulum relies on the plant VAP33 homolog PVA12. in vitro In vitro pull down pull down anti tag western anti tag western blot anti tag western Saravanan R., Slabaugh E., Singh VR., Lapidus LJ., Haas T., Brandizzi F. Recombinant purified His6 tagged ORP3a was incubated with liposomes containing stigmasterol and sitosterol the liposomes were purified and then western blot analysis of the pellets with anti-His serum was carried out. bradizz@msu.edu Plant J. (0960-7412) 2009 Only protein-protein interactions imex curation saravanan-2009-2 Targeting of the oxysterol-binding protein ORP3a to the endoplasmic reticulum relies on the plant VAP33 homolog PVA12. tobac-leaf Nicotiana tabacum (tobacco) leaf leaf leaf bifc bimolecular fluorescence complementation bifc tag fluorescence tag visualisation by fluorescence tag fluorescence Saravanan R., Slabaugh E., Singh VR., Lapidus LJ., Haas T., Brandizzi F. Recombinant purified His6 tagged ORP3a was incubated with liposomes containing stigmasterol and sitosterol the liposomes were purified and then western blot analysis of the pellets with anti-His serum was carried out. bradizz@msu.edu Plant J. (0960-7412) 2009 Only protein-protein interactions imex curation saravanan-2009-3 Targeting of the oxysterol-binding protein ORP3a to the endoplasmic reticulum relies on the plant VAP33 homolog PVA12. in vitro In vitro interaction detect interaction detection method interaction detect anti tag western anti tag western blot anti tag western Saravanan R., Slabaugh E., Singh VR., Lapidus LJ., Haas T., Brandizzi F. Recombinant purified His6 tagged ORP3a was incubated with liposomes containing stigmasterol and sitosterol the liposomes were purified and then western blot analysis of the pellets with anti-His serum was carried out. bradizz@msu.edu Plant J. (0960-7412) 2009 Only protein-protein interactions imex curation saravanan-2009-4 Targeting of the oxysterol-binding protein ORP3a to the endoplasmic reticulum relies on the plant VAP33 homolog PVA12. in vitro In vitro anti tag coip anti tag coimmunoprecipitation anti tag coip predetermined predetermined participant predetermined Saravanan R., Slabaugh E., Singh VR., Lapidus LJ., Haas T., Brandizzi F. Recombinant purified His6 tagged ORP3a was incubated with liposomes containing stigmasterol and sitosterol the liposomes were purified and then western blot analysis of the pellets with anti-His serum was carried out. bradizz@msu.edu Plant J. (0960-7412) 2009 Only protein-protein interactions imex curation vap12_arath Vesicle-associated protein 1-2 PVA12 VAP12 Plant VAP homolog 12 VAMP-associated protein 1-2 T14P1.5 At2g45140 protein protein arath Arabidopsis thaliana (Mouse-ear cress) Mouse-ear cress MSNELLTIDPVDLQFPFELKKQISCSLYLGNKTDNYVAFKVKTTNPKKYCVRPNTGVVHPRSSSEVLVTMQAQKEAPADLQCKDKFLLQCVVASPGATPKDVTHEMFSKEAGHRVEETKLRVVYVAPPRPPSPVREGSEEGSSPRASVSDNGNASDFTAAPRFSADRVDAQDNSSEARALVTKLTEEKNSAVQLNNRLQQELDQLRRESKRSKSGGIPFMYVLLVGLIGLILGYIMKRT orp3a_arath Oxysterol-binding protein-related protein 3A UNE18 OSBP-related protein 3A Protein UNFERTILIZED EMBRYO SAC 18 T7H20.150 ORP3A At5g02100 protein protein arath Arabidopsis thaliana (Mouse-ear cress) Mouse-ear cress MASNDPKNGGGFFASLASSITNFGSAMSKSVNGLMGYEGLEVINPEGSTDDAEEEAGRGRWKQEERDGYWKMMQKYIGSDVTSMVTLPVIIFEPMTMLQKMAELMEYSYLLDMADKTEDPYMRMVYASSWAISVYYAYQRTWKPFNPILGETYEMTNHNGINFIAEQVCHHPPMSAGHAENEHFAYDCTSKLKTKFLGNSIDVYPVGRTRVTLKRDGVVLDLVPPLTKVHNLIFGRTWVDSPGEMVMTNLTTGDKVVLYFQPCGWFGSGRYEVDGYVYNSAEEPKMLMTGKWNESLSYQPCDAEGEPLPGTELKEVWKVAEAPKNDKYQYTHFAHKINSFDTAPKKLLSSDSRLRPDRYALEMGDMSKSGFEKSSLEDRQRAEKKSREEKGQKFAPKWFDETEEVTPTPWGDLEVYQFNGKYSVHRATAENSEDTTDVKLTQFNPWQFQDLSA sitosterol stigmast-5-en-3β-ol small molecule small molecule chemical synthesis Chemical synthesis (Chemical synthesis) pva12-orp3a PVA12 interacts with ORP3a 802178 n/a 802182 unspecified role unspecified role bait bait region gst tag glutathione s tranferase tag glutathione S-tranferase tag glutathione S-tranferase tag gst tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined false ecoli Escherichia coli (strain K12) n/a 802183 unspecified role unspecified role prey prey c-terminal cfp tag cyan fluorescent protein tag CFP cyan fluorescent protein cyan fluorescent protein CFP cfp tag c-term range C-terminal range c-term range c-term range C-terminal range c-term range false tobac Nicotiana tabacum (Common tobacco) Common tobacco physical association physical association false false false Figure 2b The authors state that the "cytosolic domain" of PVA12 protein was used in the GST tagged construct. However the co-ordiantes of the protein region are not provided. The UniProt record does not clearly define this region. Therefore this information could not be annotated in the entry. pva12-orp3a-2 PVA12 interacts with ORP3a 802178 n/a 802183 unspecified role unspecified role prey prey region his tag his tag Hexa-His-tag 6-His-tag Histidine-tag Hexa-His-tag Histidine-tag 6-His-tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined false Q9LZM1:p.[Trp398Ala;Phe399Ala;Asp400Gly;Glu401Gly;Glu403Gly] trp398ala phe399ala asp400gly glu401gly glu403g;y mutation disrupting mutation disrupting interaction mutation disrupting certain certain sequence position certain certain certain sequence position certain false certain certain sequence position certain certain certain sequence position certain false certain certain sequence position certain certain certain sequence position certain false certain certain sequence position certain certain certain sequence position certain false certain certain sequence position certain certain certain sequence position certain false ecoli Escherichia coli (strain K12) n/a 802182 unspecified role unspecified role bait bait region gst tag glutathione s tranferase tag glutathione S-tranferase tag glutathione S-tranferase tag gst tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined false ecoli Escherichia coli (strain K12) direct interaction direct interaction false false false Figure 4 The authors state that the "cytosolic domain" of PVA12 protein was used in the GST tagged construct. However the co-ordiantes of the protein region are not provided. The UniProt record does not clearly define this region. Therefore this information could not be annotated in the entry. The authors conclude "our results indicate that disruption of the PVA12-ORP3a398AAGGTG403 interaction is specific to the mutations of the WFDE region, rather than resulting from secondary effects of the mutations on protein folding". orp3a-pva12 PVA12 interacts with ORP3a 802179 n/a 802182 unspecified role unspecified role neutral component neutral component n-terminal N-terminal YFP yfp tag yellow fluorescent protein tag YFP yellow fluorescent protein yellow fluorescent protein YFP yfp tag n-term range n-terminal range n-term range n-term range n-terminal range n-term range false n/a 802183 unspecified role unspecified role neutral component neutral component c-terminal C-terminal YFP yfp tag yellow fluorescent protein tag YFP yellow fluorescent protein yellow fluorescent protein YFP yfp tag c-term range C-terminal range c-term range c-term range C-terminal range c-term range false Q9LZM1:p.[Trp398Ala;Phe399Ala;Asp400Gly;Glu401Gly;Glu403Gly] trp398ala phe399ala asp400gly glu401gly glu403gly mutation disrupting mutation disrupting interaction mutation disrupting certain certain sequence position certain certain certain sequence position certain false certain certain sequence position certain certain certain sequence position certain false certain certain sequence position certain certain certain sequence position certain false certain certain sequence position certain certain certain sequence position certain false certain certain sequence position certain certain certain sequence position certain false physical association physical association false false false Figure 3 The authors state that the proteins interacted within the endoplasmic reticulum. orp3a-sitosterol 802180 n/a 802183 unspecified role unspecified role neutral component neutral component region his tag his tag Hexa-His-tag 6-His-tag Histidine-tag Hexa-His-tag Histidine-tag 6-His-tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined false ecoli Escherichia coli (strain K12) n/a 802184 unspecified role unspecified role neutral component neutral component direct interaction direct interaction false false false Figure 7a orp3a-sitosterol-1 802181 n/a 802184 unspecified role unspecified role neutral component neutral component n/a 802183 unspecified role unspecified role neutral component neutral component region his tag his tag Hexa-His-tag 6-His-tag Histidine-tag Hexa-His-tag Histidine-tag 6-His-tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined false ecoli Escherichia coli (strain K12) physical association physical association false false false Figure 7b Figure 7c Sitosterol was labelled with tritium isotope.