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/ Phosphorylation of SARS-CoV-2 Orf9b Regulates Its Targeting to Two Binding Sites in TOM70 and Recruitment of Hsp90. Phosphorylation of SARS-CoV-2 Orf9b Regulates Its Targeting to Two Binding Sites in TOM70 and Recruitment of Hsp90. International journal of molecular sciences 2021 imex curation Brandherm L., Kobaš AM., Klöhn M., Brüggemann Y., Pfaender S., Rassow J., Kreimendahl S. Virus - Publications including interactions involving viral proteins Coronavirus - Interactions investigated in the context of Coronavirus joachim.rassow@rub.de Only protein-protein interactions imex curation in vitro In vitro enzymatic footprint enzymatic footprinting enzymatic footprint weight autoradiogra molecular weight estimation by autoradiography weight autoradiogra Brandherm L., Kobaš AM., Klöhn M., Brüggemann Y., Pfaender S., Rassow J., Kreimendahl S. Virus - Publications including interactions involving viral proteins joachim.rassow@rub.de Accepted 2021-OCT-21 AT 09:05 BST AT 09:05 BST by ORCHARD International journal of molecular sciences 2021 Only protein-protein interactions imex curation imex curation Phosphorylation of SARS-CoV-2 Orf9b Regulates Its Targeting to Two Binding Sites in TOM70 and Recruitment of Hsp90. Phosphorylation of SARS-CoV-2 Orf9b Regulates Its Targeting to Two Binding Sites in TOM70 and Recruitment of Hsp90. International journal of molecular sciences 2021 imex curation Brandherm L., Kobaš AM., Klöhn M., Brüggemann Y., Pfaender S., Rassow J., Kreimendahl S. Virus - Publications including interactions involving viral proteins Coronavirus - Interactions investigated in the context of Coronavirus joachim.rassow@rub.de Only protein-protein interactions imex curation rabit-reticulocyte Oryctolagus cuniculus reticulocyte reticulocyte Reticulocytes bn-page blue native page bn-page weight autoradiogra molecular weight estimation by autoradiography weight autoradiogra Brandherm L., Kobaš AM., Klöhn M., Brüggemann Y., Pfaender S., Rassow J., Kreimendahl S. Virus - Publications including interactions involving viral proteins joachim.rassow@rub.de Accepted 2021-OCT-21 AT 09:05 BST AT 09:05 BST by ORCHARD International journal of molecular sciences 2021 Only protein-protein interactions imex curation imex curation Phosphorylation of SARS-CoV-2 Orf9b Regulates Its Targeting to Two Binding Sites in TOM70 and Recruitment of Hsp90. Phosphorylation of SARS-CoV-2 Orf9b Regulates Its Targeting to Two Binding Sites in TOM70 and Recruitment of Hsp90. International journal of molecular sciences 2021 imex curation Brandherm L., Kobaš AM., Klöhn M., Brüggemann Y., Pfaender S., Rassow J., Kreimendahl S. Virus - Publications including interactions involving viral proteins Coronavirus - Interactions investigated in the context of Coronavirus joachim.rassow@rub.de Only protein-protein interactions imex curation ecoli Escherichia coli (strain K12) molecular sieving molecular sieving Gel Filtration Size Exclusion Chromatography Size Exclusion Chromatography Gel Filtration Sizing column predetermined predetermined participant predetermined Brandherm L., Kobaš AM., Klöhn M., Brüggemann Y., Pfaender S., Rassow J., Kreimendahl S. Virus - Publications including interactions involving viral proteins joachim.rassow@rub.de Accepted 2021-OCT-21 AT 09:05 BST AT 09:05 BST by ORCHARD International journal of molecular sciences 2021 Only protein-protein interactions imex curation imex curation Phosphorylation of SARS-CoV-2 Orf9b Regulates Its Targeting to Two Binding Sites in TOM70 and Recruitment of Hsp90. Phosphorylation of SARS-CoV-2 Orf9b Regulates Its Targeting to Two Binding Sites in TOM70 and Recruitment of Hsp90. International journal of molecular sciences 2021 imex curation Brandherm L., Kobaš AM., Klöhn M., Brüggemann Y., Pfaender S., Rassow J., Kreimendahl S. Virus - Publications including interactions involving viral proteins Coronavirus - Interactions investigated in the context of Coronavirus joachim.rassow@rub.de Only protein-protein interactions imex curation in vitro In vitro pull down pull down weight autoradiogra molecular weight estimation by autoradiography weight autoradiogra Brandherm L., Kobaš AM., Klöhn M., Brüggemann Y., Pfaender S., Rassow J., Kreimendahl S. Virus - Publications including interactions involving viral proteins joachim.rassow@rub.de Accepted 2021-OCT-21 AT 09:05 BST AT 09:05 BST by ORCHARD International journal of molecular sciences 2021 Only protein-protein interactions imex curation imex curation hs90b_human Heat shock protein HSP 90-beta HSP90AB1 HSP90B HSPC2 HSPCB Heat shock 84 kDa protein protein human Homo sapiens Human MPEEVHHGEEEVETFAFQAEIAQLMSLIINTFYSNKEIFLRELISNASDALDKIRYESLTDPSKLDSGKELKIDIIPNPQERTLTLVDTGIGMTKADLINNLGTIAKSGTKAFMEALQAGADISMIGQFGVGFYSAYLVAEKVVVITKHNDDEQYAWESSAGGSFTVRADHGEPIGRGTKVILHLKEDQTEYLEERRVKEVVKKHSQFIGYPITLYLEKEREKEISDDEAEEEKGEKEEEDKDDEEKPKIEDVGSDEEDDSGKDKKKKTKKIKEKYIDQEELNKTKPIWTRNPDDITQEEYGEFYKSLTNDWEDHLAVKHFSVEGQLEFRALLFIPRRAPFDLFENKKKKNNIKLYVRRVFIMDSCDELIPEYLNFIRGVVDSEDLPLNISREMLQQSKILKVIRKNIVKKCLELFSELAEDKENYKKFYEAFSKNLKLGIHEDSTNRRRLSELLRYHTSQSGDEMTSLSEYVSRMKETQKSIYYITGESKEQVANSAFVERVRKRGFEVVYMTEPIDEYCVQQLKEFDGKSLVSVTKEGLELPEDEEEKKKMEESKAKFENLCKLMKEILDKKVEKVTISNRLVSSPCCIVTSTYGWTANMERIMKAQALRDNSTMGYMMAKKHLEINPDHPIVETLRQKAEADKNDKAVKDLVVLLFETALLSSGFSLEDPQTHSNRIYRMIKLGLGIDEDEVAAEEPNAAVPDEIPPLEGDEDASRMEEVD DIP protein P08238 original sequence version: 91 DIP protein P08238 original sequence version: 149 DIP protein P08238 original sequence version: 165 DIP protein P08238 original sequence version: 167 DIP protein P08238 original sequence version: 210 DIP protein P08238 original sequence version: 187 A93118C214D03810 orf9b_sars2 ORF9b protein Accessory protein 9b ORF-9b Protein 9b 9b protein protein SARS-CoV-2 SARS-CoV-2 SARS-CoV-2 Severe acute respiratory syndrome coronavirus 2 2019-nCoV MDPKISEMHPALRLVDPQIQLAVTRMENAVGRDQNNVGPKVYPIILRLGSPLSLNMARKTLNSLEDKAFQLTPIAVQMTKLATTEELPDEFVVVTVK tom70_human Mitochondrial import receptor subunit TOM70 TOMM70 KIAA0719 TOM70 TOMM70A Translocase of outer membrane 70 kDa subunit Translocase of outer mitochondrial membrane protein 70 Mitochondrial precursor proteins import receptor protein protein human Homo sapiens Human MAASKPVEAAVVAAAVPSSGSGVGGGGTAGPGTGGLPRWQLALAVGAPLLLGAGAIYLWSRQQRRREARGRGDASGLKRNSERKTPEGRASPAPGSGHPEGPGAHLDMNSLDRAQAAKNKGNKYFKAGKYEQAIQCYTEAISLCPTEKNVDLSTFYQNRAAAFEQLQKWKEVAQDCTKAVELNPKYVKALFRRAKAHEKLDNKKECLEDVTAVCILEGFQNQQSMLLADKVLKLLGKEKAKEKYKNREPLMPSPQFIKSYFSSFTDDIISQPMLKGEKSDEDKDKEGEALEVKENSGYLKAKQYMEEENYDKIISECSKEIDAEGKYMAEALLLRATFYLLIGNANAAKPDLDKVISLKEANVKLRANALIKRGSMYMQQQQPLLSTQDFNMAADIDPQNADVYHHRGQLKILLDQVEEAVADFDECIRLRPESALAQAQKCFALYRQAYTGNNSSQIQAAMKGFEEVIKKFPRCAEGYALYAQALTDQQQFGKADEMYDKCIDLEPDNATTYVHKGLLQLQWKQDLDRGLELISKAIEIDNKCDFAYETMGTIEVQRGNMEKAIDMFNKAINLAKSEMEMAHLYSLCDAAHAQTEVAKKYGLKPPTL 5AAF5CAAA8582480 adt3_human ADP/ATP translocase 3 SLC25A6 ANT3 CDABP0051 Adenine nucleotide translocator 3 ADP,ATP carrier protein 3 Solute carrier family 25 member 6 ADP,ATP carrier protein, isoform T2 AAC3 protein protein human Homo sapiens Human MTEQAISFAKDFLAGGIAAAISKTAVAPIERVKLLLQVQHASKQIAADKQYKGIVDCIVRIPKEQGVLSFWRGNLANVIRYFPTQALNFAFKDKYKQIFLGGVDKHTQFWRYFAGNLASGGAAGATSLCFVYPLDFARTRLAADVGKSGTEREFRGLGDCLVKITKSDGIRGLYQGFSVSVQGIIIYRAAYFGVYDTAKGMLPDPKNTHIVVSWMIAQTVTAVAGVVSYPFDTVRRRMMMQSGRKGADIMYTGTVDCWRKIFRDEGGKAFFKGAWSNVLRGMGGAFVLVLYDELKKVI 18534E9F0E49672F tomm70-orf9b_sars2-1 4 6 unspecified role unspecified role prey prey in vitro translated in vitro translated protein in vitro translated P0DTD2:p.Ser53Glu mutation decreasing strength mutation decreasing interaction strength mutation decreasing strength certain certain sequence position certain certain certain sequence position certain S E Tue Oct 05 19:19:56 BST 2021 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Ser53Glu' P0DTD2:p.Ser53Ala mutation with no effect mutation with no effect mutation with no effect mutation not affecting interaction certain certain sequence position certain certain certain sequence position certain S A Tue Oct 05 18:51:25 BST 2021 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Ser53Ala' region 35s radiolabel 35s radiolabel 35S s35 radiolabelled S35 s35 radiolabelled 35S S35 undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined rabit-reticulocyte Oryctolagus cuniculus reticulocyte reticulocyte Reticulocytes 7 unspecified role unspecified role bait bait purified purified O94826:p.Glu477Ala mutation decreasing strength mutation decreasing interaction strength mutation decreasing strength certain certain sequence position certain certain certain sequence position certain E A Tue Oct 05 18:43:12 BST 2021 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Glu477Ala' n-terminus his tag his tag Hexa-His-tag 6-His-tag Histidine-tag Hexa-His-tag Histidine-tag 6-His-tag n-term range n-terminal range n-term range n-term range n-terminal range n-term range cytosolic domain sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain ecoli Escherichia coli (strain K12) physical association physical association 1C/I/K A mutation of either amino acid residue (TOM70-E477A OR Of9b-S53E) drastically reduces the association of the two proteins. The incubation of phosphomimetic Orf9bS53E with isolated TOM70E477A did not lead to a reduction in comparison to TOM70WT. orf9b_sars2-24 3 6 unspecified role unspecified role neutral component neutral component purified purified P0DTD2:p.Ser53Glu mutation with no effect mutation with no effect mutation with no effect mutation not affecting interaction certain certain sequence position certain certain certain sequence position certain S E Tue Oct 05 18:51:30 BST 2021 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Ser53Glu' direct interaction direct interaction 1F Both Orf9b variants were eluted as dimers of a molecular mass of approximately 28.7 kDa. tomm70-hsp90ab1-1 4 5 unspecified role unspecified role prey prey in vitro translated in vitro translated protein in vitro translated region 35s radiolabel 35s radiolabel 35S s35 radiolabelled S35 s35 radiolabelled 35S S35 undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined rabit-reticulocyte Oryctolagus cuniculus reticulocyte reticulocyte Reticulocytes 7 unspecified role unspecified role bait bait purified purified O94826:p.Glu477Ala mutation decreasing strength mutation decreasing interaction strength mutation decreasing strength certain certain sequence position certain certain certain sequence position certain E A Tue Oct 05 19:23:47 BST 2021 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Glu477Ala' n-terminus his tag his tag Hexa-His-tag 6-His-tag Histidine-tag Hexa-His-tag Histidine-tag 6-His-tag n-term range n-terminal range n-term range n-term range n-terminal range n-term range cytosolic domain sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain O94826:p.Arg192Ala mutation decreasing strength mutation decreasing interaction strength mutation decreasing strength certain certain sequence position certain certain certain sequence position certain R A Tue Oct 05 18:45:16 BST 2021 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Arg192Ala' ecoli Escherichia coli (strain K12) physical association physical association Figs. 3 B/C, 4C Orf9b (P0DTD2), Wild type only. No effect was observed when TOM70 was pre-incubated with the phosphomimetic variant Orf9bS53E. tomm70-slc25a6-1 4 8 unspecified role unspecified role prey prey in vitro translated in vitro translated protein in vitro translated region 35s radiolabel 35s radiolabel 35S s35 radiolabelled S35 s35 radiolabelled 35S S35 undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined rabit-reticulocyte Oryctolagus cuniculus reticulocyte reticulocyte Reticulocytes 7 unspecified role unspecified role bait bait purified purified O94826:p.Glu477Ala mutation decreasing strength mutation decreasing interaction strength mutation decreasing strength certain certain sequence position certain certain certain sequence position certain E A Tue Oct 05 19:15:00 BST 2021 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Glu477Ala' n-terminus his tag his tag Hexa-His-tag 6-His-tag Histidine-tag Hexa-His-tag Histidine-tag 6-His-tag n-term range n-terminal range n-term range n-term range n-terminal range n-term range cytosolic domain sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain ecoli Escherichia coli (strain K12) physical association physical association Fig. 2D Orf9b (P0DTD2). tomm70-orf9b_sars2-2 4 6 unspecified role unspecified role prey prey in vitro translated in vitro translated protein in vitro translated region sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain region sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain region 35s radiolabel 35s radiolabel 35S s35 radiolabelled S35 s35 radiolabelled 35S S35 undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined rabit-reticulocyte Oryctolagus cuniculus reticulocyte reticulocyte Reticulocytes 7 unspecified role unspecified role bait bait purified purified O94826:p.Arg192Ala mutation decreasing strength mutation decreasing interaction strength mutation decreasing strength certain certain sequence position certain certain certain sequence position certain R A Tue Oct 05 19:30:55 BST 2021 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Arg192Ala' n-terminus his tag his tag Hexa-His-tag 6-His-tag Histidine-tag Hexa-His-tag Histidine-tag 6-His-tag n-term range n-terminal range n-term range n-term range n-terminal range n-term range cytosolic domain sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain ecoli Escherichia coli (strain K12) physical association physical association 4E/F/G entered as a separate interaction as only Wt of the viral protein is tested. 400 mM KCl, but only for WT interaction. tomm70-orf9b_sars2-3 1 6 unspecified role unspecified role neutral component neutral component in vitro translated in vitro translated protein in vitro translated Dihydrofolate reductase tag tag n-term range n-terminal range n-term range n-term range n-terminal range n-term range P0DTD2:p.Ser53Glu mutation decreasing strength mutation decreasing interaction strength mutation decreasing strength certain certain sequence position certain certain certain sequence position certain S E Tue Oct 05 18:47:47 BST 2021 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Ser53Glu' P0DTD2:p.Ser53Ala mutation with no effect mutation with no effect mutation with no effect mutation not affecting interaction certain certain sequence position certain certain certain sequence position certain S A Tue Oct 05 19:08:58 BST 2021 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Ser53Ala' region 35s radiolabel 35s radiolabel 35S s35 radiolabelled S35 s35 radiolabelled 35S S35 undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined rabit-reticulocyte Oryctolagus cuniculus reticulocyte reticulocyte Reticulocytes 7 unspecified role unspecified role neutral component neutral component purified purified n-terminus his tag his tag Hexa-His-tag 6-His-tag Histidine-tag Hexa-His-tag Histidine-tag 6-His-tag n-term range n-terminal range n-term range n-term range n-terminal range n-term range cytosolic domain sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain ecoli Escherichia coli (strain K12) physical association physical association Supp. figure 2 Orf9bWT-DHFR is efficiently bound to the C-terminal domain of TOM70, with this part of the construct being shielded from trypsin degradation. The Orf9b S53A mutant showed the same fragmentation pattern as the WT, but Orf9b S53E showed much less protection in the presence of TOM70, indicating reduced binding of both proteins. orf9b_sars2-orf9b_sars2-1 2 6 unspecified role unspecified role neutral component neutral component in vitro translated in vitro translated protein in vitro translated region 35s radiolabel 35s radiolabel 35S s35 radiolabelled S35 s35 radiolabelled 35S S35 undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined 6 unspecified role unspecified role neutral component neutral component in vitro translated in vitro translated protein in vitro translated region 35s radiolabel 35s radiolabel 35S s35 radiolabelled S35 s35 radiolabelled 35S S35 undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined physical association physical association Supp. figure 3 not "direct" interaction as several other things will be present. Most of the Orf9b was found as a component of a high molecular weight complex of 220–240 kDa.