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 Tim8-Tim13 complex has multiple substrate binding sites and binds cooperatively to Tim23. The Tim8-Tim13 complex has multiple substrate binding sites and binds cooperatively to Tim23. J. Mol. Biol. (0022-2836) 2008 imex curation Beverly K.N., Sawaya M.R., Schmid E., Koehler C.M. koehler@chem.ucla.edu 10-Mar-2014: Contacted by IntAct-Help. Only protein-protein interactions in vitro In vitro cd circular dichroism CD CD cd predetermined predetermined participant predetermined Beverly K.N., Sawaya M.R., Schmid E., Koehler C.M. koehler@chem.ucla.edu Accepted 2014-FEB-13 by PPORRAS J. Mol. Biol. (0022-2836) 2008 Only protein-protein interactions imex curation The Tim8-Tim13 complex has multiple substrate binding sites and binds cooperatively to Tim23. The Tim8-Tim13 complex has multiple substrate binding sites and binds cooperatively to Tim23. J. Mol. Biol. (0022-2836) 2008 imex curation Beverly K.N., Sawaya M.R., Schmid E., Koehler C.M. koehler@chem.ucla.edu 10-Mar-2014: Contacted by IntAct-Help. Only protein-protein interactions in vitro In vitro comigration in sds comigration in sds page comigration in sds weight by comassie molecular weight estimation by coomasie staining weight by comassie Beverly K.N., Sawaya M.R., Schmid E., Koehler C.M. koehler@chem.ucla.edu Accepted 2014-FEB-13 by PPORRAS J. Mol. Biol. (0022-2836) 2008 Only protein-protein interactions imex curation The Tim8-Tim13 complex has multiple substrate binding sites and binds cooperatively to Tim23. The Tim8-Tim13 complex has multiple substrate binding sites and binds cooperatively to Tim23. J. Mol. Biol. (0022-2836) 2008 imex curation Beverly K.N., Sawaya M.R., Schmid E., Koehler C.M. koehler@chem.ucla.edu 10-Mar-2014: Contacted by IntAct-Help. Only protein-protein interactions in vitro In vitro spr surface plasmon resonance Optical biosensor BIAcore(r) BIAcore(r) Optical biosensor spr predetermined predetermined participant predetermined Beverly K.N., Sawaya M.R., Schmid E., Koehler C.M. I've separated Fig. 3 and Table 3 to properly capture the Kd koehler@chem.ucla.edu Accepted 2014-FEB-13 by PPORRAS J. Mol. Biol. (0022-2836) 2008 Only protein-protein interactions imex curation The Tim8-Tim13 complex has multiple substrate binding sites and binds cooperatively to Tim23. The Tim8-Tim13 complex has multiple substrate binding sites and binds cooperatively to Tim23. J. Mol. Biol. (0022-2836) 2008 imex curation Beverly K.N., Sawaya M.R., Schmid E., Koehler C.M. koehler@chem.ucla.edu 10-Mar-2014: Contacted by IntAct-Help. Only protein-protein interactions in vitro In vitro bn-page blue native page bn-page predetermined predetermined participant predetermined Beverly K.N., Sawaya M.R., Schmid E., Koehler C.M. koehler@chem.ucla.edu Accepted 2014-FEB-13 by PPORRAS J. Mol. Biol. (0022-2836) 2008 Only protein-protein interactions imex curation The Tim8-Tim13 complex has multiple substrate binding sites and binds cooperatively to Tim23. The Tim8-Tim13 complex has multiple substrate binding sites and binds cooperatively to Tim23. J. Mol. Biol. (0022-2836) 2008 imex curation Beverly K.N., Sawaya M.R., Schmid E., Koehler C.M. koehler@chem.ucla.edu 10-Mar-2014: Contacted by IntAct-Help. Only protein-protein interactions in vitro In vitro x-ray diffraction x-ray crystallography X-ray X-ray x-ray diffraction predetermined predetermined participant predetermined Beverly K.N., Sawaya M.R., Schmid E., Koehler C.M. koehler@chem.ucla.edu Accepted 2014-FEB-13 by PPORRAS J. Mol. Biol. (0022-2836) 2008 Only protein-protein interactions imex curation Mock publication and experiment for abstract interactions that are not interaction evidences. Mock publication and experiment for abstract interactions that are not interaction evidences. unspecified method tim13_yeast Mitochondrial import inner membrane translocase subunit TIM13 G7157 YGR181W TIM13 protein protein yeast Saccharomyces cerevisiae Baker's yeast MGLSSIFGGGAPSQQKEAATTAKTTPNPIAKELKNQIAQELAVANATELVNKISENCFEKCLTSPYATRNDACIDQCLAKYMRSWNVISKAYISRIQNASASGEI B1DDFE87AFDC52A3 tim8_yeast Mitochondrial import inner membrane translocase subunit TIM8 TIM8 YJR135BW YJR135W-A protein protein yeast Saccharomyces cerevisiae Baker's yeast MSSLSTSDLASLDDTSKKEIATFLEGENSKQKVQMSIHQFTNICFKKCVESVNDSNLSSQEEQCLSNCVNRFLDTNIRIVNGLQNTR B91D1DD41707EA24 tim23_yeast Mitochondrial import inner membrane translocase subunit TIM23 MIM23 MPI3 N3180 YNR017W TIM23 MAS6 Mitochondrial protein import protein 3 Mitochondrial protein import protein MAS6 Membrane import machinery protein MIM23 protein protein yeast Saccharomyces cerevisiae Baker's yeast MSWLFGDKTPTDDANAAVGGQDTTKPKELSLKQSLGFEPNINNIISGPGGMHVDTARLHPLAGLDKGVEYLDLEEEQLSSLEGSQGLIPSRGWTDDLCYGTGAVYLLGLGIGGFSGMMQGLQNIPPNSPGKLQLNTVLNHITKRGPFLGNNAGILALSYNIINSTIDALRGKHDTAGSIGAGALTGALFKSSKGLKPMGYSSAMVAAACAVWCSVKKRLLEK 08E5AB4F400F675B 08E5AB4F400F675B tim8-tim13 5 7 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 ecoli Escherichia coli (strain K12) 8 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 ecoli Escherichia coli (strain K12) 14 12 direct interaction direct interaction Table 1 Rworking = 0.24 Rfree = 0.29 2.6A Sequence coordinates taken from the PDB submission file, however these do not correspond to the statement in the publication "Several N-terminal and a few C-terminal residues were invisible in the electron density map due to disorder. These include Tim8 residues 1–27 and 87 and Tim13 residues 1–45 and 98–104." so it is possible that the constructs were full length. tim8-tim13-1 1 7 unspecified role unspecified role neutral component neutral component ecoli Escherichia coli (strain K12) 8 unspecified role unspecified role neutral component neutral component c-terminal his tag his tag Hexa-His-tag 6-His-tag Histidine-tag Hexa-His-tag Histidine-tag 6-His-tag c-term range C-terminal range c-term range c-term range C-terminal range c-term range ecoli Escherichia coli (strain K12) direct interaction direct interaction Fig. 2C tim8-tim13-2 4 7 western blot western blot Immuno blot Immuno blot predetermined predetermined participant predetermined unspecified role unspecified role neutral component neutral component ecoli Escherichia coli (strain K12) 8 unspecified role unspecified role neutral component neutral component c-terminal his tag his tag Hexa-His-tag 6-His-tag Histidine-tag Hexa-His-tag Histidine-tag 6-His-tag c-term range C-terminal range c-term range c-term range C-terminal range c-term range ecoli Escherichia coli (strain K12) direct interaction direct interaction Fig. 2E tim13-tim23 3 7 unspecified role unspecified role bait bait c-terminal his tag his tag Hexa-His-tag 6-His-tag Histidine-tag Hexa-His-tag Histidine-tag 6-His-tag c-term range C-terminal range c-term range c-term range C-terminal range c-term range ecoli Escherichia coli (strain K12) 9 unspecified role unspecified role prey prey 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) 25 27 physical association physical association Fig. 3 tim8_tim13-tim23 3 9 unspecified role unspecified role prey prey 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) 32 unspecified role unspecified role bait bait 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 ecoli Escherichia coli (strain K12) physical association physical association Fig. 4, 5a, Supp. Fig. 2 tim13-tim8_tim13 3 32 unspecified role unspecified role bait bait ecoli Escherichia coli (strain K12) 9 unspecified role unspecified role prey prey 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 chemical synthesis Chemical synthesis (Chemical synthesis) physical association physical association Figs. 5 and 6 The addition of pep91– 103 with pep136–148 increased the binding of pep136–148 approximately three times above that of pep136–148 alone. This positive cooperativity implies that binding at one site on the complex may influence binding at other sites. tim8-tim13-1 2 7 unspecified role unspecified role neutral component neutral component ecoli Escherichia coli (strain K12) 8 unspecified role unspecified role neutral component neutral component c-terminal his tag his tag Hexa-His-tag 6-His-tag Histidine-tag Hexa-His-tag Histidine-tag 6-His-tag c-term range C-terminal range c-term range c-term range C-terminal range c-term range ecoli Escherichia coli (strain K12) direct interaction direct interaction Supp. Fig. 1A tim8-tim13_yeast TIM8-TIM13 mitochondrial intermembrane space protein transporter complex 3xTIM8:3xTIM13 7 unspecified role unspecified role tim8 binding domain sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain 8 unspecified role unspecified role tim13 binding domain sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain 44 46 direct interaction direct interaction yeast Saccharomyces cerevisiae Baker's yeast stable complex stable complex physical interaction evidence used in manual assertion physical interaction evidence used in manual assertion IPI inferred from physical interaction Facilitates transport of hydrophobic precursors of a distinct subgroup of inner membrane proteins through the aqueous intermembrane space as they exit the TOM40 channel complex (CPX-474) in the outer membrane. Plays a role in the import of a distinct subgroup of inner membrane proteins, functioning as a chaperone to maintain the hydrophobic membrane proteins in an import competent state and escort substrates to the TIM22 insertion complex (CPX-1629), which mediates protein insertion into the membrane. Cross-links to the COOH-terminal domain of the essential import translocase protein TIM23 (P32897) and plays a key role in TIM23 import to the mitochondrial inner membrane. Each subunit contains a central loop flanked by disulfide bonds that separate two extended N- and C-terminal helices. Buried salt-bridges between highly conserved lysine and glutamate residues connect alternating subunits. Mol. wt. = 70kDa. Heterohexamer created with intact-jami-dbscripts