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/ An adenosine deaminase that generates inosine at the wobble position of tRNAs. An adenosine deaminase that generates inosine at the wobble position of tRNAs. Science (0036-8075) 1999 imex curation Gerber AP., Keller W. walter.keller@unibas.ch 23-Feb-2010: Contacted by JYOTI. jyoti Only protein-protein interactions in vitro In vitro molecular sieving molecular sieving Gel Filtration Size Exclusion Chromatography Size Exclusion Chromatography Gel Filtration Sizing column predetermined predetermined participant predetermined Gerber AP., Keller W. example of participant determination by enzyme assay, to be updated when correct CV is available? Superdex 200 gel filtration column. Presence of participants demonstrated by running a tRNA-specific deamination assay on each fraction. walter.keller@unibas.ch Accepted 2010-FEB-12 by JYOTI. 23-Feb-2010: Contacted by JYOTI. Science (0036-8075) 1999 jyoti Only protein-protein interactions imex curation An adenosine deaminase that generates inosine at the wobble position of tRNAs. An adenosine deaminase that generates inosine at the wobble position of tRNAs. Science (0036-8075) 1999 imex curation Gerber AP., Keller W. walter.keller@unibas.ch 23-Feb-2010: Contacted by JYOTI. jyoti Only protein-protein interactions yeasx Saccharomyces cerevisiae (Baker's yeast) Baker's yeast pull down pull down ms participant Identification by mass spectrometry ms participant Gerber AP., Keller W. walter.keller@unibas.ch Accepted 2010-FEB-12 by JYOTI. 23-Feb-2010: Contacted by JYOTI. Science (0036-8075) 1999 jyoti Only protein-protein interactions imex curation tad2_yeast tRNA-specific adenosine deaminase subunit TAD2 TAD2 YJL035C J1246 tRNA-specific adenosine-34 deaminase subunit TAD2 protein protein yeast Saccharomyces cerevisiae Baker's yeast MQHIKHMRTAVRLARYALDHDETPVACIFVHTPTGQVMAYGMNDTNKSLTGVAHAEFMGIDQIKAMLGSRGVVDVFKDITLYVTVEPCIMCASALKQLDIGKVVFGCGNERFGGNGTVLSVNHDTCTLVPKNNSAAGYESIPGILRKEAIMLLRYFYVRQNERAPKPRSKSDRVLDKNTFPPMEWSKYLNEEAFIETFGDDYRTCFANKVDLSSNSVDWDLIDSHQDNIIQELEEQCKMFKFNVHKKSKV C774A1E50AA68106 tad3_yeast tRNA-specific adenosine deaminase subunit TAD3 tRNA-specific adenosine-34 deaminase subunit TAD3 YLR316C TAD3 protein protein yeast Saccharomyces cerevisiae Baker's yeast MVKKVNNPLKIDYQNGIIENRLLQIRNFKDVNTPKLINVWSIRIDPRDSKKVIELIRNDFQKNDPVSLRHLKRIRKDIETSTLEVVLCSKEYICDEGEINNKLKSIWVGTKKYELSDDIEVPEFAPSTKELNNAWSVKYWPLIWNGNPNDQILNDYKIDMQEVRNELSRASTLSVKMATAGKQFPMVSVFVDPSRKKDKVVAEDGRNCENSLPIDHSVMVGIRAVGERLREGVDEDANSYLCLDYDVYLTHEPCSMCSMALIHSRVRRVVFLTEMQRTGSLKLTSGDGYCMNDNKQLNSTYEAFQWIGEEYPVGQVDRDVCC 08D0878F10B36403 tad2-tad3-1 2 4 unspecified role unspecified role prey prey 3 unspecified role unspecified role bait bait n-terminal flag tag flag tag FLAG-tagged FLAG DYKDDDDKV epitope tag DYKDDDDKV epitope tag FLAG FLAG-tagged n-term range n-terminal range n-term range n-term range n-terminal range n-term range 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 physical association physical association To isolate this factor, we cultured tad2Delta [pGal-FLIS6-Tad2] cells and purified the Tad2 fusion protein (15). A protein of 38 kD copurified in a 1:1 ratio with the tagged Tad2p (Fig. 1, A and B), Fig. 1 tad2-tad3-2 1 3 unspecified role unspecified role neutral component neutral component n-terminal flag tag flag tag FLAG-tagged FLAG DYKDDDDKV epitope tag DYKDDDDKV epitope tag FLAG FLAG-tagged n-term range n-terminal range n-term range n-term range n-terminal range n-term range 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) 4 unspecified role unspecified role neutral component neutral component ecoli Escherichia coli (strain K12) direct interaction direct interaction To further analyze the Tad2p-Tad3p complex, we combined the recombinant proteins and ran them on a sizing column. rTad2p-rTad3p coeluted at an estimated molecular mass of 70 kD together with tRNA:A34 deaminase activity (2), indicating that Tad2p and Tad3p form a heterodimeric complex. Supp. fig. 2 Complex m.w.=70kDa