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/ Vps51p links the VFT complex to the SNARE Tlg1p. Vps51p links the VFT complex to the SNARE Tlg1p. J. Biol. Chem. (0021-9258) 2002 imex curation Siniossoglou S., Pelham HR. hp@mrc-lmb.cam.ac.uk 09-April-2009: Contacted by JYOTI. orchard Only protein-protein interactions in vitro In vitro pull down pull down anti tag western anti tag western blot anti tag western Siniossoglou S., Pelham HR. hp@mrc-lmb.cam.ac.uk Accepted 2009-APR-13 by ORCHARD. 09-April-2009: Contacted by JYOTI. J. Biol. Chem. (0021-9258) 2002 orchard Only protein-protein interactions imex curation Vps51p links the VFT complex to the SNARE Tlg1p. Vps51p links the VFT complex to the SNARE Tlg1p. J. Biol. Chem. (0021-9258) 2002 imex curation Siniossoglou S., Pelham HR. hp@mrc-lmb.cam.ac.uk 09-April-2009: Contacted by JYOTI. orchard Only protein-protein interactions yeasx Saccharomyces cerevisiae (Baker's yeast) Baker's yeast pull down pull down western blot western blot Immuno blot Immuno blot Siniossoglou S., Pelham HR. hp@mrc-lmb.cam.ac.uk Accepted 2009-APR-13 by ORCHARD. 09-April-2009: Contacted by JYOTI. J. Biol. Chem. (0021-9258) 2002 orchard Only protein-protein interactions imex curation vps52_yeast Vacuolar protein sorting-associated protein 52 VPS52 YDR484W D8035.27 SAC2 Suppressor of actin mutation protein 2 protein protein yeast Saccharomyces cerevisiae Baker's yeast MDVLKEVLSLDQDKFDQLKETSRDKTNETDDPFENYLKDCKFKAPSNKDQSPFAKLKSLQETHSNNEAAINIIIPQLIDYLTEFTNRLSNYTQDLDFIKKKSNELQSLLEYNSTKLAHISPMVNDLMIPPELIDDIIKGKINESWQDNITFIADKEEIYNKYRSNNLDQDNKDAENSAMLAPKDFDKLCQLLDILKNVILERSKRLIISKIKTLRSHNPVPSQRIQNKLLKVQKIFPFIRDNNLSLALELRQAYCYTMKWYYREYFSRYIRSLTILQFQQIDSQFALGNGLSTTSVSGFNNSPSLFFSNYLTTSASNAFYNKLPVTDEKIDKYFQIKKRLNILTQEDNTVMVSQIAENNTTKNYIEIGFKNLNLAILDNCTVEYHFLKDFFAMNGDNFEEINGLLEQIFQPTFDEATTYTQQLIQYNYDIFGVLISIRVANQLQFESERRGIPSMFDSFLNGQLIQLWPRFQQLVDFQCESLRKAAITTNVAKYAGNSSTSNSSPLTSPHELTVQFGKFLSSFLTLAITHKQSIDERSEPLYNSIIRLRNDFETVMTKCSKKTKSPERFLATNYMYLYNNLQQLHLHLNINDSDAQNYNFDSAENVGTKVANDDDNDSSVPLIIRETENHFKTLVEAFTRN F2BF669A72A712C1 F2BF669A72A712C1 vps53_yeast Vacuolar protein sorting-associated protein 53 J1258 YJL029C VPS53 protein protein yeast Saccharomyces cerevisiae Baker's yeast MLEGTVDYDPLEDITNILFSKESLNNIDELISITRSYKKQLQEDILKEENELKEHPKNSAEIEASLRKVFQDFKETQDVSASTELTISNLTEGISYLDIAKKNLTHSLTLFQNLKILTDSYIQCNELLSQGSFKKMVSPYKIMCSLAENTFISYKSLDEINYLLSSISRLKGDTLSKIKQNYNALFSGGNISEHDTALTMELREGACELLDCDTSTRAQMIDWCLDKLLFEMKEIFRVDDEAGSLENLSRRYIYFKKILNNFNSKFADYFLKDWEMAVRLTTTFYHITHKDLQTLLKREFKDKNPSIDLFMTALQSTLDFEKYIDVRFSKKIKEPKLSSCFEPYLTLWVSHQNQMMEKKFLSYMSEPKYPSNETESLVLPSSADLFRTYRSVLTQTLELIDNNANDSILTSLANFFSRWLQTYSQKILLPLLLPDNIEVQDKLEAAKYTVLLINTADYCATTIDQLEDKLSEFSGNREKLANSFTKTKNIYDDLLAKGTSFLLNRVIPLDLNFVWREFINNDWSNAAIEDYSRYMVTLKSVLKMPALTDASIKQQQEQPSTLAFILSQFNRDVYKWNFLDKVIDIITTNFVSNTIRLLQPVPPFSLAGSKRKFETRTVVNIGEQLLLDLELLKEIFHTLPESVSNDSDLRENTSYKRVKRHADNNIDQLLKFIKLLMAPLDSADDYYETYSKLTNNNPDSAVWSFVLALKGIPWDLALWKKLWSAYNLETDDTDEGSRPDSNRDLFIFKWDKVLLGQFENNLARMQDPNWSKFVRQDLKISPPVMKRIVSTPQIQQQKEEQKKQSLSVKDFVSHSRFFNRGT C75F19FA57157481 vps51_yeast Vacuolar protein sorting-associated protein 51 YKR020W Apical bud growth protein 3 VPS51 API3 VPS67 WHI6 protein protein yeast Saccharomyces cerevisiae Baker's yeast MAEQISHKKSLRVSSLNKDRRLLLREFYNLENEPNKGRQEARIGEKASEAHSGEEQVTDVNIDTEANTEKPVKDDELSATEEDLKEGSEDAEEEIKNLPFKRLVQIHNKLLGKETETNNSIKNTIYENYYDLIKVNDLLKEITNANEDQINKLKQTVESLIKEL 7E76264F654AC452 ypt6_yeast GTP-binding protein YPT6 L8479.11 YLR262C YPT6 protein protein yeast Saccharomyces cerevisiae Baker's yeast MSRSGKSLTKYKIVFLGEQGVGKTSLITRFMYDTFDDHYQATIGIDFLSKTMYLDDKTIRLQLWDTAGQERFRSLIPSYIRDSRVAIIVYDITKRKSFEYIDKWIEDVKNERGDENVILCIVGNKSDLSDERQISTEEGEKKAKLLGAKIFMETSTKAGYNVKALFKKIAKSLPEFQNSESTPLDSENANSANQNKPGVIDISTAEEQEQSACQC A8E6211FF24FAE0B A8E6211FF24FAE0B vps54_yeast Vacuolar protein sorting-associated protein 54 VPS54 CGP1 CPF1 genetically-interacting protein 1 Temperature-sensitive clathrin synthetic mutation protein 3 LUV1 RKI1 TCS3 YDR027C PZF889 YD9813.05C protein protein yeast Saccharomyces cerevisiae Baker's yeast MSISETPHNKSQGLQKAAGRPKIVVPEGSPSRNSDSGSFTIEGDTSLNDDLLSISGSVTPRARRSSRLSLDSITPRRSFDSRTLSVANSRSFGFENETHSGSMDFSPLGNNSIYEIVMNTRRKNWLNYPTVADIPQVSLSKNDLDDHWKTHVIEYVKNIKSDYQIFQSTNNIRNMNQMEQLKELREGENMHEESFEANLRQGDAELINSIPDFYFSDKFQLDNPRTFHKVLDAIDLFLTKLDMKRQAERDEAFSELRDRLNDFLDIVETLLVTEISKSSHKFFHALSEVDNIQKRALDTMSELKELAQNIKTIDAENIRKKISHLEMIFKRKNVEKLEQGLLQAKLVLNKTDECKSMYEENKLDNCLELIKSIDYLIKGDDSINEDVQSWTRCWPYKLSNLRTIPALSATREFLTNMKIEIGGKFSLQLSILLIDDLRSFCKSIKPKETLHRIQTGSNDKKQTIFTDNFSSKITELIVRLNRCEELTSAFDLYREKSITELKSIIKIYLPTENAHADNNHDEKHLNNGSTSGSKLSRLIKEQTPAEFQSMLVNIFTHALEALRRLYGHQKLLLDISLNELASVKSPNENQHNMITQLDIRTGINEIIRIIQLRTGKIIAVRRELNLSLRYDYFLKFYAICVIFIQECEVLSGEFLTKYLSNVLASQIKHYANAQSSKNYRNIKKKIDAEEWIPYIVDSSIQSDVNDIVSSIDIDPLSWTTILDMVGGSHDCENGRSEDKEKDEGNETYQGHRKSVVVGDKTFVASSSLLATIEVIKELMVLSINLPSIYLSNFEKLCYDALQYYNSSAMASVTQPGNSLLKTGRNLSIMGESLDCLAEFVIIVQRFYQRLSNSNRDFEPFDASHYTTLLGQFQASSNKIYMANAPPPPV 67E930C34E8CC5AA tlg1_yeast t-SNARE affecting a late Golgi compartment protein 1 Syntaxin TLG1 TLG1 YDR468C D8035.11 protein protein yeast Saccharomyces cerevisiae Baker's yeast MNNSEDPFQQVVKDTKEQLNRINNYITRHNTAGDDDQEEEIQDILKDVEETIVDLDRSIIVMKRDENEDVSGREAQVKNIKQQLDALKLRFDRRIQESTQTTIPLEETVENSTLNTSMAENNDGGMSNPFQEQMLREQDVHLDGIHKTMQNLHIQAQTMGDELENQGQLLDNMDEGMDGVVNKLARGRRQLEWVYEKNKEKYDDCCIGLLIVVLIVLLVLAFIA B33251C18645EF74 B33251C18645EF74 tlg1-vps51 1 8 unspecified role unspecified role prey prey yeasx Saccharomyces cerevisiae (Baker's yeast) Baker's yeast 5 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 region required to bind necessary binding region required to bind deletion analysis deletion analysis certain certain sequence position certain certain certain sequence position certain ecoli Escherichia coli (strain K12) physical association physical association Figure 4 Vps51p binds to Tlg1p. 4B, the Tlg1p binding site mapped not to the C-terminal coiled-coil, but to the N terminus of Vps51p. More detailed mapping showed that residues 1�30, which are highly conserved, bound to Tlg1p as efficiently as did the full-length protein. vps54-vps51 2 5 unspecified role unspecified role bait bait region protein a tag protein a tag protein A protein A undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined region required to bind necessary binding region required to bind certain certain sequence position certain certain certain sequence position certain 7 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 association association 5c As shown in Fig. 5C, in each case removal of the coiled-coil domain from a subunit prevented its association with the VFT complex. vps54-vps52-2 2 3 unspecified role unspecified role bait bait region protein a tag protein a tag protein A protein A undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined region required to bind necessary binding region required to bind certain certain sequence position certain certain certain sequence position certain 7 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 association association 5c As shown in Fig. 5C, in each case removal of the coiled-coil domain from a subunit prevented its association with the VFT complex. vps54-vps53-1 2 7 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 4 unspecified role unspecified role bait bait region protein a tag protein a tag protein A protein A undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined region required to bind necessary binding region required to bind certain certain sequence position certain certain certain sequence position certain association association 5c As shown in Fig. 5C, in each case removal of the coiled-coil domain from a subunit prevented its association with the VFT complex. vps54-vps53-2 2 7 unspecified role unspecified role bait bait region protein a tag protein a tag protein A protein A undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined region required to bind necessary binding region required to bind certain certain sequence position certain certain certain sequence position certain 4 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 association association 5c As shown in Fig. 5C, in each case removal of the coiled-coil domain from a subunit prevented its association with the VFT complex. vps52-ypt6 1 3 unspecified role unspecified role prey prey region protein a tag protein a tag protein A protein A undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined yeasx Saccharomyces cerevisiae (Baker's yeast) Baker's yeast 6 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 ecoli Escherichia coli (strain K12) physical association physical association Figure 6 Vps52p had the ability to bind to Ypt6p and retained a preference for the GTP form.