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/ Structure of protein phosphatase 2A core enzyme bound to tumor-inducing toxins. Structure of protein phosphatase 2A core enzyme bound to tumor-inducing toxins. Cell (0092-8674) 2006 imex curation Xing Y., Xu Y., Chen Y., Jeffrey PD., Chao Y., Lin Z., Li Z., Strack S., Stock JB., Shi Y. Cancer - Interactions investigated in the context of cancer ygshi@princeton.edu 13-Nov-2007: Contacted by JYOTI. Only protein-protein interactions 2 in vitro In vitro molecular sieving molecular sieving Gel Filtration Size Exclusion Chromatography Size Exclusion Chromatography Gel Filtration Sizing column predetermined predetermined participant predetermined Xing Y., Xu Y., Chen Y., Jeffrey PD., Chao Y., Lin Z., Li Z., Strack S., Stock JB., Shi Y. Cancer - Interactions investigated in the context of cancer The PP2A core enzyme was methylated by LCMT in the presence of S-adenosyl methionine. The fully methylated or unmethylated PP2A core enzyme was incubated with an excess amount of the B"/PR72 protein and the mixtures were examined by gel filtration chromatography. ygshi@princeton.edu Accepted 2007-DEC-20 by DAVET. 13-Nov-2007: Contacted by JYOTI. Cell (0092-8674) 2006 Only protein-protein interactions 2 imex curation Structure of protein phosphatase 2A core enzyme bound to tumor-inducing toxins. Structure of protein phosphatase 2A core enzyme bound to tumor-inducing toxins. Cell (0092-8674) 2006 imex curation Xing Y., Xu Y., Chen Y., Jeffrey PD., Chao Y., Lin Z., Li Z., Strack S., Stock JB., Shi Y. Cancer - Interactions investigated in the context of cancer ygshi@princeton.edu 13-Nov-2007: Contacted by JYOTI. Only protein-protein interactions 2 in vitro In vitro x-ray diffraction x-ray crystallography X-ray X-ray x-ray diffraction predetermined predetermined participant predetermined Xing Y., Xu Y., Chen Y., Jeffrey PD., Chao Y., Lin Z., Li Z., Strack S., Stock JB., Shi Y. Cancer - Interactions investigated in the context of cancer ygshi@princeton.edu Accepted 2007-DEC-20 by DAVET. Cell (0092-8674) 2006 Only protein-protein interactions 2 imex curation okadaic acid Okadaic acid small molecule small molecule chemical synthesis Chemical synthesis (Chemical synthesis) microcystin-peptide Microcystin-LR (MCLR) heptapeptide peptide peptide polypeptide oligopeptide oligopeptide polypeptide chemical synthesis Chemical synthesis (Chemical synthesis) DRXAAAL An amino acid sequence which is not represented in UniProt This peptide is a tumour inducer. It is an inhibitor of the PP2A phosphatase. 672DDDDDCAF40AC0 2aaa_human Serine/threonine-protein phosphatase 2A 65 kDa regulatory subunit A alpha isoform PPP2R1A PP2A subunit A isoform PR65-alpha PP2A subunit A isoform R1-alpha Medium tumor antigen-associated 61 kDa protein protein protein human Homo sapiens Human MAAADGDDSLYPIAVLIDELRNEDVQLRLNSIKKLSTIALALGVERTRSELLPFLTDTIYDEDEVLLALAEQLGTFTTLVGGPEYVHCLLPPLESLATVEETVVRDKAVESLRAISHEHSPSDLEAHFVPLVKRLAGGDWFTSRTSACGLFSVCYPRVSSAVKAELRQYFRNLCSDDTPMVRRAAASKLGEFAKVLELDNVKSEIIPMFSNLASDEQDSVRLLAVEACVNIAQLLPQEDLEALVMPTLRQAAEDKSWRVRYMVADKFTELQKAVGPEITKTDLVPAFQNLMKDCEAEVRAAASHKVKEFCENLSADCRENVIMSQILPCIKELVSDANQHVKSALASVIMGLSPILGKDNTIEHLLPLFLAQLKDECPEVRLNIISNLDCVNEVIGIRQLSQSLLPAIVELAEDAKWRVRLAIIEYMPLLAGQLGVEFFDEKLNSLCMAWLVDHVYAIREAATSNLKKLVEKFGKEWAHATIIPKVLAMSGDPNYLHRMTTLFCINVLSEVCGQDITTKHMLPTVLRMAGDPVANVRFNVAKSLQKIGPILDNSTLQSEVKPILEKLTQDQDVDVKYFAQEALTVLSLA DIP protein P30153 original sequence version: 153 DIP protein P30153 original sequence version: 71 DIP protein P30153 original sequence version: 105 DIP protein P30153 original sequence version: 107 DIP protein P30153 original sequence version: 151 DIP protein P30153 original sequence version: 156 DIP protein P30153 original sequence version: 155 DIP protein P30153 original sequence version: 169 5174EBE94D537836 p2r3a_human Serine/threonine-protein phosphatase 2A regulatory subunit B'' subunit alpha PPP2R3A PPP2R3 Serine/threonine-protein phosphatase 2A 72/130 kDa regulatory subunit B PP2A subunit B isoforms B''-PR72/PR130 PP2A subunit B isoforms B72/B130 PP2A subunit B isoform PR72/PR130 PP2A subunit B isoform R3 isoform protein protein human Homo sapiens Human MAATYRLVVSTVNHYSSVVIDRRFEQAIHYCTGTCHTFTHGIDCIVVHHSVCADLLHIPVSQFKDADLNSMFLPHENGLSSAEGDYPQQAFTGIPRVKRGSTFQNTYNLKDIAGEAISFASGKIKEFSFEKLKNSNHAAYRKGRKVKSDSFNRRSVDLDLLCGHYNNDGNAPSFGLLRSSSVEEKPLSHRNSLDTNLTSMFLQNFSEEDLVTQILEKHKIDNFSSGTDIKMCLDILLKCSEDLKKCTDIIKQCIKKKSGSSISEGSGNDTISSSETVYMNVMTRLASYLKKLPFEFMQSGNNEALDLTELISNMPSLQLTPFSPVFGTEQPPKYEDVVQLSASDSGRFQTIELQNDKPNSRKMDTVQSIPNNSTNSLYNLEVNDPRTLKAVQVQSQSLTMNPLENVSSDDLMETLYIEEESDGKKALDKGQKTENGPSHELLKVNEHRAEFPEHATHLKKCPTPMQNEIGKIFEKSFVNLPKEDCKSKVSKFEEGDQRDFTNSSSQEEIDKLLMDLESFSQKMETSLREPLAKGKNSNFLNSHSQLTGQTLVDLEPKSKVSSPIEKVSPSCLTRIIETNGHKIEEEDRALLLRILESIEDFAQELVECKSSRGSLSQEKEMMQILQETLTTSSQANLSVCRSPVGDKAKDTTSAVLIQQTPEVIKIQNKPEKKPGTPLPPPATSPSSPRPLSPVPHVNNVVNAPLSINIPRFYFPEGLPDTCSNHEQTLSRIETAFMDIEEQKADIYEMGKIAKVCGCPLYWKAPMFRAAGGEKTGFVTAQSFIAMWRKLLNNHHDDASKFICLLAKPNCSSLEQEDFIPLLQDVVDTHPGLTFLKDAPEFHSRYITTVIQRIFYTVNRSWSGKITSTEIRKSNFLQTLALLEEEEDINQITDYFSYEHFYVIYCKFWELDTDHDLYISQADLSRYNDQASSSRIIERIFSGAVTRGKTIQKEGRMSYADFVWFLISEEDKRNPTSIEYWFRCMDVDGDGVLSMYELEYFYEEQCERMEAMGIEPLPFHDLLCQMLDLVKPAVDGKITLRDLKRCRMAHIFYDTFFNLEKYLDHEQRDPFAVQKDVENDGPEPSDWDRFAAEEYETLVAEESAQAQFQEGFEDYETDEPASPSEFGNKSNKILSASLPEKCGKLQSVDEE DIP protein Q06190 original sequence version: 138 DIP protein Q06190 original sequence version: 63 97A31BA4206518A3 pp2aa_human Serine/threonine-protein phosphatase 2A catalytic subunit alpha isoform PPP2CA Replication protein C protein protein human Homo sapiens Human MDEKVFTKELDQWIEQLNECKQLSESQVKSLCEKAKEILTKESNVQEVRCPVTVCGDVHGQFHDLMELFRIGGKSPDTNYLFMGDYVDRGYYSVETVTLLVALKVRYRERITILRGNHESRQITQVYGFYDECLRKYGNANVWKYFTDLFDYLPLTALVDGQIFCLHGGLSPSIDTLDHIRALDRLQEVPHEGPMCDLLWSDPDDRGGWGISPRGAGYTFGQDISETFNHANGLTLVSRAHQLVMEGYNWCHDRNVVTIFSAPNYCYRCGNQAAIMELDDTLKYSFLQFDPAPRRGEPHVTRRTPDYFL DIP protein P67775 original sequence version: 112 DIP protein P67775-1 original sequence version: 98 DIP protein P67775 original sequence version: 29 DIP protein P67775 original sequence version: 102 DIP protein P67775 original sequence version: 65 DIP protein P67775 original sequence version: 67 DIP protein P67775-1 original sequence version: 114 DIP protein P67775 original sequence version: 113 DIP protein P67775-1 original sequence version: 122 C602291F78F34555 ppp2ca-ppp2r1a-1 2 3 unspecified role unspecified role neutral component neutral component 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 ile71 binding region binding-associated region binding region certain certain sequence position certain certain certain sequence position certain arg70 binding region binding-associated region binding region certain certain sequence position certain certain certain sequence position certain glu67 binding region binding-associated region binding region certain certain sequence position certain certain certain sequence position certain lys74 binding region binding-associated region binding region certain certain sequence position certain certain certain sequence position certain asp280 binding region binding-associated region binding region certain certain sequence position certain certain certain sequence position certain asp280 binding region binding-associated region binding region certain certain sequence position certain certain certain sequence position certain arg70 binding region binding-associated region binding region certain certain sequence position certain certain certain sequence position certain asp280 binding region binding-associated region binding region certain certain sequence position certain certain certain sequence position certain asn79 binding region binding-associated region binding region certain certain sequence position certain certain certain sequence position certain pro76 binding region binding-associated region binding region certain certain sequence position certain certain certain sequence position certain asp77 binding region binding-associated region binding region certain certain sequence position certain certain certain sequence position certain pro51 binding region binding-associated region binding region certain certain sequence position certain certain certain sequence position certain trini-high_5 Trichoplusia ni cell line from eggs high_5 Insect cell line from Trichoplusia ni egg cells Invitrogen supplies this cell line. Invitrogen supply this cell line. 5 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 leu454 binding region binding-associated region binding region certain certain sequence position certain certain certain sequence position certain arg417 binding region binding-associated region binding region certain certain sequence position certain certain certain sequence position certain arg417 binding region binding-associated region binding region certain certain sequence position certain certain certain sequence position certain tyr455 binding region binding-associated region binding region certain certain sequence position certain certain certain sequence position certain pro492 binding region binding-associated region binding region certain certain sequence position certain certain certain sequence position certain trp416 binding region binding-associated region binding region certain certain sequence position certain certain certain sequence position certain tyr494 binding region binding-associated region binding region certain certain sequence position certain certain certain sequence position certain arg497 binding region binding-associated region binding region certain certain sequence position certain certain certain sequence position certain arg497 binding region binding-associated region binding region certain certain sequence position certain certain certain sequence position certain asp530 binding region binding-associated region binding region certain certain sequence position certain certain certain sequence position certain asp497 binding region binding-associated region binding region certain certain sequence position certain certain certain sequence position certain asp491 binding region binding-associated region binding region certain certain sequence position certain certain certain sequence position certain arg497 binding region binding-associated region binding region certain certain sequence position certain certain certain sequence position certain pro492 binding region binding-associated region binding region certain certain sequence position certain certain certain sequence position certain asn534 binding region binding-associated region binding region certain certain sequence position certain certain certain sequence position certain asn534 binding region binding-associated region binding region certain certain sequence position certain certain certain sequence position certain asn534 binding region binding-associated region binding region certain certain sequence position certain certain certain sequence position certain val532 binding region binding-associated region binding region certain certain sequence position certain certain certain sequence position certain ecobb Escherichia coli (strain B / BL21) 12 26 13 27 14 28 29 15 32 16 33 17 31 18 19 30 40 20 21 41 42 22 23 43 34 35 37 36 38 39 direct interaction direct interaction Table 1, Figure 3, Figure 4 The PP2A core enzyme was separately incubated with 1.2 molar equivalence of OA or MCLR for crystallization. After extensive trials, we succeeded in obtaining crystals of the 100 kDa PP2A core enzyme separately bound to OA and MCLR under similar conditions. The structures were determined by molecular replacement and refined to 2.6 and 2.8 A resolution, respectively, for the OA- and MCLR-bound PP2A core enzyme 2.5 A Rwork 0.226; Rfree 0.270 The regulatory subunit sequence used in the experiment had an initial Met whereas the UniprotKB sequence does not. The regulatory subunit sequence used has an initial Met and an Arg at position 259, whereas the sequence for P30153 has no initial Met and an Ala at position 258. ppp2ca-ppp2r1a-2 2 4 unspecified role unspecified role neutral component neutral component chemical synthesis Chemical synthesis (Chemical synthesis) 7 unspecified role unspecified role neutral component neutral component arg70 binding region binding-associated region binding region certain certain sequence position certain certain certain sequence position certain asp280 binding region binding-associated region binding region certain certain sequence position certain certain certain sequence position certain asp280 binding region binding-associated region binding region certain certain sequence position certain certain certain sequence position certain arg70 binding region binding-associated region binding region certain certain sequence position certain certain certain sequence position certain ile71 binding region binding-associated region binding region certain certain sequence position certain certain certain sequence position certain glu67 binding region binding-associated region binding region certain certain sequence position certain certain certain sequence position certain lys74 binding region binding-associated region binding region certain certain sequence position certain certain certain sequence position certain asp280 binding region binding-associated region binding region certain certain sequence position certain certain certain sequence position certain asn79 binding region binding-associated region binding region certain certain sequence position certain certain certain sequence position certain pro76 binding region binding-associated region binding region certain certain sequence position certain certain certain sequence position certain asp77 binding region binding-associated region binding region certain certain sequence position certain certain certain sequence position certain pro51 binding region binding-associated region binding region 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 trini-high_5 Trichoplusia ni cell line from eggs high_5 Insect cell line from Trichoplusia ni egg cells Invitrogen supplies this cell line. Invitrogen supply this cell line. 5 unspecified role unspecified role neutral component neutral component arg417 binding region binding-associated region binding region certain certain sequence position certain certain certain sequence position certain arg497 binding region binding-associated region binding region certain certain sequence position certain certain certain sequence position certain arg497 binding region binding-associated region binding region certain certain sequence position certain certain certain sequence position certain pro492 binding region binding-associated region binding region certain certain sequence position certain certain certain sequence position certain asn534 binding region binding-associated region binding region certain certain sequence position certain certain certain sequence position certain asn534 binding region binding-associated region binding region 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 trp416 binding region binding-associated region binding region certain certain sequence position certain certain certain sequence position certain leu454 binding region binding-associated region binding region certain certain sequence position certain certain certain sequence position certain arg417 binding region binding-associated region binding region certain certain sequence position certain certain certain sequence position certain tyr455 binding region binding-associated region binding region certain certain sequence position certain certain certain sequence position certain pro492 binding region binding-associated region binding region certain certain sequence position certain certain certain sequence position certain tyr494 binding region binding-associated region binding region certain certain sequence position certain certain certain sequence position certain arg497 binding region binding-associated region binding region certain certain sequence position certain certain certain sequence position certain asp530 binding region binding-associated region binding region certain certain sequence position certain certain certain sequence position certain arg497 binding region binding-associated region binding region certain certain sequence position certain certain certain sequence position certain asp491 binding region binding-associated region binding region certain certain sequence position certain certain certain sequence position certain asn534 binding region binding-associated region binding region certain certain sequence position certain certain certain sequence position certain val532 binding region binding-associated region binding region certain certain sequence position certain certain certain sequence position certain ecobb Escherichia coli (strain B / BL21) 47 61 48 73 49 74 50 68 69 51 52 70 53 71 72 54 78 55 65 56 66 57 58 79 77 62 64 63 76 75 direct interaction direct interaction Table 1, 3, 4 The PP2A core enzyme was separately incubated with 1.2 molar equivalence of OA or MCLR for crystallization. After extensive trials, we succeeded in obtaining crystals of the 100 kDa PP2A core enzyme separately bound to OA and MCLR under similar conditions. The structures were determined by molecular replacement and refined to 2.6 and 2.8 A resolution, respectively, for the OA- and MCLR-bound PP2A core enzyme The regulatory subunit sequence used has an initial Met and an Arg at position 259, whereas the sequence for P30153 has no initial Met and an Ala at position 258. The catalytic subunit sequence used has an Val at position 5 whereas the sequence for P67775 has a Leu at position 5. Microcystin-LR is an inhibitor of PP2A and is an tumour-inducing agent. 2.8 A Rwork 0.222; Rfree 0.264 ppp2ca-ppp2r1a 1 6 unspecified role unspecified role neutral component neutral component ecobb Escherichia coli (strain B / BL21) 7 unspecified role unspecified role neutral component neutral component leu309 meres methylated residue MeRes Methylation (N terminus, N epsilon of Lysine, O of Serine, Threonine or C terminus, N of Asparagine) certain certain sequence position certain certain certain sequence position certain trini Trichoplusia ni (Cabbage looper) Cabbage looper 5 unspecified role unspecified role neutral component neutral component ecobb Escherichia coli (strain B / BL21) physical association physical association 6 The methylated PP2A core enzyme coeluted with the B00/PR72 protein in a single peak, which corresponds to an apparent molecular mass of approximately 160 kDa (Figure 6A, top right panel). Importantly, the elution volume for this peak is smaller than that of the PP2A core enzyme or the B00/PR72 protein alone. This result unambiguously demonstrates that the methylated PP2A core enzyme forms a stable holoenzyme with the B00/PR72 protein. In contrast, the unmethylated PP2A core enzyme failed to form a stable complex with the B00/PR72 protein, as the PP2A core enzyme eluted from gel filtration at a volume identical to that of the free PP2A core enzyme The interaction of the core PP2A enzyme (P67775 and P30153) with PR72 (Q06190) requires the methylated of Leu-309 of the catalytic subunit of the core enzyme.