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/ HNPCC mutations in hMSH2 result in reduced hMSH2-hMSH6 molecular switch functions. HNPCC mutations in hMSH2 result in reduced hMSH2-hMSH6 molecular switch functions. Cancer Cell (1535-6108) 2002 Heinen CD., Wilson T., Mazurek A., Berardini M., Butz C., Fishel R. Cancer - Interactions investigated in the context of cancer consulted Gradia et al 1997 to find out cell type recombinant proteins expressed in fishel@lac.jci.tju.edu 02-MAR-2006: Contacted by JYOTI. Returned mail. in vitro In vitro light scattering light scattering weight autoradiogra molecular weight estimation by autoradiography weight autoradiogra Heinen CD., Wilson T., Mazurek A., Berardini M., Butz C., Fishel R. Cancer - Interactions investigated in the context of cancer Total internal reflectance occurs when light strikes a medium boundary at a steep angle. If refractive index is lower on other side of the boundary no light can pass through, so all light is reflected (Wikipedia). The interaction between hMSH2-hMSH6 and the 41 bp DNA duplex with a G/T mismatch - attached via a biotin-streptavidin linkage to an IAsys cuvette - was measured using an IAsys Biosensor. The protein was incubated with mismatched DNA until saturation was reached. For protein dissociation studies, after saturation reached, unbound protein removed and replaced with protein-free buffer with or without ATP. consulted Gradia et al 1997 to find out cell type recombinant proteins expressed in fishel@lac.jci.tju.edu Accepted 2007-FEB-14 by DAVET. 02-MAR-2006: Contacted by JYOTI. Returned mail. Cancer Cell (1535-6108) 2002 HNPCC mutations in hMSH2 result in reduced hMSH2-hMSH6 molecular switch functions. HNPCC mutations in hMSH2 result in reduced hMSH2-hMSH6 molecular switch functions. Cancer Cell (1535-6108) 2002 Heinen CD., Wilson T., Mazurek A., Berardini M., Butz C., Fishel R. Cancer - Interactions investigated in the context of cancer consulted Gradia et al 1997 to find out cell type recombinant proteins expressed in fishel@lac.jci.tju.edu 02-MAR-2006: Contacted by JYOTI. Returned mail. in vitro In vitro emsa electrophoretic mobility shift assay Gel retardation assay band shift band shift Gel retardation assay emsa weight autoradiogra molecular weight estimation by autoradiography weight autoradiogra Heinen CD., Wilson T., Mazurek A., Berardini M., Butz C., Fishel R. Cancer - Interactions investigated in the context of cancer consulted Gradia et al 1997 to find out cell type recombinant proteins expressed in fishel@lac.jci.tju.edu Accepted 2007-FEB-14 by DAVET. 02-MAR-2006: Contacted by JYOTI. Returned mail. Cancer Cell (1535-6108) 2002 msh6_human DNA mismatch repair protein Msh6 MSH6 MutS-alpha 160 kDa subunit G/T mismatch-binding protein GTBP MutS protein homolog 6 protein protein human Homo sapiens Human MSRQSTLYSFFPKSPALSDANKASARASREGGRAAAAPGASPSPGGDAAWSEAGPGPRPLARSASPPKAKNLNGGLRRSVAPAAPTSCDFSPGDLVWAKMEGYPWWPCLVYNHPFDGTFIREKGKSVRVHVQFFDDSPTRGWVSKRLLKPYTGSKSKEAQKGGHFYSAKPEILRAMQRADEALNKDKIKRLELAVCDEPSEPEEEEEMEVGTTYVTDKSEEDNEIESEEEVQPKTQGSRRSSRQIKKRRVISDSESDIGGSDVEFKPDTKEEGSSDEISSGVGDSESEGLNSPVKVARKRKRMVTGNGSLKRKSSRKETPSATKQATSISSETKNTLRAFSAPQNSESQAHVSGGGDDSSRPTVWYHETLEWLKEEKRRDEHRRRPDHPDFDASTLYVPEDFLNSCTPGMRKWWQIKSQNFDLVICYKVGKFYELYHMDALIGVSELGLVFMKGNWAHSGFPEIAFGRYSDSLVQKGYKVARVEQTETPEMMEARCRKMAHISKYDRVVRREICRIITKGTQTYSVLEGDPSENYSKYLLSLKEKEEDSSGHTRAYGVCFVDTSLGKFFIGQFSDDRHCSRFRTLVAHYPPVQVLFEKGNLSKETKTILKSSLSCSLQEGLIPGSQFWDASKTLRTLLEEEYFREKLSDGIGVMLPQVLKGMTSESDSIGLTPGEKSELALSALGGCVFYLKKCLIDQELLSMANFEEYIPLDSDTVSTTRSGAIFTKAYQRMVLDAVTLNNLEIFLNGTNGSTEGTLLERVDTCHTPFGKRLLKQWLCAPLCNHYAINDRLDAIEDLMVVPDKISEVVELLKKLPDLERLLSKIHNVGSPLKSQNHPDSRAIMYEETTYSKKKIIDFLSALEGFKVMCKIIGIMEEVADGFKSKILKQVISLQTKNPEGRFPDLTVELNRWDTAFDHEKARKTGLITPKAGFDSDYDQALADIRENEQSLLEYLEKQRNRIGCRTIVYWGIGRNRYQLEIPENFTTRNLPEEYELKSTKKGCKRYWTKTIEKKLANLINAEERRDVSLKDCMRRLFYNFDKNYKDWQSAVECIAVLDVLLCLANYSRGGDGPMCRPVILLPEDTPPFLELKGSRHPCITKTFFGDDFIPNDILIGCEEEEQENGKAYCVLVTGPNMGGKSTLMRQAGLLAVMAQMGCYVPAEVCRLTPIDRVFTRLGASDRIMSGESTFFVELSETASILMHATAHSLVLVDELGRGTATFDGTAIANAVVKELAETIKCRTLFSTHYHSLVEDYSQNVAVRLGHMACMVENECEDPSQETITFLYKFIKGACPKSYGFNAARLANLPEEVIQKGHRKAREFEKMNQSLRLFREVCLASERSTVDAEAVHKLLTLIKEL DIP protein P52701 original sequence version: 128 4A4AA9F8ECB8FFE9 41_bp_heteroduplex 41 bp heteroduplex used with a G/T mismatch at position 21 ds dna double stranded deoxyribonucleic acid ds DNA ds dna chemical synthesis Chemical synthesis (Chemical synthesis) GCTTAGGATCATCGAGGATCGAGCTCGGTGCAATTCAGCGG 7D03729F45E598B0 msh2_human DNA mismatch repair protein Msh2 MSH2 MutS protein homolog 2 protein protein human Homo sapiens Human MAVQPKETLQLESAAEVGFVRFFQGMPEKPTTTVRLFDRGDFYTAHGEDALLAAREVFKTQGVIKYMGPAGAKNLQSVVLSKMNFESFVKDLLLVRQYRVEVYKNRAGNKASKENDWYLAYKASPGNLSQFEDILFGNNDMSASIGVVGVKMSAVDGQRQVGVGYVDSIQRKLGLCEFPDNDQFSNLEALLIQIGPKECVLPGGETAGDMGKLRQIIQRGGILITERKKADFSTKDIYQDLNRLLKGKKGEQMNSAVLPEMENQVAVSSLSAVIKFLELLSDDSNFGQFELTTFDFSQYMKLDIAAVRALNLFQGSVEDTTGSQSLAALLNKCKTPQGQRLVNQWIKQPLMDKNRIEERLNLVEAFVEDAELRQTLQEDLLRRFPDLNRLAKKFQRQAANLQDCYRLYQGINQLPNVIQALEKHEGKHQKLLLAVFVTPLTDLRSDFSKFQEMIETTLDMDQVENHEFLVKPSFDPNLSELREIMNDLEKKMQSTLISAARDLGLDPGKQIKLDSSAQFGYYFRVTCKEEKVLRNNKNFSTVDIQKNGVKFTNSKLTSLNEEYTKNKTEYEEAQDAIVKEIVNISSGYVEPMQTLNDVLAQLDAVVSFAHVSNGAPVPYVRPAILEKGQGRIILKASRHACVEVQDEIAFIPNDVYFEKDKQMFHIITGPNMGGKSTYIRQTGVIVLMAQIGCFVPCESAEVSIVDCILARVGAGDSQLKGVSTFMAEMLETASILRSATKDSLIIIDELGRGTSTYDGFGLAWAISEYIATKIGAFCMFATHFHELTALANQIPTVNNLHVTALTTEETLTMLYQVKKGVCDQSFGIHVAELANFPKHVIECAKQKALELEEFQYIGESQGYDIMEPAAKKCYLEREQGEKIIQEFLSKVKQMPFTEMSEENITIKLKQLKAEVIAKNNSFVNEIISRIKVTT DIP protein P43246 original sequence version: 156 DIP protein P43246 original sequence version: 123 DIP protein P43246 original sequence version: 138 664A058C78242E05 msh2-msh6-2 2 5 unspecified role unspecified role neutral component neutral component P43246:p.Pro622Leu mutation disrupting mutation disrupting interaction mutation disrupting mutation analysis mutation analysis certain certain sequence position certain certain certain sequence position certain P L Wed May 02 18:19:22 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'pro622leu' Tue Jun 30 15:45:01 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Pro622Leu' P43246:p.Arg524Pro mutation disrupting mutation disrupting interaction mutation disrupting mutation analysis mutation analysis certain certain sequence position certain certain certain sequence position certain R P Wed May 02 14:37:48 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'arg524pro' Tue Jun 30 19:32:13 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Arg524Pro' spofr-sf_9 Spodoptera frugiperda insect cells Fall armyworm sf_9 insect cells Derived from pupal ovarian tissue of the fall armyworm Cloned from the parent cell line, IPLB-SF-21-AE (SF-21) derived from immature ovaries of fall armyworm Spodoptera frugiperda pupae cells are susceptible to Baculovirus infection 3 unspecified role unspecified role neutral component neutral component n-terminal 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 spofr-sf_9 Spodoptera frugiperda insect cells Fall armyworm sf_9 insect cells Derived from pupal ovarian tissue of the fall armyworm Cloned from the parent cell line, IPLB-SF-21-AE (SF-21) derived from immature ovaries of fall armyworm Spodoptera frugiperda pupae cells are susceptible to Baculovirus infection 4 unspecified role unspecified role neutral component neutral component region 32p radiolabel 32p radiolabel P32 32P 32P P32 undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined g_t_mismatch G/T mismatch biological feature biological feature certain certain sequence position certain certain certain sequence position certain association association Compared to the wild-type protein, the hMSH2(P622L)- hMSH6 and hMSH2(R524P)-hMSH6 proteins failed to elicit a spethe cific gel shift of the labeled G/T mispair (Figure 2A). These results suggest that the hMSH2(P622L)-hMSH6 and hMSH2(R524P)- hMSH6 proteins confer a fundamental defect in mismatch rec- ognition. 2 A msh2-msh6-3 1 3 unspecified role unspecified role prey prey n-terminal 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 spofr-sf_9 Spodoptera frugiperda insect cells Fall armyworm sf_9 insect cells Derived from pupal ovarian tissue of the fall armyworm Cloned from the parent cell line, IPLB-SF-21-AE (SF-21) derived from immature ovaries of fall armyworm Spodoptera frugiperda pupae cells are susceptible to Baculovirus infection 5 unspecified role unspecified role prey prey P43246:p.Lys393Met mutation decreasing mutation decreasing interaction hotspot hotspot mutation decreasing mutation analysis mutation analysis certain certain sequence position certain certain certain sequence position certain K M Wed May 02 18:00:07 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'lys393met' Tue Jun 30 18:41:20 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Lys393Met' P43246:p.Arg524Pro mutation decreasing mutation decreasing interaction hotspot hotspot mutation decreasing mutation analysis mutation analysis certain certain sequence position certain certain certain sequence position certain R P Wed May 02 19:31:17 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'arg524pro' Tue Jun 30 18:14:26 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Arg524Pro' P43246:p.Pro622Leu mutation decreasing mutation decreasing interaction hotspot hotspot mutation decreasing mutation analysis mutation analysis certain certain sequence position certain certain certain sequence position certain P L Wed May 02 16:13:52 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'pro622leu' Tue Jun 30 17:05:46 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Pro622Leu' P43246:p.Gly674Ser mutation decreasing mutation decreasing interaction hotspot hotspot mutation decreasing mutation analysis mutation analysis certain certain sequence position certain certain certain sequence position certain G S Wed May 02 17:06:21 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'gly674ser' Tue Jun 30 17:10:27 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Gly674Ser' P43246:p.Thr905Arg mutation decreasing mutation decreasing interaction hotspot hotspot mutation decreasing mutation analysis mutation analysis certain certain sequence position certain certain certain sequence position certain T R Wed May 02 15:25:46 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'thr905arg' Tue Jun 30 18:58:27 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Thr905Arg' P43246:p.Asp167His mutation decreasing mutation decreasing interaction hotspot hotspot mutation decreasing mutation analysis mutation analysis certain certain sequence position certain certain certain sequence position certain D H Wed May 02 18:59:50 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'asp167his' Tue Jun 30 18:13:06 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Asp167His' spofr-sf_9 Spodoptera frugiperda insect cells Fall armyworm sf_9 insect cells Derived from pupal ovarian tissue of the fall armyworm Cloned from the parent cell line, IPLB-SF-21-AE (SF-21) derived from immature ovaries of fall armyworm Spodoptera frugiperda pupae cells are susceptible to Baculovirus infection 4 unspecified role unspecified role bait bait g_t_mismatch G/T mismatch biological feature biological feature certain certain sequence position certain certain certain sequence position certain region a single 3'-biotin biotin tag biotin tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined physical association physical association 2 B, Table 1, 4 B, C none of the hMSH2-hMSH6 missense mutant proteins retain the maximum binding (Bmax) of the wild-type protein. Moreover, between a 2- and 8-fold reduction in KD(G/T) was observed KDs for binding to the nucleic acid by heterodimers with the following mutations in hMSH2: D167H 6.05 E-8 M; K393M 4.38 E-8 M; R524P 7.21 E-8 M; N596Delta 8.8 E-8 M; P622L 9.71 E-8 M; G674S 3.61 E-8 M; T905R 2.57 E-8 M Addition of ATP increased the rate of heterodimer release approx. 165-fold. Addition of ATP increased the rate of heterodimer release. Koff 10-3 sec-1: Wild-type Koff 2.0 Koff(ATP) 329.0; D167H Koff 2.2 Koff(ATP) 389.0; K393M Koff 2.1 Koff(ATP) 186.0; R524P Koff 27.0 Koff(ATP) 42.3; N596Delta Koff 2.1 Koff(ATP) 314.0; P622L Koff 5.3 Koff(ATP) 5.0; G674S Koff 2.0 Koff(ATP) 305.0; T905R Koff 1.8 Koff(ATP) 311.0