DIP Database of Interacting Proteins 611 Young Drive East; Los Angeles CA 90095; USA http://dip.doe-mbi.ucla.edu/ dip@mbi.ucla.edu Identification of an unconventional E3 binding surface on the UbcH5 ~ Ub conjugate recognized by a pathogenic bacterial E3 ligase. Identification of an unconventional E3 binding surface on the UbcH5 ~ Ub conjugate recognized by a pathogenic bacterial E3 ligase. Proceedings of the National Academy of Sciences of the United States of America 2010 imex curation Levin I., Eakin C., Blanc MP., Klevit RE., Miller SI., Brzovic PS., Levin I., Eakin C., Blanc MP., Klevit RE., Miller SI., Brzovic PS., Levin I., Eakin C., Blanc MP., Klevit RE., Miller SI., Brzovic PS., Levin I., Eakin C., Blanc MP., Klevit RE., Miller SI., Brzovic PS. klevit@u.washington.edu 22-Feb-2016: Contacted by IntAct-Help. Only protein-protein interactions in vitro In vitro ubiquitinase assay ubiquitinase assay ubiquitinase assay western blot western blot Immuno blot Immuno blot Levin I., Eakin C., Blanc MP., Klevit RE., Miller SI., Brzovic PS., Levin I., Eakin C., Blanc MP., Klevit RE., Miller SI., Brzovic PS., Levin I., Eakin C., Blanc MP., Klevit RE., Miller SI., Brzovic PS., Levin I., Eakin C., Blanc MP., Klevit RE., Miller SI., Brzovic PS. In Fig S2 a His-tag is stated to be present on Ssph2 OR E1 SspH2-catalyzed reactions were performed with either purified UbcH5 ∼ Ub conjugates or in reaction mixtures containing E1 and ubiquitin (origins unknown). klevit@u.washington.edu Accepted 2016-JAN-20 AT 09:29 GMT AT 09:29 GMT by MGT Proceedings of the National Academy of Sciences of the United States of America 2010 imex curation Identification of an unconventional E3 binding surface on the UbcH5 ~ Ub conjugate recognized by a pathogenic bacterial E3 ligase. Identification of an unconventional E3 binding surface on the UbcH5 ~ Ub conjugate recognized by a pathogenic bacterial E3 ligase. Proceedings of the National Academy of Sciences of the United States of America 2010 imex curation Levin I., Eakin C., Blanc MP., Klevit RE., Miller SI., Brzovic PS., Levin I., Eakin C., Blanc MP., Klevit RE., Miller SI., Brzovic PS., Levin I., Eakin C., Blanc MP., Klevit RE., Miller SI., Brzovic PS., Levin I., Eakin C., Blanc MP., Klevit RE., Miller SI., Brzovic PS. klevit@u.washington.edu 22-Feb-2016: Contacted by IntAct-Help. Only protein-protein interactions in vitro In vitro ubiquitinase assay ubiquitinase assay ubiquitinase assay ms participant Identification by mass spectrometry ms participant Levin I., Eakin C., Blanc MP., Klevit RE., Miller SI., Brzovic PS., Levin I., Eakin C., Blanc MP., Klevit RE., Miller SI., Brzovic PS., Levin I., Eakin C., Blanc MP., Klevit RE., Miller SI., Brzovic PS., Levin I., Eakin C., Blanc MP., Klevit RE., Miller SI., Brzovic PS. SspH2-catalyzed reactions were performed with either purified UbcH5 ∼ Ub conjugates or in reaction mixtures containing E1 and ubiquitin (origins unknown). klevit@u.washington.edu Accepted 2016-JAN-20 AT 09:29 GMT AT 09:29 GMT by MGT Proceedings of the National Academy of Sciences of the United States of America 2010 imex curation Identification of an unconventional E3 binding surface on the UbcH5 ~ Ub conjugate recognized by a pathogenic bacterial E3 ligase. Identification of an unconventional E3 binding surface on the UbcH5 ~ Ub conjugate recognized by a pathogenic bacterial E3 ligase. Proceedings of the National Academy of Sciences of the United States of America 2010 imex curation Levin I., Eakin C., Blanc MP., Klevit RE., Miller SI., Brzovic PS., Levin I., Eakin C., Blanc MP., Klevit RE., Miller SI., Brzovic PS., Levin I., Eakin C., Blanc MP., Klevit RE., Miller SI., Brzovic PS., Levin I., Eakin C., Blanc MP., Klevit RE., Miller SI., Brzovic PS. klevit@u.washington.edu 22-Feb-2016: Contacted by IntAct-Help. Only protein-protein interactions in vitro In vitro ubiquitinase assay ubiquitinase assay ubiquitinase assay weight by comassie molecular weight estimation by coomasie staining weight by comassie Levin I., Eakin C., Blanc MP., Klevit RE., Miller SI., Brzovic PS., Levin I., Eakin C., Blanc MP., Klevit RE., Miller SI., Brzovic PS., Levin I., Eakin C., Blanc MP., Klevit RE., Miller SI., Brzovic PS., Levin I., Eakin C., Blanc MP., Klevit RE., Miller SI., Brzovic PS. SspH2-catalyzed reactions were performed with either purified UbcH5 ∼ Ub conjugates or in reaction mixtures containing E1 and ubiquitin (origins unknown). klevit@u.washington.edu Accepted 2016-JAN-20 AT 09:29 GMT AT 09:29 GMT by MGT Proceedings of the National Academy of Sciences of the United States of America 2010 imex curation Identification of an unconventional E3 binding surface on the UbcH5 ~ Ub conjugate recognized by a pathogenic bacterial E3 ligase. Identification of an unconventional E3 binding surface on the UbcH5 ~ Ub conjugate recognized by a pathogenic bacterial E3 ligase. Proceedings of the National Academy of Sciences of the United States of America 2010 imex curation Levin I., Eakin C., Blanc MP., Klevit RE., Miller SI., Brzovic PS., Levin I., Eakin C., Blanc MP., Klevit RE., Miller SI., Brzovic PS., Levin I., Eakin C., Blanc MP., Klevit RE., Miller SI., Brzovic PS., Levin I., Eakin C., Blanc MP., Klevit RE., Miller SI., Brzovic PS. klevit@u.washington.edu 22-Feb-2016: Contacted by IntAct-Help. Only protein-protein interactions in vitro In vitro nmr nuclear magnetic resonance NMR NMR nmr predetermined predetermined participant predetermined Levin I., Eakin C., Blanc MP., Klevit RE., Miller SI., Brzovic PS., Levin I., Eakin C., Blanc MP., Klevit RE., Miller SI., Brzovic PS., Levin I., Eakin C., Blanc MP., Klevit RE., Miller SI., Brzovic PS., Levin I., Eakin C., Blanc MP., Klevit RE., Miller SI., Brzovic PS. klevit@u.washington.edu Accepted 2016-JAN-20 AT 09:29 GMT AT 09:29 GMT by MGT Proceedings of the National Academy of Sciences of the United States of America 2010 imex curation ub2d3_human Ubiquitin-conjugating enzyme E2 D3 UBC5C Ubiquitin-protein ligase D3 Ubiquitin carrier protein D3 Ubiquitin-conjugating enzyme E2-17 kDa 3 Ubiquitin-conjugating enzyme E2(17)KB 3 UBE2D3 UBCH5C (E3-independent) E2 ubiquitin-conjugating enzyme D3 E2 ubiquitin-conjugating enzyme D3 protein protein human Homo sapiens Human MALKRINKELSDLARDPPAQCSAGPVGDDMFHWQATIMGPNDSPYQGGVFFLTIHFPTDYPFKPPKVAFTTRIYHPNINSNGSICLDILRSQWSPALTISKVLLSICSLLCDPNPDDPLVPEIARIYKTDRDKYNRISREWTQKYAM ADD74A8A708EFEE3 ssph2_salty E3 ubiquitin-protein ligase SspH2 Salmonella secreted protein H2 Secreted effector protein sspH2 STM2241 sspH2 RING-type E3 ubiquitin transferase SspH2 protein protein salty Salmonella typhimurium MPFHIGSGCLPATISNRRIYRIAWSDTPPEMSSWEKMKEFFCSTHQTEALECIWTICHPPAGTTREDVINRFELLRTLAYAGWEESIHSGQHGENYFCILDEDSQEILSVTLDDAGNYTVNCQGYSETHRLTLDTAQGEEGTGHAEGASGTFRTSFLPATTAPQTPAEYDAVWSAWRRAAPAEESRGRAAVVQKMRACLNNGNAVLNVGESGLTTLPDCLPAHITTLVIPDNNLTSLPALPPELRTLEVSGNQLTSLPVLPPGLLELSIFSNPLTHLPALPSGLCKLWIFGNQLTSLPVLPPGLQELSVSDNQLASLPALPSELCKLWAYNNQLTSLPMLPSGLQELSVSDNQLASLPTLPSELYKLWAYNNRLTSLPALPSGLKELIVSGNRLTSLPVLPSELKELMVSGNRLTSLPMLPSGLLSLSVYRNQLTRLPESLIHLSSETTVNLEGNPLSERTLQALREITSAPGYSGPIIRFDMAGASAPRETRALHLAAADWLVPAREGEPAPADRWHMFGQEDNADAFSLFLDRLSETENFIKDAGFKAQISSWLAQLAEDEALRANTFAMATEATSSCEDRVTFFLHQMKNVQLVHNAEKGQYDNDLAALVATGREMFRLGKLEQIAREKVRTLALVDEIEVWLAYQNKLKKSLGLTSVTSEMRFFDVSGVTVTDLQDAELQVKAAEKSEFREWILQWGPLHRVLERKAPERVNALREKQISDYEETYRMLSDTELRPSGLVGNTDAERTIGARAMESAKKTFLDGLRPLVEEMLGSYLNVQWRRN E12C103027294080 ssph1_salt1 E3 ubiquitin-protein ligase SspH1 sspH1 Salmonella secreted protein H1 Secreted effector protein SspH1 STM14_1483 RING-type E3 ubiquitin transferase SspH1 protein protein salt1 Salmonella typhimurium (strain 14028s / SGSC 2262) MFNIRNTQPSVSMQAIAGAAAPEASPEEIVWEKIQVFFPQENYEEAQQCLAELCHPARGMLPDHISSQFARLKALTFPAWEENIQCNRDGINQFCILDAGSKEILSITLDDAGNYTVNCQGYSEAHDFIMDTEPGEECTEFAEGASGTSLRPATTVSQKAAEYDAVWSKWERDAPAGESPGRAAVVQEMRDCLNNGNPVLNVGASGLTTLPDRLPPHITTLVIPDNNLTSLPELPEGLRELEVSGNLQLTSLPSLPQGLQKLWAYNNWLASLPTLPPGLGDLAVSNNQLTSLPEMPPALRELRVSGNNLTSLPALPSGLQKLWAYNNRLTSLPEMSPGLQELDVSHNQLTRLPQSLTGLSSAARVYLDGNPLSVRTLQALRDIIGHSGIRIHFDMAGPSVPREARALHLAVADWLTSAREGEAAQADRWQAFGLEDNAAAFSLVLDRLRETENFKKDAGFKAQISSWLTQLAEDAALRAKTFAMATEATSTCEDRVTHALHQMNNVQLVHNAEKGEYDNNLQGLVSTGREMFRLATLEQIAREKAGTLALVDDVEVYLAFQNKLKESLELTSVTSEMRFFDVSGVTVSDLQAAELQVKTAENSGFSKWILQWGPLHSVLERKVPERFNALREKQISDYEDTYRKLYDEVLKSSGLVDDTDAERTIGVSAMDSAKKEFLDGLRALVDEVLGSYLTARWRLN 141612145C01D582 ssph2-ubch5-1 4 6 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 5 unspecified role unspecified role neutral component neutral component cys85 oglycoser O-glycosyl-L-serine OGlycoSer certain certain sequence position certain certain certain sequence position certain P61077:p.Cys85Ser mutation mutation certain certain sequence position certain certain certain sequence position certain C S Wed May 02 18:14:01 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'cys85ser' Tue Jun 30 17:38:07 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Cys85Ser' P61077:p.Ala96Asp mutation decreasing mutation decreasing interaction hotspot hotspot mutation decreasing certain certain sequence position certain certain certain sequence position certain A D Wed May 02 16:56:10 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'ala96asp' Tue Jun 30 16:16:22 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Ala96Asp' 15n label 15n label 15n label 15N N15 15N N15 undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined ecoli Escherichia coli (strain K12) physical association physical association 2 and 3A SspH2 preferentially binds the substrate (i.e., UbcH5 ∼ Ub) as opposed to the product (i.e., free UbcH5) of the Ub-transfer reaction. The UbcH5 molecule has an active site C85S mutation, and through this site is linked to Ubiquitin via an ester link, rather than through a thio-ester link. This UbcH5-O-Ub conjugate is stable. ssph2-ube2d3-2 1 6 unspecified role unspecified role neutral component neutral component 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 region sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain 5 unspecified role unspecified role neutral component neutral component ecoli Escherichia coli (strain K12) ubiquitination ubiquitination reaction ubiquitination S1 and S2 In Fig S2 a His-tag is stated to be present on Ssph2 OR E1 ssph2-ube2d3-4 1 6 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 5 unspecified role unspecified role neutral component neutral component P61077:p.Cys85Ser mutation mutation certain certain sequence position certain certain certain sequence position certain C S Wed May 02 17:29:31 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'cys85ser' Tue Jun 30 19:22:49 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Cys85Ser' cys85 oglycoser O-glycosyl-L-serine OGlycoSer certain certain sequence position certain certain certain sequence position certain ecoli Escherichia coli (strain K12) ubiquitination ubiquitination reaction ubiquitination 5 The UbcH5 molecule has an active site C85S mutation, and through this site is linked to Ubiquitin via an ester link, rather than through a thio-ester link. This UbcH5-O-Ub conjugate is stable. ssph2-ube2d3-3 1 6 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 5 unspecified role unspecified role neutral component neutral component P61077:p.Ala96Asp mutation decreasing rate mutation decreasing interaction rate mutation decreasing rate certain certain sequence position certain certain certain sequence position certain A D Wed May 02 15:08:15 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'ala96asp' Tue Jun 30 18:23:16 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Ala96Asp' P61077:p.Phe62Ala mutation decreasing rate mutation decreasing interaction rate mutation decreasing rate certain certain sequence position certain certain certain sequence position certain F A Wed Jul 06 17:07:42 BST 2016 Sequence change details about this feature cannot be ascertained or do not fit with the current version of the referenced protein, so they have been deleted as a result of our quality control procedures. The original label was 'phe62ala' Wed May 02 17:25:54 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'phe62ala' Tue Jun 30 17:37:07 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Phe62Ala' ecoli Escherichia coli (strain K12) ubiquitination ubiquitination reaction ubiquitination 1B, 4, S6 and 3B ssph2-ube2d3-5 3 6 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 5 unspecified role unspecified role neutral component neutral component ecoli Escherichia coli (strain K12) ubiquitination ubiquitination reaction ubiquitination S3A The use of K48R-Ub generates high MW ubiquitylation products ssph2-ube2d3-6 2 6 unspecified role unspecified role neutral component neutral component 5 unspecified role unspecified role neutral component neutral component ecoli Escherichia coli (strain K12) ubiquitination ubiquitination reaction ubiquitination S3B ssph1-ube2d3-1 1 7 unspecified role unspecified role neutral component neutral component 5 unspecified role unspecified role neutral component neutral component ecoli Escherichia coli (strain K12) ubiquitination ubiquitination reaction ubiquitination S2 ssph2-ube2d3-7 1 6 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 P0CE12:p.Cys580Ala mutation decreasing rate mutation decreasing interaction rate mutation decreasing rate certain certain sequence position certain certain certain sequence position certain C A Wed May 02 15:16:53 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'cys580ala' Tue Jun 30 17:09:12 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Cys580Ala' 5 unspecified role unspecified role neutral component neutral component ecoli Escherichia coli (strain K12) ubiquitination ubiquitination reaction ubiquitination S2,6