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/ Critical glycosylated residues in exon three of erythrocyte glycophorin A engage Plasmodium falciparum EBA-175 and define receptor specificity. Critical glycosylated residues in exon three of erythrocyte glycophorin A engage Plasmodium falciparum EBA-175 and define receptor specificity. mBio 2014 imex curation Salinas ND., Paing MM., Tolia NH. GPA (ecd):- Sequence, as shown in Figure 1A appears to be that of GenBank: AAA52624.1. This sequence has been remapped to P02724 due to 'Erroneous initiation' of AAA52624.1. EBA-175 P. falciparum protein:- Ref 7 states this is a protein of 616 amino acid and PDBe entries give UniProt ID =Q25735 for the protein. No strain of the parasite is stated, and Q25735 has been remapped to reviewed entry P19214 (aas 145-760) with which it has 99% homology. tolia@wustl.edu 15-Nov-2017: Contacted by IntAct-Help. Only protein-protein interactions imex curation in vitro In vitro solid phase assay solid phase assay tag fluorescence tag visualisation by fluorescence tag fluorescence Salinas ND., Paing MM., Tolia NH. terminal sialic acid was removed by neuraminidase treatment. GPA (ecd):- Sequence, as shown in Figure 1A appears to be that of GenBank: AAA52624.1. This sequence has been remapped to P02724 due to 'Erroneous initiation' of AAA52624.1. EBA-175 P. falciparum protein:- Ref 7 states this is a protein of 616 amino acid and PDBe entries give UniProt ID =Q25735 for the protein. No strain of the parasite is stated, and Q25735 has been remapped to reviewed entry P19214 (aas 145-760) with which it has 99% homology. tolia@wustl.edu Accepted 2017-OCT-04 AT 14:32 BST AT 14:32 BST by ORCHARD mBio 2014 imex curation Email sent 29/817 to you ,Sylvie+ Pablo - regarding carbohydrate PTMs, as suggested by Pablo. EBI-15486608 the Peanut agglutinin has a different specificity than Soyabean Lectin (probably binding to beta-galactose or beta-galactose linked to N-Acetylgalactosamine) whereas Soyabean Lectin just binds to alpha and beta N-acetyl-galactosamine, according to this publication; the publication also states "...are known to bind different sugar moieties on glycosylated proteins" - so I did not think I should add the same carbohydrate residue for both agglutinins, even if both agglutinins DO react with N-acetyl-galactosamine? I have altered the "glycores" to a "Pre-requisite PTM" for BOTH interactions and added a comment regarding specificity, and removed CHEBI:28800 from EBI-15486561. Hope this is OK. ; I have shown the dimerisation by linking the two Fc tags (- cannot link using "cys2" as a feature) Critical glycosylated residues in exon three of erythrocyte glycophorin A engage Plasmodium falciparum EBA-175 and define receptor specificity. Critical glycosylated residues in exon three of erythrocyte glycophorin A engage Plasmodium falciparum EBA-175 and define receptor specificity. mBio 2014 imex curation Salinas ND., Paing MM., Tolia NH. GPA (ecd):- Sequence, as shown in Figure 1A appears to be that of GenBank: AAA52624.1. This sequence has been remapped to P02724 due to 'Erroneous initiation' of AAA52624.1. EBA-175 P. falciparum protein:- Ref 7 states this is a protein of 616 amino acid and PDBe entries give UniProt ID =Q25735 for the protein. No strain of the parasite is stated, and Q25735 has been remapped to reviewed entry P19214 (aas 145-760) with which it has 99% homology. tolia@wustl.edu 15-Nov-2017: Contacted by IntAct-Help. Only protein-protein interactions imex curation in vitro In vitro pull down pull down western blot western blot Immuno blot Immuno blot Salinas ND., Paing MM., Tolia NH. P19214: this is the only reviewed entry for EBA-175 for P. falciparum GPA (ecd):- Sequence, as shown in Figure 1A appears to be that of GenBank: AAA52624.1. This sequence has been remapped to P02724 due to 'Erroneous initiation' of AAA52624.1. EBA-175 P. falciparum protein:- Ref 7 states this is a protein of 616 amino acid and PDBe entries give UniProt ID =Q25735 for the protein. No strain of the parasite is stated, and Q25735 has been remapped to reviewed entry P19214 (aas 145-760) with which it has 99% homology. tolia@wustl.edu Accepted 2017-OCT-04 AT 14:32 BST AT 14:32 BST by ORCHARD mBio 2014 imex curation Email sent 29/817 to you ,Sylvie+ Pablo - regarding carbohydrate PTMs, as suggested by Pablo. EBI-15486608 the Peanut agglutinin has a different specificity than Soyabean Lectin (probably binding to beta-galactose or beta-galactose linked to N-Acetylgalactosamine) whereas Soyabean Lectin just binds to alpha and beta N-acetyl-galactosamine, according to this publication; the publication also states "...are known to bind different sugar moieties on glycosylated proteins" - so I did not think I should add the same carbohydrate residue for both agglutinins, even if both agglutinins DO react with N-acetyl-galactosamine? I have altered the "glycores" to a "Pre-requisite PTM" for BOTH interactions and added a comment regarding specificity, and removed CHEBI:28800 from EBI-15486561. Hope this is OK. ; I have shown the dimerisation by linking the two Fc tags (- cannot link using "cys2" as a feature) lecg_arahy Galactose-binding lectin Agglutinin PNA protein protein arahy Arachis hypogaea (Peanut) MKPFCVFLTFFLLLAASSKKVDSAETVSFNFNSFSEGNPAINFQGDVTVLSNGNIQLTNLNKVNSVGRVLYAMPVRIWSSATGNVASFLTSFSFEMKDIKDYDPADGIIFFIAPEDTQIPAGSIGGGTLGVSDTKGAGHFVGVEFDTYSNSEYNDPPTDHVGIDVNSVDSVKTVPWNSVSGAVVKVTVIYDSSTKTLSVAVTNDNGDITTIAQVVDLKAKLPERVKFGFSASGSLGGRQIHLIRSWSFTSTLITTTRRSIDNNEKKIMNMASA 05A0B1A8FAC7B159 eba1_plafc Erythrocyte-binding antigen 175 protein protein isolate Camp / Malaysia Plasmodium falciparum MKCNISIYFFASFFVLYFAKARNEYDIKENEKFLDVYKEKFNELDKKKYGNVQKTDKKIFTFIENKLDILNNSKFNKRWKSYGTPDNIDKNMSLINKHNNEEMFNNNYQSFLSTSSLIKQNKYVPINAVRVSRILSFLDSRINNGRNTSSNNEVLSNCREKRKGMKWDCKKKNDRSNYVCIPDRRIQLCIVNLSIIKTYTKETMKDHFIEASKKESQLLLKKNDNKYNSKFCNDLKNSFLDYGHLAMGNDMDFGGYSTKAENKIQEVFKGAHGEISEHKIKNFRKEWWNEFREKLWEAMLSEHKNNINNCKNIPQEELQITQWIKEWHGEFLLERDNRSKLPKSKCKNNTLYEACEKECIDPCMKYRDWIIRSKFEWHTLSKEYETQKVPKENAENYLIKISENKNDAKVSLLLNNCDAEYSKYCDCKHTTTLVKSVLNGNDNTIKEKREHIDLDDFSKFGCDKNSVDTNTKVWECKNPYILSTKDVCVPPRRQELCLGNIDRIYDKNLLMIKEHILAIAIYESRILKRKYKNKDDKEVCKIINKTFADIRDIIGGTDYWNDLSNRKLVGKINTNSKYVHRNKKNDKLFRDEWWKVIKKDVWNVISWVFKDKTVCKEDDIENIPQFFRWFSEWGDDYCQDKTKMIETLKVECKEKPCEDDNCKSKCNSYKEWISKKKEEYNKQAKQYQEYQKGNNYKMYSEFKSIKPEVYLKKYSEKCSNLNFEDEFKEELHSDYKNKCTMCPEVKDVPISIIRNNEQTSQEAVPEENTEIAHRTETPSISEGPKGNEQKERDDDSLSKISVSPENSRPETDAKDTSNLLKLKGDVDISMPKAVIGSSPNDNINVTEQGDNISGVNSKPLSDDVRPDKKELEDQNSDESEETVVNHISKSPSINNGDDSGSGSATVSESSSSNTGLSIDDDRNGDTFVRTQDTANTEDVIRKENADKDEDEKGADEERHSTSESLSSPEEKMLTDNEGGNSLNHEEVKEHTSNSDNVQQSGGIVNMNVEKELKDTLENPSSSLDEGKAHEELSEPNLSSDQDMSNTPGPLDNTSEETTERISNNEYKVNEREDERTLTKEYEDIVLKSHMNRESDDGELYDENSDLSTVNDESEDAEAKMKGNDTSEMSHNSSQHIESDQQKNDMKTVGDLGTTHVQNEISVPVTGEIDEKLRESKESKIHKAEEERLSHTDIHKINPEDRNSNTLHLKDIRNEENERHLTNQNINISQERDLQKHGFHTMNNLHGDGVSERSQINHSHHGNRQDRGGNSGNVLNMRSNNNNFNNIPSRYNLYDKKLDLDLYENRNDSTTKELIKKLAEINKCENEISVKYCDHMIHEEIPLKTCTKEKTRNLCCAVSDYCMSYFTYDSEEYYNCTKREFDDPSYTCFRKEAFSSMIFKFLITNKIYYYFYTYKTAKVTIKKINFSLIFFFFFSF 32A4309021B1C3D6 glpa_human Glycophorin-A PAS-2 Sialoglycoprotein alpha MN sialoglycoprotein GYPA GPA protein protein human Homo sapiens Human MYGKIIFVLLLSEIVSISASSTTGVAMHTSTSSSVTKSYISSQTNDTHKRDTYAATPRAHEVSEISVRTVYPPEEETGERVQLAHHFSEPEITLIIFGVMAGVIGTILLISYGIRRLIKKSPSDVKPLPSPDTDVPLSSVEIENPETSDQ 48A5450E22FA99C9 lec_soybn Lectin LE1 Agglutinin SBA protein protein soybn Glycine max (Soybean) Glycine hispida MATSKLKTQNVVVSLSLTLTLVLVLLTSKANSAETVSFSWNKFVPKQPNMILQGDAIVTSSGKLQLNKVDENGTPKPSSLGRALYSTPIHIWDKETGSVASFAASFNFTFYAPDTKRLADGLAFFLAPIDTKPQTHAGYLGLFNENESGDQVVAVEFDTFRNSWDPPNPHIGINVNSIRSIKTTSWDLANNKVAKVLITYDASTSLLVASLVYPSQRTSNILSDVVDLKTSLPEWVRIGFSAATGLDIPGESHDVLSWSFASNLPHASSNIDPLDLTSFVLHEAI B3704533C9315C52 gypa-le1-1 1 6 unspecified role unspecified role prey prey region alexa 488 label alexa 488 label undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined soybn Glycine max (Soybean) Glycine hispida 5 unspecified role unspecified role bait bait purified purified region glycores glycosylated residue GlycoRes undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined prerequisite-ptm prerequisite-ptm prerequisite ptm required ptm prerequisite-ptm required-ptm c_terminus 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 c_terminus fc-igg tag fc-igg tag c-term range C-terminal range c-term range c-term range C-terminal range c-term range ecd_delta_exon3 sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain human-293f Homo sapiens HEK293F cell line 293f embryonic kidney cell 293f physical association physical association Figure 3 C/D SBA from Glycine max binds to alpha- and beta-N-acetyl-galactosamine (GalNAc). galactose-lecg_arahy-1 1 3 unspecified role unspecified role prey prey region alexa 488 label alexa 488 label undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined arahy Arachis hypogaea (Peanut) 5 unspecified role unspecified role bait bait purified purified region glycores glycosylated residue GlycoRes undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined prerequisite-ptm prerequisite-ptm prerequisite ptm required ptm prerequisite-ptm required-ptm c_terminus 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 c_terminus fc-igg tag fc-igg tag c-term range C-terminal range c-term range c-term range C-terminal range c-term range ecd_delta_exon3 sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain human-293f Homo sapiens HEK293F cell line 293f embryonic kidney cell 293f physical association physical association Figure 3 e/f It is stated that PNA from Arachis hypogaea is specific for beta-galactose (or beta-galactose linked to N-Acetylgalactosamine). gypa-lecg_arahy-1 2 4 unspecified role unspecified role prey prey P19214:p.[Asn147Gln;Ser194Ala;Ser339Ala;Thr350Ala;Asn544Gln] mutation mutation certain certain sequence position certain certain certain sequence position certain N Q certain certain sequence position certain certain certain sequence position certain S A certain certain sequence position certain certain certain sequence position certain S A certain certain sequence position certain certain certain sequence position certain T A certain certain sequence position certain certain certain sequence position certain N Q Thu Aug 17 12:25:25 BST 2017 This feature has been corrected as a result of our quality control procedures. The original label was 'asn147glu,ser194ala,ser339ala,thr350ala,asn544glu' Wed May 02 16:29:09 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'asn147gln,ser194ala,ser339ala,thr350ala,asn544gln' Tue Jun 30 19:08:10 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.[Asn147Gln;Ser194Ala;Ser339Ala;Thr350Ala;Asn544Gln]' RII_region sufficient to bind sufficient binding region sufficient to bind range range range range picpa Pichia pastoris (Yeast) Yeast Pichia pastoris 5 unspecified role unspecified role bait bait purified purified P02724:p.Ser66Ala mutation decreasing strength mutation decreasing interaction strength mutation decreasing strength certain certain sequence position certain certain certain sequence position certain S A Wed May 02 14:31:12 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'ser66ala' Tue Jun 30 19:22:11 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Ser66Ala' P02724:p.Thr69Ala mutation decreasing strength mutation decreasing interaction strength mutation decreasing strength certain certain sequence position certain certain certain sequence position certain T A Wed May 02 18:02:09 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'thr69ala' Tue Jun 30 16:34:23 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Thr69Ala' region glycores glycosylated residue GlycoRes undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined c_terminus 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 c_terminus fc-igg tag fc-igg tag c-term range C-terminal range c-term range c-term range C-terminal range c-term range ecd_delta_exon3 sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain P02724:p.Ser63Ala mutation with no effect mutation with no effect mutation with no effect mutation not affecting interaction certain certain sequence position certain certain certain sequence position certain S A Wed May 02 19:19:35 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'ser63ala' Tue Jun 30 16:13:15 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Ser63Ala' P02724:p.Thr69Ala mutation decreasing strength mutation decreasing interaction strength mutation decreasing strength certain certain sequence position certain certain certain sequence position certain T A Wed May 02 17:46:17 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'thr69ala' Tue Jun 30 18:17:12 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Thr69Ala' P02724:p.[Ser63Ala;Ser66Ala;Thr69Ala] mutation disrupting strength mutation disrupting interaction strength mutation disrupting strength certain certain sequence position certain certain certain sequence position certain S A certain certain sequence position certain certain certain sequence position certain S A certain certain sequence position certain certain certain sequence position certain T A Wed May 02 17:39:05 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'ser63ala,ser66ala,thr69ala' Tue Jun 30 17:56:56 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.[Ser63Ala;Ser66Ala;Thr69Ala]' exon3 required to bind necessary binding region required to bind certain certain sequence position certain certain certain sequence position certain human-293f Homo sapiens HEK293F cell line 293f embryonic kidney cell 293f physical association physical association Figure 4, Supp fig 3 neuraminidase treatment