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/ http://www.ebi.ac.uk/ melnyk-2006-1 Structural determinants for the binding of anthrax lethal factor to oligomeric protective antigen. in vitro In vitro ms of complexes mass spectrometry studies of complexes ms of complexes predetermined predetermined participant predetermined Melnyk RA., Hewitt KM., Lacy DB., Lin HC., Gessner CR., Li S., Woods VL., Collier RJ. Hydrogen-deuterium exchange MS was performed. The number of deuterons incorporated into proteins was determined by MS of peptides obtained by passing protein through an immobilised protease column. Mutation mapping shows that the "mature" protein - minus the signal sequence was used. vwoods@ucsd.edu,jcollier@hms.harvard.edu J. Biol. Chem. (0021-9258) 2006 Only protein-protein interactions imex curation pag_bacan Protective antigen PA-83 Anthrax toxins translocating protein pagA pag pXO1-110 BXA0164 GBAA_pXO1_0164 protein protein bacan Bacillus anthracis MKKRKVLIPLMALSTILVSSTGNLEVIQAEVKQENRLLNESESSSQGLLGYYFSDLNFQAPMVVTSSTTGDLSIPSSELENIPSENQYFQSAIWSGFIKVKKSDEYTFATSADNHVTMWVDDQEVINKASNSNKIRLEKGRLYQIKIQYQRENPTEKGLDFKLYWTDSQNKKEVISSDNLQLPELKQKSSNSRKKRSTSAGPTVPDRDNDGIPDSLEVEGYTVDVKNKRTFLSPWISNIHEKKGLTKYKSSPEKWSTASDPYSDFEKVTGRIDKNVSPEARHPLVAAYPIVHVDMENIILSKNEDQSTQNTDSQTRTISKNTSTSRTHTSEVHGNAEVHASFFDIGGSVSAGFSNSNSSTVAIDHSLSLAGERTWAETMGLNTADTARLNANIRYVNTGTAPIYNVLPTTSLVLGKNQTLATIKAKENQLSQILAPNNYYPSKNLAPIALNAQDDFSSTPITMNYNQFLELEKTKQLRLDTDQVYGNIATYNFENGRVRVDTGSNWSEVLPQIQETTARIIFNGKDLNLVERRIAAVNPSDPLETTKPDMTLKEALKIAFGFNEPNGNLQYQGKDITEFDFNFDQQTSQNIKNQLAELNATNIYTVLDKIKLNAKMNILIRDKRFHYDRNNIAVGADESVVKEAHREVINSSTEGLLLNIDKDIRKILSGYIVEIEDTEGLKEVINDRYDMLNISSLRQDGKTFIDFKKYNDKLPLYISNPNYKVNVYAVTKENTIINPSENGDTSTNGIKKILIFSKKGYEIG lef_bacan Lethal factor Anthrax lethal toxin endopeptidase component lef pXO1-107 BXA0172 GBAA_pXO1_0172 protein protein bacan Bacillus anthracis MNIKKEFIKVISMSCLVTAITLSGPVFIPLVQGAGGHGDVGMHVKEKEKNKDENKRKDEERNKTQEEHLKEIMKHIVKIEVKGEEAVKKEAAEKLLEKVPSDVLEMYKAIGGKIYIVDGDITKHISLEALSEDKKKIKDIYGKDALLHEHYVYAKEGYEPVLVIQSSEDYVENTEKALNVYYEIGKILSRDILSKINQPYQKFLDVLNTIKNASDSDGQDLLFTNQLKEHPTDFSVEFLEQNSNEVQEVFAKAFAYYIEPQHRDVLQLYAPEAFNYMDKFNEQEINLSLEELKDQRMLARYEKWEKIKQHYQHWSDSLSEEGRGLLKKLQIPIEPKKDDIIHSLSQEEKELLKRIQIDSSDFLSTEEKEFLKKLQIDIRDSLSEEEKELLNRIQVDSSNPLSEKEKEFLKKLKLDIQPYDINQRLQDTGGLIDSPSINLDVRKQYKRDIQNIDALLHQSIGSTLYNKIYLYENMNINNLTATLGADLVDSTDNTKINRGIFNEFKKNFKYSISSNYMIVDINERPALDNERLKWRIQLSPDTRAGYLENGKLILQRNIGLEIKDVQIIKQSEKEYIRIDAKVVPKSKIDTKIQEAQLNINQEWNKALGLPKYTKLITFNVHNRYASNIVESAYLILNEWKNNIQSDLIKKVTNYLVDGNGRFVFTDITLPNIAEQYTHQDEIYEQVHSKGLYVPESRSILLHGPSKGVELRNDSEGFIHEFGHAVDDYAGYLLDKNQSDLVTNSKKFIDIFKEEGSNLTSYGRTNEAEFFAEAFRLMHSTDHAERLKVQKNAPKTFQFINDQIKFIINS lef-paga 502981 N/A 502982 unspecified role unspecified role neutral component neutral component c-terminal PA83 sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain range range false P13423:p.Asp541Lys mutation mutation certain certain sequence position certain certain certain sequence position certain false P13423:p.[Lys228Glu;Arg497Ala;Arg499Asp] mutation mutation certain certain sequence position certain certain certain sequence position certain false certain certain sequence position certain certain certain sequence position certain false certain certain sequence position certain certain certain sequence position certain false ecoli Escherichia coli (strain K12) Stoichiometry: 2.0 N/A 502983 unspecified role unspecified role neutral component neutral component P15917:p.Asn173Ala mutation decreasing mutation decreasing interaction hotspot hotspot mutation decreasing certain certain sequence position certain certain certain sequence position certain false P15917:p.Lys176Ala mutation decreasing mutation decreasing interaction hotspot hotspot mutation decreasing certain certain sequence position certain certain certain sequence position certain false region Only the mutant proteins were radiolabeled at Met residues. 35s radiolabel 35s radiolabel 35S s35 radiolabelled S35 s35 radiolabelled 35S S35 undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined false P15917:p.Asp169Lys mutation decreasing mutation decreasing interaction hotspot hotspot mutation decreasing certain certain sequence position certain certain certain sequence position certain false n-terminal LFN sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain false P15917:p.Asp139Ala mutation decreasing mutation decreasing interaction hotspot hotspot mutation decreasing certain certain sequence position certain certain certain sequence position certain false P15917:p.Tyr141Lys mutation decreasing mutation decreasing interaction hotspot hotspot mutation decreasing certain certain sequence position certain certain certain sequence position certain false P15917:p.Glu168Ala mutation decreasing mutation decreasing interaction hotspot hotspot mutation decreasing certain certain sequence position certain certain certain sequence position certain false P15917:p.Glu168Lys mutation decreasing mutation decreasing interaction hotspot hotspot mutation decreasing certain certain sequence position certain certain certain sequence position certain false P15917:p.Asp169Ala mutation decreasing mutation decreasing interaction hotspot hotspot mutation decreasing certain certain sequence position certain certain certain sequence position certain false ecoli Escherichia coli (strain K12) Stoichiometry: 1.0 physical association physical association false false false A paper describing the plasmid pET22b-PA indicates that PA is only 735 residues long but the Uniprot entry is 764 residues. This paper does not specify the length. The plasmid pET22b-PA does not contain the sequence coding for the signal peptide. Thus the mutations described in the paper (asp512lys and lys199glu&arg468ala&arg470asp) are mapped to residues 29 residues later in the Uniprot entry. They first examined LFN bound to the heptameric pre-pore, but because of the inherent tendency of these complexes to aggregate at high concentrations, they turned to a dimeric formof PA as an alternative binding partner for LFN. When two mutated,oligomerization-deficient forms of PA are combined in the presence of LFN, a ternary complex is formed containing one molecule of each species. PA was activated by incubation with trypsin at a final trypsin. Four discrete sites in LFN (i.e. residues 95�120, DX site 1; 137�147, DX site 2; 177�189, DX site 3; and 223�235, DX site 4) became significantly more protected from solvent exchange in the presence of PA. 2 and 3 Mapping all of the binding-defective mutants onto the four sites on the structure of LFN revealed the breadth of the binding surface defined by residues in DX sites 2 and 3. The distance between the most distal bindingdefective mutants, Asp-182 on DX site 3 to Glu-135 on DX site 2, was seen to be 40 A. The mutations in LFN listed above are those the authors specify as causing a defect in binding, however, from Fig. 3 A it seems that some of the other mutations did cause a decrease in binding.