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/ Cysteine 73 in bleomycin hydrolase is critical for amyloid precursor protein processing. Cysteine 73 in bleomycin hydrolase is critical for amyloid precursor protein processing. Biochem. Biophys. Res. Commun. (0006-291X) 2001 imex curation Lefterov IM., Koldamova RP., Lefterova MI., Schwartz DR., Lazo JS. Neurodegeneration - Publications depicting interactions involved in neurodegenerative diseases. Apoptosis - Interactions involving proteins with a function related to apoptosis Alzheimers - Interactions investigated in the context of Alzheimers disease In vitro binding assays performed using GST-hBH fusion proteins immobilized on glutathione-Sepharose beads and incubated with APP proteins transcribed and translated in vitro as S-35 labelled or non labelled proteins using TnT system (Promega). As a negative control APP was incubated with GST alone. The bound proteins were eluted and resolved on SDS page and probed with an N terminal APP antibody LN27. The binding was quantified by scanning the radioactive gels. lazo@pitt.edu 27-Oct-2007: Contacted by JYOTI. Only protein-protein interactions yeasx Saccharomyces cerevisiae (Baker's yeast) Baker's yeast 2 hybrid two hybrid 2H 2h Gal4 transcription regeneration two-hybrid yeast two hybrid 2-hybrid classical two hybrid classical two hybrid 2H two-hybrid 2-hybrid Gal4 transcription regeneration 2h yeast two hybrid Y2H Y-2H 2 hybrid nucleotide sequence nucleotide sequence identification sequence cloning sequence cloning nucleotide sequence Lefterov IM., Koldamova RP., Lefterova MI., Schwartz DR., Lazo JS. Apoptosis - Interactions involving proteins with a function related to apoptosis The authors do not describe how clone identity was confirmed. They simply say that "5 of the true positive clones were deltaBH". This is assumed to be by sequence verification. In vitro binding assays performed using GST-hBH fusion proteins immobilized on glutathione-Sepharose beads and incubated with APP proteins transcribed and translated in vitro as S-35 labelled or non labelled proteins using TnT system (Promega). As a negative control APP was incubated with GST alone. The bound proteins were eluted and resolved on SDS page and probed with an N terminal APP antibody LN27. The binding was quantified by scanning the radioactive gels. lazo@pitt.edu Accepted 2007-MAY-14 by ORCHARD. Biochem. Biophys. Res. Commun. (0006-291X) 2001 Only protein-protein interactions imex curation Cysteine 73 in bleomycin hydrolase is critical for amyloid precursor protein processing. Cysteine 73 in bleomycin hydrolase is critical for amyloid precursor protein processing. Biochem. Biophys. Res. Commun. (0006-291X) 2001 imex curation Lefterov IM., Koldamova RP., Lefterova MI., Schwartz DR., Lazo JS. Neurodegeneration - Publications depicting interactions involved in neurodegenerative diseases. Apoptosis - Interactions involving proteins with a function related to apoptosis Alzheimers - Interactions investigated in the context of Alzheimers disease In vitro binding assays performed using GST-hBH fusion proteins immobilized on glutathione-Sepharose beads and incubated with APP proteins transcribed and translated in vitro as S-35 labelled or non labelled proteins using TnT system (Promega). As a negative control APP was incubated with GST alone. The bound proteins were eluted and resolved on SDS page and probed with an N terminal APP antibody LN27. The binding was quantified by scanning the radioactive gels. lazo@pitt.edu 27-Oct-2007: Contacted by JYOTI. Only protein-protein interactions in vitro In vitro pull down pull down western blot western blot Immuno blot Immuno blot Lefterov IM., Koldamova RP., Lefterova MI., Schwartz DR., Lazo JS. Apoptosis - Interactions involving proteins with a function related to apoptosis In vitro binding assays performed using GST-hBH fusion proteins immobilized on glutathione-Sepharose beads and incubated with APP proteins transcribed and translated in vitro as S-35 labelled or non labelled proteins using TnT system (Promega). As a negative control APP was incubated with GST alone. The bound proteins were eluted and resolved on SDS page and probed with an N terminal APP antibody LN27. The binding was quantified by scanning the radioactive gels. lazo@pitt.edu Accepted 2007-MAY-14 by ORCHARD. 27-Oct-2007: Contacted by JYOTI. Biochem. Biophys. Res. Commun. (0006-291X) 2001 Only protein-protein interactions imex curation p05067-4 Amyloid-beta precursor protein APP A4 AD1 ABPP Alzheimer disease amyloid protein Cerebral vascular amyloid peptide PreA4 Protease nexin-II APPI PreA4 695 Amyloid precursor protein Amyloid-beta A4 protein Alzheimer disease amyloid A4 protein homolog Amyloid-beta (A4) precursor protein protein protein human Homo sapiens Human MLPGLALLLLAAWTARALEVPTDGNAGLLAEPQIAMFCGRLNMHMNVQNGKWDSDPSGTKTCIDTKEGILQYCQEVYPELQITNVVEANQPVTIQNWCKRGRKQCKTHPHFVIPYRCLVGEFVSDALLVPDKCKFLHQERMDVCETHLHWHTVAKETCSEKSTNLHDYGMLLPCGIDKFRGVEFVCCPLAEESDNVDSADAEEDDSDVWWGGADTDYADGSEDKVVEVAEEEEVAEVEEEEADDDEDDEDGDEVEEEAEEPYEEATERTTSIATTTTTTTESVEEVVRVPTTAASTPDAVDKYLETPGDENEHAHFQKAKERLEAKHRERMSQVMREWEEAERQAKNLPKADKKAVIQHFQEKVESLEQEAANERQQLVETHMARVEAMLNDRRRLALENYITALQAVPPRPRHVFNMLKKYVRAEQKDRQHTLKHFEHVRMVDPKKAAQIRSQVMTHLRVIYERMNQSLSLLYNVPAVAEEIQDEVDELLQKEQNYSDDVLANMISEPRISYGNDALMPSLTETKTTVELLPVNGEFSLDDLQPWHSFGADSVPANTENEVEPVDARPAADRGLTTRPGSGLTNIKTEEISEVKMDAEFRHDSGYEVHHQKLVFFAEDVGSNKGAIIGLMVGGVVIATVIVITLVMLKKKQYTSIHHGVVEVDAAVTPEERHLSKMQQNGYENPTYKFFEQMQN DIP protein P05067-4 original sequence version: 183 DIP protein P05067-4 original sequence version: 198 DIP protein P05067-4 original sequence version: 224 A major isoform. 4F6EA0139F969D56 blmh_human Bleomycin hydrolase BLMH protein protein human Homo sapiens Human MSSSGLNSEKVAALIQKLNSDPQFVLAQNVGTTHDLLDICLKRATVQRAQHVFQHAVPQEGKPITNQKSSGRCWIFSCLNVMRLPFMKKLNIEEFEFSQSYLFFWDKVERCYFFLSAFVDTAQRKEPEDGRLVQFLLMNPANDGGQWDMLVNIVEKYGVIPKKCFPESYTTEATRRMNDILNHKMREFCIRLRNLVHSGATKGEISATQDVMMEEIFRVVCICLGNPPETFTWEYRDKDKNYQKIGPITPLEFYREHVKPLFNMEDKICLVNDPRPQHKYNKLYTVEYLSNMVGGRKTLYNNQPIDFLKKMVAASIKDGEAVWFGCDVGKHFNSKLGLSDMNLYDHELVFGVSLKNMNKAERLTFGESLMTHAMTFTAVSEKDDQDGAFTKWRVENSWGEDHGHKGYLCMTDEWFSEYVYEVVVDRKHVPEEVLAVLEQEPIILPAWDPMGALAE 577E86241EB0D460 blmh-aplp1 2 4 unspecified role unspecified role bait bait region gst tag glutathione s tranferase tag glutathione S-tranferase tag glutathione S-tranferase tag gst 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 ecoli Escherichia coli (strain K12) 3 unspecified role unspecified role prey prey region 35s radiolabel 35s radiolabel 35S s35 radiolabelled S35 s35 radiolabelled 35S S35 undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined in vitro In vitro direct interaction direct interaction No interaction was detected between any of the bleomycin hydrolase constructs and a clone including only the extracellular domain of APP. Additionally no interaction was detected between bleomycin truncation variant (170-455) and full length APP. The authors state that this is consistant with bleomycin hydrolase interacting with the cytoplasmic rather than the extracellular domain of APP. Figure 1 aplp1-blmh 1 3 unspecified role unspecified role bait bait 4 unspecified role unspecified role prey prey blmh 14-455 sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain physical association physical association The authors note that full length BH was not found among the positive clones examined "which may be due to inhibition of productive heterologous interactions in the yeast two hybrid environment when BH can oligomerize.".