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/ reczek-2007-3 LIMP-2 Is a Receptor for Lysosomal Mannose-6-Phosphate-Independent Targeting of beta-Glucocerebrosidase. mouse-embryonic_fibroblast Mus musculus embryonic fibroblasts embryonic_fibroblast Embryonic fibroblast Fibroblasts taken from an embryonic as opposed to a foetal or adult source. Foetus is defined as the stage at which tissue diferentiation is initiated. Tissue taken from an embryonic as opposed to a foetal or adult source. Embryo is defined as the development stage prior to organ formation. fluorescence imaging fluorescence microscopy fluorescence imaging antibody detection identification by antibody antibody detection Reczek D., Schwake M., Schroder J., Hughes H., Blanz J., Jin X., Brondyk W., Van Patten S., Edmunds T., Saftig P. Purified recombinant Gba was covalently coupled to CNBr-activated Sepharose 4B psaftig@biochem.uni-kiel.de; david.reczek@genzyme.com Cell (0092-8674) 2007 Only protein-protein interactions 6 imex curation reczek-2007-4 LIMP-2 Is a Receptor for Lysosomal Mannose-6-Phosphate-Independent Targeting of beta-Glucocerebrosidase. chlae-cos_7 Cercopithecus aethiops simian cells transformed with SV40 Grivet cos_7 cercopithecus aethiops, by transformation with SV40 anti tag coip anti tag coimmunoprecipitation anti tag coip western blot western blot Immuno blot Immuno blot Reczek D., Schwake M., Schroder J., Hughes H., Blanz J., Jin X., Brondyk W., Van Patten S., Edmunds T., Saftig P. Purified recombinant Gba was covalently coupled to CNBr-activated Sepharose 4B psaftig@biochem.uni-kiel.de; david.reczek@genzyme.com Cell (0092-8674) 2007 Only protein-protein interactions 6 imex curation reczek-2007-1 LIMP-2 Is a Receptor for Lysosomal Mannose-6-Phosphate-Independent Targeting of beta-Glucocerebrosidase. in vitro In vitro pull down pull down de novo protein sequence de novo protein sequencing by mass spectrometry de novo protein sequence Reczek D., Schwake M., Schroder J., Hughes H., Blanz J., Jin X., Brondyk W., Van Patten S., Edmunds T., Saftig P. Purified recombinant Gba was covalently coupled to CNBr-activated Sepharose 4B psaftig@biochem.uni-kiel.de; david.reczek@genzyme.com Cell (0092-8674) 2007 Only protein-protein interactions 6 imex curation reczek-2007-2 LIMP-2 Is a Receptor for Lysosomal Mannose-6-Phosphate-Independent Targeting of beta-Glucocerebrosidase. in vitro In vitro pull down pull down weight by staining molecular weight estimation by staining weight by staining Reczek D., Schwake M., Schroder J., Hughes H., Blanz J., Jin X., Brondyk W., Van Patten S., Edmunds T., Saftig P. Purified recombinant Gba was covalently coupled to CNBr-activated Sepharose 4B psaftig@biochem.uni-kiel.de; david.reczek@genzyme.com Cell (0092-8674) 2007 Only protein-protein interactions 6 imex curation glcm_mouse Lysosomal acid glucosylceramidase Beta-glucocerebrosidase Acid beta-glucosidase D-glucosyl-N-acylsphingosine glucohydrolase Gba Cholesterol glucosyltransferase Cholesteryl-beta-glucosidase protein protein mouse Mus musculus Mouse MAARLIGFFLFQAVSWAYGAQPCIPKSFGYSSVVCVCNASYCDSLDPVTLPALGTFSRYESTRRGRRMELSVGAIQANRTGTGLLLTLQPEKKFQKVKGFGGAMTDATALNILALSPPTQKLLLRSYFSTNGIEYNIIRVPMASCDFSIRVYTYADTPNDFQLSNFSLPEEDTKLKIPLIHQALKMSSRPISLFASPWTSPTWLKTNGRVNGKGSLKGQPGDIFHQTWANYFVKFLDAYAKYGLRFWAVTAENEPTAGLFTGYPFQCLGFTPEHQRDFISRDLGPALANSSHDVKLLMLDDQRLLLPRWAEVVLSDPEAAKYVHGIAVHWYMDFLAPAKATLGETHRLFPNTMLFASEACVGSKFWEQSVRLGSWDRGMQYSHSIITNLLYHVTGWTDWNLALNPEGGPNWVRNFVDSPIIVDIPKDAFYKQPMFYHLGHFSKFIPEGSQRVALVASESTDLETVALLRPDGSAVVVVLNRSSEDVPLTISDPDLGFLETVSPGYSIHTYLWRRQ scrb2_mouse Lysosome membrane protein 2 Lysosome membrane protein II Scavenger receptor class B member 2 85 kDa lysosomal membrane sialoglycoprotein Scarb2 protein protein mouse Mus musculus Mouse MGRCCFYTAGTLSLLLLVTSVTLLVARVFQKAVDQTIEKNMVLQNGTKVFNSWEKPPLPVYIQFYFFNVTNPEEILQGEIPLLEEVGPYTYRELRNKANIQFGENGTTISAVTNKAYVFERNQSVGDPNVDLIRTINIPLLTVVDLAQLTLLRELIEAMLKAYQQKLFVIHTVHELLWGYKDEILSLVHIFKPDVSPNFGLFYERNGTNDGEYVFLTGEDNYLNFSKIVEWNGKTSLDWWTTDTCNMINGTDGDSFHPLISKDEVLYLFPSDLCRSVHITFSSFENVEGLPAFRYKVPAEILANTSENAGFCIPEGNCMDSGVLNISICKNGAPIIMSFPHFYQADEKFVSAIKGMHPNKEEHESFVDINPLTGIILRGAKRFQINTYVRKLDDFVETGDIRTMVFPVMYLNESVLIDKETANQLKSVINTTLVVTNIPYIIMALGVFFGLVFTWLACRGQGSMDEGTADERAPLIRT lamp2_mouse Lysosome-associated membrane glycoprotein 2 Lysosomal membrane glycoprotein type B CD107 antigen-like family member B Lamp2 Lamp-2 protein protein mouse Mus musculus Mouse MCLSPVKGAKLILIFLFLGAVQSNALIVNLTDSKGTCLYAEWEMNFTITYETTNQTNKTITIAVPDKATHDGSSCGDDRNSAKIMIQFGFAVSWAVNFTKEASHYSIHDIVLSYNTSDSTVFPGAVAKGVHTVKNPENFKVPLDVIFKCNSVLTYNLTPVVQKYWGIHLQAFVQNGTVSKNEQVCEEDQTPTTVAPIIHTTAPSTTTTLTPTSTPTPTPTPTPTVGNYSIRNGNTTCLLATMGLQLNITEEKVPFIFNINPATTNFTGSCQPQSAQLRLNNSQIKYLDFIFAVKNEKRFYLKEVNVYMYLANGSAFNISNKNLSFWDAPLGSSYMCNKEQVLSVSRAFQINTFNLKVQPFNVTKGQYSTAQDCSADEDNFLVPIAVGAALGGVLILVLLAYFIGLKRHHTGYEQF glcm_cerae_protein African green monkey Glucosylceramidase protein protein chlae Chlorocebus aethiops Cercopithecus aethiops Grivet Green monkey An amino acid sequence which is not represented in UniProt glcm_human Lysosomal acid glucosylceramidase Beta-glucocerebrosidase Acid beta-glucosidase D-glucosyl-N-acylsphingosine glucohydrolase Alglucerase Imiglucerase GBA GC GLUC Cholesterol glucosyltransferase Cholesteryl-beta-glucosidase protein protein human Homo sapiens Human MEFSSPSREECPKPLSRVSIMAGSLTGLLLLQAVSWASGARPCIPKSFGYSSVVCVCNATYCDSFDPPTFPALGTFSRYESTRSGRRMELSMGPIQANHTGTGLLLTLQPEQKFQKVKGFGGAMTDAAALNILALSPPAQNLLLKSYFSEEGIGYNIIRVPMASCDFSIRTYTYADTPDDFQLHNFSLPEEDTKLKIPLIHRALQLAQRPVSLLASPWTSPTWLKTNGAVNGKGSLKGQPGDIYHQTWARYFVKFLDAYAEHKLQFWAVTAENEPSAGLLSGYPFQCLGFTPEHQRDFIARDLGPTLANSTHHNVRLLMLDDQRLLLPHWAKVVLTDPEAAKYVHGIAVHWYLDFLAPAKATLGETHRLFPNTMLFASEACVGSKFWEQSVRLGSWDRGMQYSHSIITNLLYHVVGWTDWNLALNPEGGPNWVRNFVDSPIIVDITKDTFYKQPMFYHLGHFSKFIPEGSQRVGLVASQKNDLDAVALMHPDGSAVVVVLNRSSKDVPLTIKDPAVGFLETISPGYSIHTYLWRRQ scrb2_human Lysosome membrane protein 2 Lysosome membrane protein II Scavenger receptor class B member 2 85 kDa lysosomal membrane sialoglycoprotein CD36 antigen-like 2 SCARB2 CD36L2 LIMPII LIMP2 protein protein human Homo sapiens Human MGRCCFYTAGTLSLLLLVTSVTLLVARVFQKAVDQSIEKKIVLRNGTEAFDSWEKPPLPVYTQFYFFNVTNPEEILRGETPRVEEVGPYTYRELRNKANIQFGDNGTTISAVSNKAYVFERDQSVGDPKIDLIRTLNIPVLTVIEWSQVHFLREIIEAMLKAYQQKLFVTHTVDELLWGYKDEILSLIHVFRPDISPYFGLFYEKNGTNDGDYVFLTGEDSYLNFTKIVEWNGKTSLDWWITDKCNMINGTDGDSFHPLITKDEVLYVFPSDFCRSVYITFSDYESVQGLPAFRYKVPAEILANTSDNAGFCIPEGNCLGSGVLNVSICKNGAPIIMSFPHFYQADERFVSAIEGMHPNQEDHETFVDINPLTGIILKAAKRFQINIYVKKLDDFVETGDIRTMVFPVMYLNESVHIDKETASRLKSMINTTLIITNIPYIIMALGVFFGLVFTWLACKGQGSMDEGTADERAPLIRT gba-scarb2-3 Interaction between Gba and Scarb2 identified by colocalization 682734 N/A 682738 unspecified role unspecified role neutral component neutral component N/A 682739 unspecified role unspecified role neutral component neutral component region myc tag myc tag EUKLISEED epitope tag EUKLISEED epitope tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined false colocalization colocalization false false false Fig. 3C O35114 transfection gba-scarb2-4 Interaction between Gba, Lamp2 and Scarb2 identified by colocalization 682734 N/A 682738 unspecified role unspecified role neutral component neutral component N/A 682740 unspecified role unspecified role neutral component neutral component N/A 682739 unspecified role unspecified role neutral component neutral component region myc tag myc tag EUKLISEED epitope tag EUKLISEED epitope tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined false colocalization colocalization false false false Fig. 3G O35114 transfection gba-scarb2-5 Interaction between Gba and Scarb2 identified by coimmunoprecipitation 682735 N/A 682741 unspecified role unspecified role prey prey N/A 682739 unspecified role unspecified role bait bait region myc tag myc tag EUKLISEED epitope tag EUKLISEED epitope tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined false O35114:p.Leu160Pro mutation decreasing mutation decreasing interaction hotspot hotspot mutation decreasing mutation analysis mutation analysis certain certain sequence position certain certain certain sequence position certain false coiled coil region required to bind necessary binding region required to bind deletion analysis deletion analysis certain certain sequence position certain certain certain sequence position certain false physical association physical association false false false Fig. 4G and 5E A Scar2 mutant fused to an endoplasmic retention signal was also coimmunoprecipitated with Gba. O35114 transfection gba-scarb2-6 Interaction between Gba and Scarb2 identified by coimmunoprecipitation 682735 N/A 682739 unspecified role unspecified role bait bait region myc tag myc tag EUKLISEED epitope tag EUKLISEED epitope tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined false N/A 682738 unspecified role unspecified role prey prey physical association physical association false false false Fig. S5A P17439; O35114 transfection gba-scarb2-1 Interaction between Gba and Scarb2 identified by pull down 682736 N/A 682742 unspecified role unspecified role bait bait spofr Spodoptera frugiperda (Fall armyworm) Fall armyworm N/A 682739 unspecified role unspecified role prey prey mouse-liver Mus musculus liver liver physical association physical association false false false Fig. 1A, 1B Scarb2 competed off by excess GBA gba-scarb2-2 Interaction between GBA and SCARB2 identified by pull down 682737 N/A 682738 unspecified role unspecified role bait bait hpc4 tagged region tag tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined false spofr Spodoptera frugiperda (Fall armyworm) Fall armyworm N/A 682743 unspecified role unspecified role prey prey lumenal region sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain false spofr Spodoptera frugiperda (Fall armyworm) Fall armyworm direct interaction direct interaction false false false Fig. 1D, 1E Binding of the SCARB2 lumenal domain to the GBA resin was favored at neutral pH and attenuated as the pH approached that typical of the lysosomal lumen.