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/ muda-2013-6 Parkinson-related LRRK2 mutation R1441C/G/H impairs PKA phosphorylation of LRRK2 and disrupts its interaction with 14-3-3. in vitro In vitro fps fluorescence polarization spectroscopy Fluorescence anisotropy FPS FPS Fluorescence anisotropy fps tag fluorescence tag visualisation by fluorescence tag fluorescence Muda K., Bertinetti D., Gesellchen F., Hermann J.S., von Zweydorf F., Geerlof A., Jacob A., Ueffing M., Gloeckner C.J., Herberg F.W. Parkinsons - Interactions investigated in the context of Parkinsons disease Affinomics - Interactions curated for the Affinomics consortium. Peptides were phosphorylated by PKA preincubation. As a negative control a PKA inhibitor was used in some preincubations. The authors confirm that "PKA" is a Human protein, ID is UniProt P17612 and it was expressed in E.coli. and purified by affinity purification. herberg@uni-kassel.de Proc. Natl. Acad. Sci. U.S.A. (0027-8424) 2013 Only protein-protein interactions imex curation Neurodegeneration - Publications depicting interactions involved in neurodegenerative diseases. muda-2013-1 Parkinson-related LRRK2 mutation R1441C/G/H impairs PKA phosphorylation of LRRK2 and disrupts its interaction with 14-3-3. in vitro In vitro protein kinase assay protein kinase assay ms participant Identification by mass spectrometry ms participant Muda K., Bertinetti D., Gesellchen F., Hermann J.S., von Zweydorf F., Geerlof A., Jacob A., Ueffing M., Gloeckner C.J., Herberg F.W. Parkinsons - Interactions investigated in the context of Parkinsons disease Affinomics - Interactions curated for the Affinomics consortium. Results were obtained from either the full length LRRK2 Kinase Dead mutant alone, or the full length molecule concurrently with the truncated Delta-967 LRRK2 Kinase Dead mutant (aas 968-2527), or from the deletion construct alone. The authors confirm that "PKA" is a Human protein, ID is UniProt P17612 and it was expressed in E.coli. and purified by affinity purification. herberg@uni-kassel.de Proc. Natl. Acad. Sci. U.S.A. (0027-8424) 2013 Only protein-protein interactions imex curation Neurodegeneration - Publications depicting interactions involved in neurodegenerative diseases. muda-2013-3 Parkinson-related LRRK2 mutation R1441C/G/H impairs PKA phosphorylation of LRRK2 and disrupts its interaction with 14-3-3. in vitro In vitro protein kinase assay protein kinase assay weight autoradiogra molecular weight estimation by autoradiography weight autoradiogra Muda K., Bertinetti D., Gesellchen F., Hermann J.S., von Zweydorf F., Geerlof A., Jacob A., Ueffing M., Gloeckner C.J., Herberg F.W. Parkinsons - Interactions investigated in the context of Parkinsons disease Affinomics - Interactions curated for the Affinomics consortium. The authors confirm that "PKA" is a Human protein, ID is UniProt P17612 and it was expressed in E.coli. and purified by affinity purification. herberg@uni-kassel.de Proc. Natl. Acad. Sci. U.S.A. (0027-8424) 2013 Only protein-protein interactions imex curation Neurodegeneration - Publications depicting interactions involved in neurodegenerative diseases. muda-2013-7 Parkinson-related LRRK2 mutation R1441C/G/H impairs PKA phosphorylation of LRRK2 and disrupts its interaction with 14-3-3. in vitro In vitro spr surface plasmon resonance Optical biosensor BIAcore(r) BIAcore(r) Optical biosensor spr predetermined predetermined participant predetermined Muda K., Bertinetti D., Gesellchen F., Hermann J.S., von Zweydorf F., Geerlof A., Jacob A., Ueffing M., Gloeckner C.J., Herberg F.W. Parkinsons - Interactions investigated in the context of Parkinsons disease Affinomics - Interactions curated for the Affinomics consortium. The authors confirm that "PKA" is a Human protein, ID is UniProt P17612 and it was expressed in E.coli. and purified by affinity purification. herberg@uni-kassel.de Proc. Natl. Acad. Sci. U.S.A. (0027-8424) 2013 Only protein-protein interactions imex curation Neurodegeneration - Publications depicting interactions involved in neurodegenerative diseases. muda-2013-2 Parkinson-related LRRK2 mutation R1441C/G/H impairs PKA phosphorylation of LRRK2 and disrupts its interaction with 14-3-3. in vitro In vitro protein kinase assay protein kinase assay western blot western blot Immuno blot Immuno blot Muda K., Bertinetti D., Gesellchen F., Hermann J.S., von Zweydorf F., Geerlof A., Jacob A., Ueffing M., Gloeckner C.J., Herberg F.W. Parkinsons - Interactions investigated in the context of Parkinsons disease Affinomics - Interactions curated for the Affinomics consortium. Autophosphorylation of LRRK2 was detected by westerm Blotting using an anti-pT2483 antibody - T2483 being a previously identified LRRK2 autophosphorylation site (one of many) - see Reference 7 (Gloeckner J. Proteome Res 2010. PMID:20108944) The authors confirm that "PKA" is a Human protein, ID is UniProt P17612 and it was expressed in E.coli. and purified by affinity purification. herberg@uni-kassel.de Proc. Natl. Acad. Sci. U.S.A. (0027-8424) 2013 Only protein-protein interactions imex curation Neurodegeneration - Publications depicting interactions involved in neurodegenerative diseases. muda-2013-4 Parkinson-related LRRK2 mutation R1441C/G/H impairs PKA phosphorylation of LRRK2 and disrupts its interaction with 14-3-3. in vitro In vitro pull down pull down tag visualisation tag visualisation Muda K., Bertinetti D., Gesellchen F., Hermann J.S., von Zweydorf F., Geerlof A., Jacob A., Ueffing M., Gloeckner C.J., Herberg F.W. Parkinsons - Interactions investigated in the context of Parkinsons disease Affinomics - Interactions curated for the Affinomics consortium. The authors confirm that "PKA" is a Human protein, ID is UniProt P17612 and it was expressed in E.coli. and purified by affinity purification. herberg@uni-kassel.de Proc. Natl. Acad. Sci. U.S.A. (0027-8424) 2013 Only protein-protein interactions imex curation Neurodegeneration - Publications depicting interactions involved in neurodegenerative diseases. muda-2013-5 Parkinson-related LRRK2 mutation R1441C/G/H impairs PKA phosphorylation of LRRK2 and disrupts its interaction with 14-3-3. in vitro In vitro far western blotting far western blotting Affinity blotting Affinity blotting anti tag western anti tag western blot anti tag western Muda K., Bertinetti D., Gesellchen F., Hermann J.S., von Zweydorf F., Geerlof A., Jacob A., Ueffing M., Gloeckner C.J., Herberg F.W. Parkinsons - Interactions investigated in the context of Parkinsons disease Affinomics - Interactions curated for the Affinomics consortium. The authors confirm that "PKA" is a Human protein, ID is UniProt P17612 and it was expressed in E.coli. and purified by affinity purification. herberg@uni-kassel.de Proc. Natl. Acad. Sci. U.S.A. (0027-8424) 2013 Only protein-protein interactions imex curation Neurodegeneration - Publications depicting interactions involved in neurodegenerative diseases. muda-2013-8 Parkinson-related LRRK2 mutation R1441C/G/H impairs PKA phosphorylation of LRRK2 and disrupts its interaction with 14-3-3. in vitro In vitro protein kinase assay protein kinase assay tag visualisation tag visualisation Muda K., Bertinetti D., Gesellchen F., Hermann J.S., von Zweydorf F., Geerlof A., Jacob A., Ueffing M., Gloeckner C.J., Herberg F.W. Parkinsons - Interactions investigated in the context of Parkinsons disease Affinomics - Interactions curated for the Affinomics consortium. The authors confirm that "PKA" is a Human protein, ID is UniProt P17612 and it was expressed in E.coli. and purified by affinity purification. herberg@uni-kassel.de Proc. Natl. Acad. Sci. U.S.A. (0027-8424) 2013 Only protein-protein interactions imex curation Neurodegeneration - Publications depicting interactions involved in neurodegenerative diseases. 1433g_rat 14-3-3 protein gamma Ywhag protein protein rat Rattus norvegicus (Rat) Rat MVDREQLVQKARLAEQAERYDDMAAAMKNVTELNEPLSNEERNLLSVAYKNVVGARRSSWRVISSIEQKTSADGNEKKIEMVRAYREKIEKELEAVCQDVLSLLDNYLIKNCSETQYESKVFYLKMKGDYYRYLAEVATGEKRATVVESSEKAYSEAHEISKEHMQPTHPIRLGLALNYSVFYYEIQNAPEQACHLAKTAFDDAIAELDTLNEDSYKDSTLIMQLLRDNLTLWTSDQQDDDGGEGNN lrrk2_human Leucine-rich repeat serine/threonine-protein kinase 2 LRRK2 PARK8 Dardarin protein protein human Homo sapiens Human MASGSCQGCEEDEETLKKLIVRLNNVQEGKQIETLVQILEDLLVFTYSERASKLFQGKNIHVPLLIVLDSYMRVASVQQVGWSLLCKLIEVCPGTMQSLMGPQDVGNDWEVLGVHQLILKMLTVHNASVNLSVIGLKTLDLLLTSGKITLLILDEESDIFMLIFDAMHSFPANDEVQKLGCKALHVLFERVSEEQLTEFVENKDYMILLSALTNFKDEEEIVLHVLHCLHSLAIPCNNVEVLMSGNVRCYNIVVEAMKAFPMSERIQEVSCCLLHRLTLGNFFNILVLNEVHEFVVKAVQQYPENAALQISALSCLALLTETIFLNQDLEEKNENQENDDEGEEDKLFWLEACYKALTWHRKNKHVQEAACWALNNLLMYQNSLHEKIGDEDGHFPAHREVMLSMLMHSSSKEVFQASANALSTLLEQNVNFRKILLSKGIHLNVLELMQKHIHSPEVAESGCKMLNHLFEGSNTSLDIMAAVVPKILTVMKRHETSLPVQLEALRAILHFIVPGMPEESREDTEFHHKLNMVKKQCFKNDIHKLVLAALNRFIGNPGIQKCGLKVISSIVHFPDALEMLSLEGAMDSVLHTLQMYPDDQEIQCLGLSLIGYLITKKNVFIGTGHLLAKILVSSLYRFKDVAEIQTKGFQTILAILKLSASFSKLLVHHSFDLVIFHQMSSNIMEQKDQQFLNLCCKCFAKVAMDDYLKNVMLERACDQNNSIMVECLLLLGADANQAKEGSSLICQVCEKESSPKLVELLLNSGSREQDVRKALTISIGKGDSQIISLLLRRLALDVANNSICLGGFCIGKVEPSWLGPLFPDKTSNLRKQTNIASTLARMVIRYQMKSAVEEGTASGSDGNFSEDVLSKFDEWTFIPDSSMDSVFAQSDDLDSEGSEGSFLVKKKSNSISVGEFYRDAVLQRCSPNLQRHSNSLGPIFDHEDLLKRKRKILSSDDSLRSSKLQSHMRHSDSISSLASEREYITSLDLSANELRDIDALSQKCCISVHLEHLEKLELHQNALTSFPQQLCETLKSLTHLDLHSNKFTSFPSYLLKMSCIANLDVSRNDIGPSVVLDPTVKCPTLKQFNLSYNQLSFVPENLTDVVEKLEQLILEGNKISGICSPLRLKELKILNLSKNHISSLSENFLEACPKVESFSARMNFLAAMPFLPPSMTILKLSQNKFSCIPEAILNLPHLRSLDMSSNDIQYLPGPAHWKSLNLRELLFSHNQISILDLSEKAYLWSRVEKLHLSHNKLKEIPPEIGCLENLTSLDVSYNLELRSFPNEMGKLSKIWDLPLDELHLNFDFKHIGCKAKDIIRFLQQRLKKAVPYNRMKLMIVGNTGSGKTTLLQQLMKTKKSDLGMQSATVGIDVKDWPIQIRDKRKRDLVLNVWDFAGREEFYSTHPHFMTQRALYLAVYDLSKGQAEVDAMKPWLFNIKARASSSPVILVGTHLDVSDEKQRKACMSKITKELLNKRGFPAIRDYHFVNATEESDALAKLRKTIINESLNFKIRDQLVVGQLIPDCYVELEKIILSERKNVPIEFPVIDRKRLLQLVRENQLQLDENELPHAVHFLNESGVLLHFQDPALQLSDLYFVEPKWLCKIMAQILTVKVEGCPKHPKGIISRRDVEKFLSKKRKFPKNYMSQYFKLLEKFQIALPIGEEYLLVPSSLSDHRPVIELPHCENSEIIIRLYEMPYFPMGFWSRLINRLLEISPYMLSGRERALRPNRMYWRQGIYLNWSPEAYCLVGSEVLDNHPESFLKITVPSCRKGCILLGQVVDHIDSLMEEWFPGLLEIDICGEGETLLKKWALYSFNDGEEHQKILLDDLMKKAEEGDLLVNPDQPRLTIPISQIAPDLILADLPRNIMLNNDELEFEQAPEFLLGDGSFGSVYRAAYEGEEVAVKIFNKHTSLRLLRQELVVLCHLHHPSLISLLAAGIRPRMLVMELASKGSLDRLLQQDKASLTRTLQHRIALHVADGLRYLHSAMIIYRDLKPHNVLLFTLYPNAAIIAKIADYGIAQYCCRMGIKTSEGTPGFRAPEVARGNVIYNQQADVYSFGLLLYDILTTGGRIVEGLKFPNEFDELEIQGKLPDPVKEYGCAPWPMVEKLIKQCLKENPQERPTSAQVFDILNSAELVCLTRRILLPKNVIVECMVATHHNSRNASIWLGCGHTDRGQLSFLDLNTEGYTSEEVADSRILCLALVHLPVEKESWIVSGTQSGTLLVINTEDGKKRHTLEKMTDSVTCLYCNSFSKQSKQKNFLLVGTADGKLAIFEDKTVKLKGAAPLKILNIGNVSTPLMCLSESTNSTERNVMWGGCGTKIFSFSNDFTIQKLIETRTSQLFSYAAFSDSNIITVVVDTALYIAKQNSPVVEVWDKKTEKLCGLIDCVHFLREVMVKENKESKHKMSYSGRVKTLCLQKNTALWIGTGGGHILLLDLSTRRLIRVIYNFCNSVRVMMTAQLGSLKNVMLVLGYNRKNTEGTQKQKEIQSCLTVWDINLPHEVQNLEKHIEVRKELAEKMRRTSVE 1433t_human 14-3-3 protein theta YWHAQ 14-3-3 protein tau 14-3-3 protein T-cell Protein HS1 protein protein human Homo sapiens Human MEKTELIQKAKLAEQAERYDDMATCMKAVTEQGAELSNEERNLLSVAYKNVVGGRRSAWRVISSIEQKTDTSDKKLQLIKDYREKVESELRSICTTVLELLDKYLIANATNPESKVFYLKMKGDYFRYLAEVACGDDRKQTIDNSQGAYQEAFDISKKEMQPTHPIRLGLALNFSVFYYEILNNPELACTLAKTAFDEAIAELDTLNEDSYKDSTLIMQLLRDNLTLWTSDSAGEECDAAEGAEN 1433z_human 14-3-3 protein zeta/delta YWHAZ Protein kinase C inhibitor protein 1 protein protein human Homo sapiens Human MDKNELVQKAKLAEQAERYDDMAACMKSVTEQGAELSNEERNLLSVAYKNVVGARRSSWRVVSSIEQKTEGAEKKQQMAREYREKIETELRDICNDVLSLLEKFLIPNASQAESKVFYLKMKGDYYRYLAEVAAGDDKKGIVDQSQQAYQEAFEISKKEMQPTHPIRLGLALNFSVFYYEILNSPEKACSLAKTAFDEAIAELDTLSEESYKDSTLIMQLLRDNLTLWTSDTQGDEAEAGEGGEN kapca_human cAMP-dependent protein kinase catalytic subunit alpha PRKACA PKACA protein protein human Homo sapiens Human MGNAAAAKKGSEQESVKEFLAKAKEDFLKKWESPAQNTAHLDQFERIKTLGTGSFGRVMLVKHKETGNHYAMKILDKQKVVKLKQIEHTLNEKRILQAVNFPFLVKLEFSFKDNSNLYMVMEYVPGGEMFSHLRRIGRFSEPHARFYAAQIVLTFEYLHSLDLIYRDLKPENLLIDQQGYIQVTDFGFAKRVKGRTWTLCGTPEYLAPEIILSKGYNKAVDWWALGVLIYEMAAGYPPFFADQPIQIYEKIVSGKVRFPSHFSSDLKDLLRNLLQVDLTKRFGNLKNGVNDIKNHKWFATTDWIAIYQRKVEAPFIPKFKGPGDTSNFDDYEEEEIRVSINEKCGKEFSEF moes_human Moesin Membrane-organizing extension spike protein MSN protein protein human Homo sapiens Human MPKTISVRVTTMDAELEFAIQPNTTGKQLFDQVVKTIGLREVWFFGLQYQDTKGFSTWLKLNKKVTAQDVRKESPLLFKFRAKFYPEDVSEELIQDITQRLFFLQVKEGILNDDIYCPPETAVLLASYAVQSKYGDFNKEVHKSGYLAGDKLLPQRVLEQHKLNKDQWEERIQVWHEEHRGMLREDAVLEYLKIAQDLEMYGVNYFSIKNKKGSELWLGVDALGLNIYEQNDRLTPKIGFPWSEIRNISFNDKKFVIKPIDKKAPDFVFYAPRLRINKRILALCMGNHELYMRRRKPDTIEVQQMKAQAREEKHQKQMERAMLENEKKKREMAEKEKEKIEREKEELMERLKQIEEQTKKAQQELEEQTRRALELEQERKRAQSEAEKLAKERQEAEEAKEALLQASRDQKKTQEQLALEMAELTARISQLEMARQKKESEAVEWQQKAQMVQEDLEKTRAELKTAMSTPHVAEPAENEQDEQDENGAEASADLRADAMAKDRSEEERTTEAEKNERVQKHLKALTSELANARDESKKTANDMIHAENMRLGRDKYKTLRQIRQGNTKQRIDEFESM lrrk2-ywhag-2 1475302 n/a 1475310 unspecified role unspecified role neutral component neutral component n_terminus gst tag glutathione s tranferase tag glutathione S-tranferase tag glutathione S-tranferase tag gst tag n-term range n-terminal range n-term range n-term range n-terminal range n-term range false ecoli Escherichia coli (strain K12) n/a 1475311 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 false n_terminus fluorescein fluorescein label fluorescein n-term range n-terminal range n-term range n-term range n-terminal range n-term range false required_phosphoser1444 opser O-phospho-L-serine (2S)-2-amino-3-(phosphonooxy)propanoic acid 2-amino-3-hydroxypropanoic acid 3-phosphate 2-azanyl-3-(phosphonooxy)propanoic acid ACT_SITE Phosphoserine intermediate MOD_RES Phosphoserine O-phospho-L-serine O-phosphonoserine O-phosphorylated L-serine O3-phosphoserine serine phosphate ester OPSer Phospho Phospho Seryl Phosphorylation phosphorylations certain certain sequence position certain certain certain sequence position certain false Q5S007:p.Arg1441Gly mutation disrupting mutation disrupting interaction mutation disrupting certain certain sequence position certain certain certain sequence position certain false Q5S007:p.Ser1444Ala mutation disrupting mutation disrupting interaction mutation disrupting certain certain sequence position certain certain certain sequence position certain false chemical synthesis Chemical synthesis (Chemical synthesis) direct interaction direct interaction false false false 1475302 Figure 4iii, Supp figure 2, Table 1. Phosphorylated LRRK2 WT peptide bound to all three 14-3-3 isoforms, indicated by a high FP signal, whereas the nonphosphorylated WT LRRK2 peptide did not bind to any of the 14-3-3 isoforms. lrrk2-ywhaq-1 1475302 n/a 1475311 unspecified role unspecified role neutral component neutral component n_terminus fluorescein fluorescein label fluorescein n-term range n-terminal range n-term range n-term range n-terminal range n-term range false Q5S007:p.Ser1444Ala mutation disrupting mutation disrupting interaction mutation disrupting certain certain sequence position certain certain certain sequence position certain false required_phosphoser1444 opser O-phospho-L-serine (2S)-2-amino-3-(phosphonooxy)propanoic acid 2-amino-3-hydroxypropanoic acid 3-phosphate 2-azanyl-3-(phosphonooxy)propanoic acid ACT_SITE Phosphoserine intermediate MOD_RES Phosphoserine O-phospho-L-serine O-phosphonoserine O-phosphorylated L-serine O3-phosphoserine serine phosphate ester OPSer Phospho Phospho Seryl Phosphorylation phosphorylations certain certain sequence position certain certain certain sequence position certain false Q5S007:p.Arg1441Gly mutation disrupting mutation disrupting interaction mutation disrupting certain certain sequence position certain certain certain sequence position certain false region sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain false chemical synthesis Chemical synthesis (Chemical synthesis) n/a 1475312 unspecified role unspecified role neutral component neutral component n_terminus gst tag glutathione s tranferase tag glutathione S-tranferase tag glutathione S-tranferase tag gst tag n-term range n-terminal range n-term range n-term range n-terminal range n-term range false ecoli Escherichia coli (strain K12) direct interaction direct interaction false false false 1475302 Figure 4ii, supp figure 2, Table 1 Phosphorylated LRRK2 WT peptide bound to all three 14-3-3 isoforms, indicated by a high FP signal, whereas the nonphosphorylated WT LRRK2 peptide did not bind to any of the 14-3-3 isoforms. lrrk2-ywhaz-1 1475302 n/a 1475311 unspecified role unspecified role neutral component neutral component Q5S007:p.Arg1441Gly mutation disrupting mutation disrupting interaction mutation disrupting certain certain sequence position certain certain certain sequence position certain false n_terminus fluorescein fluorescein label fluorescein n-term range n-terminal range n-term range n-term range n-terminal range n-term range false required_phosphoser1444 opser O-phospho-L-serine (2S)-2-amino-3-(phosphonooxy)propanoic acid 2-amino-3-hydroxypropanoic acid 3-phosphate 2-azanyl-3-(phosphonooxy)propanoic acid ACT_SITE Phosphoserine intermediate MOD_RES Phosphoserine O-phospho-L-serine O-phosphonoserine O-phosphorylated L-serine O3-phosphoserine serine phosphate ester OPSer Phospho Phospho Seryl Phosphorylation phosphorylations certain certain sequence position certain certain certain sequence position certain false Q5S007:p.Ser1444Ala mutation disrupting mutation disrupting interaction mutation disrupting certain certain sequence position certain certain certain sequence position certain false region sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain false chemical synthesis Chemical synthesis (Chemical synthesis) n/a 1475313 unspecified role unspecified role neutral component neutral component n_terminus gst tag glutathione s tranferase tag glutathione S-tranferase tag glutathione S-tranferase tag gst tag n-term range n-terminal range n-term range n-term range n-terminal range n-term range false ecoli Escherichia coli (strain K12) direct interaction direct interaction false false false 1475302 Figure 4iii, Table 1, Supp fig 2. Phosphorylated LRRK2 WT peptide bound to all three 14-3-3 isoforms, indicated by a high FP signal, whereas the nonphosphorylated WT LRRK2 peptide did not bind to any of the 14-3-3 isoforms. lrrk2-ywhag-3 1475302 n/a 1475310 unspecified role unspecified role neutral component neutral component n_terminus gst tag glutathione s tranferase tag glutathione S-tranferase tag glutathione S-tranferase tag gst tag n-term range n-terminal range n-term range n-term range n-terminal range n-term range false ecoli Escherichia coli (strain K12) n/a 1475311 unspecified role unspecified role neutral component neutral component required_phosphoser910 opser O-phospho-L-serine (2S)-2-amino-3-(phosphonooxy)propanoic acid 2-amino-3-hydroxypropanoic acid 3-phosphate 2-azanyl-3-(phosphonooxy)propanoic acid ACT_SITE Phosphoserine intermediate MOD_RES Phosphoserine O-phospho-L-serine O-phosphonoserine O-phosphorylated L-serine O3-phosphoserine serine phosphate ester OPSer Phospho Phospho Seryl Phosphorylation phosphorylations certain certain sequence position certain certain certain sequence position certain false region sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain false n_terminus fluorescein fluorescein label fluorescein n-term range n-terminal range n-term range n-term range n-terminal range n-term range false chemical synthesis Chemical synthesis (Chemical synthesis) direct interaction direct interaction false false false 1475302 Table 1, figure S2 Phosphorylated LRRK2 WT peptide bound to all three 14-3-3 isoforms, indicated by a high FP signal, whereas the nonphosphorylated WT LRRK2 peptide did not bind to any of the 14-3-3 isoforms. lrrk2-ywhaz-2 1475302 n/a 1475311 unspecified role unspecified role neutral component neutral component n_terminus fluorescein fluorescein label fluorescein n-term range n-terminal range n-term range n-term range n-terminal range n-term range false region sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain false required_phosphoser910 opser O-phospho-L-serine (2S)-2-amino-3-(phosphonooxy)propanoic acid 2-amino-3-hydroxypropanoic acid 3-phosphate 2-azanyl-3-(phosphonooxy)propanoic acid ACT_SITE Phosphoserine intermediate MOD_RES Phosphoserine O-phospho-L-serine O-phosphonoserine O-phosphorylated L-serine O3-phosphoserine serine phosphate ester OPSer Phospho Phospho Seryl Phosphorylation phosphorylations certain certain sequence position certain certain certain sequence position certain false chemical synthesis Chemical synthesis (Chemical synthesis) n/a 1475313 unspecified role unspecified role neutral component neutral component n_terminus gst tag glutathione s tranferase tag glutathione S-tranferase tag glutathione S-tranferase tag gst tag n-term range n-terminal range n-term range n-term range n-terminal range n-term range false ecoli Escherichia coli (strain K12) direct interaction direct interaction false false false 1475302 Table 1, figure S2 Phosphorylated LRRK2 WT peptide bound to all three 14-3-3 isoforms, indicated by a high FP signal, whereas the nonphosphorylated WT LRRK2 peptide did not bind to any of the 14-3-3 isoforms. lrrk2-ywhaq-2 1475302 n/a 1475311 unspecified role unspecified role neutral component neutral component n_terminus fluorescein fluorescein label fluorescein n-term range n-terminal range n-term range n-term range n-terminal range n-term range false required_phosphoser910 opser O-phospho-L-serine (2S)-2-amino-3-(phosphonooxy)propanoic acid 2-amino-3-hydroxypropanoic acid 3-phosphate 2-azanyl-3-(phosphonooxy)propanoic acid ACT_SITE Phosphoserine intermediate MOD_RES Phosphoserine O-phospho-L-serine O-phosphonoserine O-phosphorylated L-serine O3-phosphoserine serine phosphate ester OPSer Phospho Phospho Seryl Phosphorylation phosphorylations certain certain sequence position certain certain certain sequence position certain false region sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain false chemical synthesis Chemical synthesis (Chemical synthesis) n/a 1475312 unspecified role unspecified role neutral component neutral component n_terminus gst tag glutathione s tranferase tag glutathione S-tranferase tag glutathione S-tranferase tag gst tag n-term range n-terminal range n-term range n-term range n-terminal range n-term range false ecoli Escherichia coli (strain K12) direct interaction direct interaction false false false 1475302 Table 1, figure S2 Phosphorylated LRRK2 WT peptide bound to all three 14-3-3 isoforms, indicated by a high FP signal, whereas the nonphosphorylated WT LRRK2 peptide did not bind to any of the 14-3-3 isoforms. lrrk2-prkaca-1 1475303 n/a 1475314 enzyme enzyme unspecified role unspecified role n/a 1475311 enzyme target enzyme target substrate substrate unspecified role unspecified role Q5S007:p.Lys1906Met mutation mutation certain certain sequence position certain certain certain sequence position certain false c_terminus_strepii/flag_tag tandem tag tandem tag c-term range C-terminal range c-term range c-term range C-terminal range c-term range false spofr-sf_9 Spodoptera frugiperda insect cells Fall armyworm sf_9 insect cells Derived from pupal ovarian tissue of the fall armyworm Cloned from the parent cell line, IPLB-SF-21-AE (SF-21) derived from immature ovaries of fall armyworm Spodoptera frugiperda pupae cells are susceptible to Baculovirus infection phosphorylation phosphorylation reaction phosphorylation false false false Supp Table 1,2. Supp fig 1. Q5S007 thr-833 O-phospho-L-threonine; Q5S007 ser-850 O-phospho-L-serine; Q5S007 ser-858 O-phospho-L-serine; Q5S007 ser-860 O-phospho-L-serine; Q5S007 ser-908 O-phospho-L-serine; Q5S007 ser-910 O-phospho-L-serine; Q5S007 ser-933 O-phospho-L-serine; Q5S007 ser-935 O-phospho-L-serine; Q5S007 ser-954 O-phospho-L-serine; Q5S007 ser-955 O-phospho-L-serine Q5S007 ser-958 O-phospho-L-serine Q5S007 ser-962 O-phospho-L-serine These phosphorylations were found using only Full Length LRRK2. Six serine residues (S910, S955, S971, S973, S976, and S979) also were found in the negative controls (Table S2), indicating that LRRK2 already was phosphorylated to some extent. lrrk2-prkaca-2 1475303 n/a 1475314 enzyme enzyme unspecified role unspecified role n/a 1475311 enzyme target enzyme target substrate substrate unspecified role unspecified role region sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain false Q5S007:p.Lys1906Met mutation mutation certain certain sequence position certain certain certain sequence position certain false c_terminus_strepii/flag_tag tandem tag tandem tag c-term range C-terminal range c-term range c-term range C-terminal range c-term range false spofr-sf_9 Spodoptera frugiperda insect cells Fall armyworm sf_9 insect cells Derived from pupal ovarian tissue of the fall armyworm Cloned from the parent cell line, IPLB-SF-21-AE (SF-21) derived from immature ovaries of fall armyworm Spodoptera frugiperda pupae cells are susceptible to Baculovirus infection phosphorylation phosphorylation reaction phosphorylation false false false Supp Table 1,2. Supp fig 1. -i Q5S007 ser-971 O-phospho-L-serine; Q5S007 ser-973 O-phospho-L-serine; Q5S007 ser-976 O-phospho-L-serine; Q5S007 ser-1443 O-phospho-L-serine; Q5S007 ser-1444 O-phospho-L-serine These phosphorylations were found using both the Full Length LRRK2 AND the N-terminal LRRK2 deletion construct. Six serine residues (S910, S955, S971, S973, S976, and S979) also were found in the negative controls (Table S2), indicating that LRRK2 already was phosphorylated to some extent. lrrk2-prkaca-3 1475303 n/a 1475314 enzyme enzyme unspecified role unspecified role n/a 1475311 enzyme target enzyme target substrate substrate unspecified role unspecified role c_terminus_strepii/flag_tag tandem tag tandem tag c-term range C-terminal range c-term range c-term range C-terminal range c-term range false region required to bind necessary binding region required to bind certain certain sequence position certain certain certain sequence position certain false Q5S007:p.Lys1906Met mutation mutation certain certain sequence position certain certain certain sequence position certain false spofr-sf_9 Spodoptera frugiperda insect cells Fall armyworm sf_9 insect cells Derived from pupal ovarian tissue of the fall armyworm Cloned from the parent cell line, IPLB-SF-21-AE (SF-21) derived from immature ovaries of fall armyworm Spodoptera frugiperda pupae cells are susceptible to Baculovirus infection phosphorylation phosphorylation reaction phosphorylation false false false Supp Table 1,2. Supp fig 1. -ii Q5S007 ser-979 O-phospho-L-serine; Q5S007 ser-2166 O-phospho-L-serine; Q5S007 thr-1849 O-phospho-L-threonine These phosphorylations were found only using the N-terminal LRRK2 deletion construct, and not using the Full length molecule. Six serine residues (S910, S955, S971, S973, S976, and S979) also were found in the negative controls (Table S2), indicating that LRRK2 already was phosphorylated to some extent. lrrk2-prkaca-5 1475304 n/a 1475314 enzyme enzyme unspecified role unspecified role n/a 1475311 enzyme target enzyme target substrate substrate unspecified role unspecified role 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 false roc_domain sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain false Q5S007:p.Arg1441Cys mutation disrupting rate mutation disrupting interaction rate mutation disrupting rate certain certain sequence position certain certain certain sequence position certain false Q5S007:p.Arg1441His mutation disrupting rate mutation disrupting interaction rate mutation disrupting rate certain certain sequence position certain certain certain sequence position certain false Q5S007:p.Ser1444Ala mutation decreasing rate mutation decreasing interaction rate mutation decreasing rate certain certain sequence position certain certain certain sequence position certain false Q5S007:p.Ser1443_Ser1444delinsAlaAla mutation disrupting rate mutation disrupting interaction rate mutation disrupting rate certain certain sequence position certain certain certain sequence position certain false Q5S007:p.Arg1441Gly mutation disrupting rate mutation disrupting interaction rate mutation disrupting rate certain certain sequence position certain certain certain sequence position certain false ecoli Escherichia coli (strain K12) phosphorylation phosphorylation reaction phosphorylation false false false Figure 2 P10644 inhibits activity of P17612 lrrk2-prkaca-6 1475304 n/a 1475315 enzyme target enzyme target substrate substrate unspecified role unspecified role 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 false ecoli Escherichia coli (strain K12) n/a 1475311 enzyme enzyme unspecified role unspecified role c_terminus_strepii/flag_tag tandem tag tandem tag c-term range C-terminal range c-term range c-term range C-terminal range c-term range false spofr-sf_9 Spodoptera frugiperda insect cells Fall armyworm sf_9 insect cells Derived from pupal ovarian tissue of the fall armyworm Cloned from the parent cell line, IPLB-SF-21-AE (SF-21) derived from immature ovaries of fall armyworm Spodoptera frugiperda pupae cells are susceptible to Baculovirus infection phosphorylation phosphorylation reaction phosphorylation false false false Fig 6A P26038 phosres lrrk2-1 1475304 n/a 1475311 putative self putative self putative self putative self c_terminus_strepii/flag_tag tandem tag tandem tag c-term range C-terminal range c-term range c-term range C-terminal range c-term range false Q5S007:p.Ser1444Ala mutation increasing rate mutation increasing interaction rate mutation increasing rate certain certain sequence position certain certain certain sequence position certain false Q5S007:p.Gly2019Ser mutation increasing rate mutation increasing interaction rate mutation increasing rate certain certain sequence position certain certain certain sequence position certain false Q5S007:p.Arg1441His mutation increasing rate mutation increasing interaction rate mutation increasing rate certain certain sequence position certain certain certain sequence position certain false spofr-sf_9 Spodoptera frugiperda insect cells Fall armyworm sf_9 insect cells Derived from pupal ovarian tissue of the fall armyworm Cloned from the parent cell line, IPLB-SF-21-AE (SF-21) derived from immature ovaries of fall armyworm Spodoptera frugiperda pupae cells are susceptible to Baculovirus infection phosphorylation phosphorylation reaction phosphorylation false true false Fig 6 A Q5S007 phosres lrrk2-ywhag-1 1475305 n/a 1475311 unspecified role unspecified role bait bait region sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain false required_phosphoser1444 opser O-phospho-L-serine (2S)-2-amino-3-(phosphonooxy)propanoic acid 2-amino-3-hydroxypropanoic acid 3-phosphate 2-azanyl-3-(phosphonooxy)propanoic acid ACT_SITE Phosphoserine intermediate MOD_RES Phosphoserine O-phospho-L-serine O-phosphonoserine O-phosphorylated L-serine O3-phosphoserine serine phosphate ester OPSer Phospho Phospho Seryl Phosphorylation phosphorylations certain certain sequence position certain certain certain sequence position certain false n_terminus biotin tag biotin tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined false chemical synthesis Chemical synthesis (Chemical synthesis) n/a 1475310 unspecified role unspecified role prey prey n_terminus gst tag glutathione s tranferase tag glutathione S-tranferase tag glutathione S-tranferase tag gst tag n-term range n-terminal range n-term range n-term range n-terminal range n-term range false ecoli Escherichia coli (strain K12) direct interaction direct interaction false false false Figure 5 lrrk2-ywhaz 1475305 n/a 1475311 unspecified role unspecified role bait bait required_phosphoser1444 opser O-phospho-L-serine (2S)-2-amino-3-(phosphonooxy)propanoic acid 2-amino-3-hydroxypropanoic acid 3-phosphate 2-azanyl-3-(phosphonooxy)propanoic acid ACT_SITE Phosphoserine intermediate MOD_RES Phosphoserine O-phospho-L-serine O-phosphonoserine O-phosphorylated L-serine O3-phosphoserine serine phosphate ester OPSer Phospho Phospho Seryl Phosphorylation phosphorylations certain certain sequence position certain certain certain sequence position certain false region sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain false n_terminus biotin tag biotin tag n-term range n-terminal range n-term range n-term range n-terminal range n-term range false chemical synthesis Chemical synthesis (Chemical synthesis) n/a 1475313 unspecified role unspecified role prey prey n_terminus gst tag glutathione s tranferase tag glutathione S-tranferase tag glutathione S-tranferase tag gst tag n-term range n-terminal range n-term range n-term range n-terminal range n-term range false ecoli Escherichia coli (strain K12) direct interaction direct interaction false false false Figure 5 lrrk2-ywhaq 1475305 n/a 1475312 unspecified role unspecified role prey prey n_terminus gst tag glutathione s tranferase tag glutathione S-tranferase tag glutathione S-tranferase tag gst tag n-term range n-terminal range n-term range n-term range n-terminal range n-term range false ecoli Escherichia coli (strain K12) n/a 1475311 unspecified role unspecified role bait bait region sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain false required_phosphoser1444 opser O-phospho-L-serine (2S)-2-amino-3-(phosphonooxy)propanoic acid 2-amino-3-hydroxypropanoic acid 3-phosphate 2-azanyl-3-(phosphonooxy)propanoic acid ACT_SITE Phosphoserine intermediate MOD_RES Phosphoserine O-phospho-L-serine O-phosphonoserine O-phosphorylated L-serine O3-phosphoserine serine phosphate ester OPSer Phospho Phospho Seryl Phosphorylation phosphorylations certain certain sequence position certain certain certain sequence position certain false n_terminus biotin tag biotin tag n-term range n-terminal range n-term range n-term range n-terminal range n-term range false chemical synthesis Chemical synthesis (Chemical synthesis) direct interaction direct interaction false false false Figure 5 lrrk2-prkaca-4 1475306 n/a 1475314 enzyme enzyme unspecified role unspecified role n/a 1475311 enzyme target enzyme target substrate substrate unspecified role unspecified role c_terminus_strepii/flag_tag tandem tag tandem tag c-term range C-terminal range c-term range c-term range C-terminal range c-term range false spofr-sf_9 Spodoptera frugiperda insect cells Fall armyworm sf_9 insect cells Derived from pupal ovarian tissue of the fall armyworm Cloned from the parent cell line, IPLB-SF-21-AE (SF-21) derived from immature ovaries of fall armyworm Spodoptera frugiperda pupae cells are susceptible to Baculovirus infection phosphorylation phosphorylation reaction phosphorylation false false false 1A Q5S007 ser-910 O-phospho-L-serine; Q5S007 ser-935 O-phospho-L-serine LRRK2 already showed basal phosphorylation on both S910 and S935, which were increased significantly upon incubation with PKA. P17612 increases binding of Q5S007 to P61981 lrrk2-1 1475306 n/a 1475311 putative self putative self putative self putative self c_terminus_strepii/flag_tag tandem tag tandem tag c-term range C-terminal range c-term range c-term range C-terminal range c-term range false Q5S007:p.Lys1906Met mutation decreasing rate mutation decreasing interaction rate mutation decreasing rate certain certain sequence position certain certain certain sequence position certain false spofr-sf_9 Spodoptera frugiperda insect cells Fall armyworm sf_9 insect cells Derived from pupal ovarian tissue of the fall armyworm Cloned from the parent cell line, IPLB-SF-21-AE (SF-21) derived from immature ovaries of fall armyworm Spodoptera frugiperda pupae cells are susceptible to Baculovirus infection phosphorylation phosphorylation reaction phosphorylation false true false Fig 6 C, Supp fig3 Q5S007 thr-2483 O-phospho-L-threonine PKA together with GST–14-3-3 gamma ywhag-lrrk2-7 1475307 n/a 1475311 competitor competitor competitor prey prey region sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain false required_phosphoser1444 opser O-phospho-L-serine (2S)-2-amino-3-(phosphonooxy)propanoic acid 2-amino-3-hydroxypropanoic acid 3-phosphate 2-azanyl-3-(phosphonooxy)propanoic acid ACT_SITE Phosphoserine intermediate MOD_RES Phosphoserine O-phospho-L-serine O-phosphonoserine O-phosphorylated L-serine O3-phosphoserine serine phosphate ester OPSer Phospho Phospho Seryl Phosphorylation phosphorylations certain certain sequence position certain certain certain sequence position certain false chemical synthesis Chemical synthesis (Chemical synthesis) n/a 1475310 unspecified role unspecified role bait bait n_terminus gst tag glutathione s tranferase tag glutathione S-tranferase tag glutathione S-tranferase tag gst tag n-term range n-terminal range n-term range n-term range n-terminal range n-term range false ecoli Escherichia coli (strain K12) n/a 1475311 unspecified role unspecified role prey prey c_terminus_strepii/flag_tag tandem tag tandem tag c-term range C-terminal range c-term range c-term range C-terminal range c-term range false region sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain false spofr-sf_9 Spodoptera frugiperda insect cells Fall armyworm sf_9 insect cells Derived from pupal ovarian tissue of the fall armyworm Cloned from the parent cell line, IPLB-SF-21-AE (SF-21) derived from immature ovaries of fall armyworm Spodoptera frugiperda pupae cells are susceptible to Baculovirus infection physical association physical association false false false Figure 3ii C LRRK2 WT Delta-967, which does not contain S910/S935, was precipitated efficiently by GST–14-3-3 gamma, strongly indicating an additional 14-3-3 docking site in LRRK2. ywhag-lrrk2-8 1475307 n/a 1475311 unspecified role unspecified role prey prey region sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain false c_terminus_strepii/flag_tag tandem tag tandem tag c-term range C-terminal range c-term range c-term range C-terminal range c-term range false spofr-sf_9 Spodoptera frugiperda insect cells Fall armyworm sf_9 insect cells Derived from pupal ovarian tissue of the fall armyworm Cloned from the parent cell line, IPLB-SF-21-AE (SF-21) derived from immature ovaries of fall armyworm Spodoptera frugiperda pupae cells are susceptible to Baculovirus infection n/a 1475310 unspecified role unspecified role bait bait n_terminus gst tag glutathione s tranferase tag glutathione S-tranferase tag glutathione S-tranferase tag gst tag n-term range n-terminal range n-term range n-term range n-terminal range n-term range false ecoli Escherichia coli (strain K12) physical association physical association false false false Figure 3B/C LRRK2 WT Delta-967, which does not contain S910/S935, was precipitated efficiently by GST–14-3-3 gamma, strongly indicating an additional 14-3-3 docking site in LRRK2. prkaca-lrrk2 1475308 n/a 1475311 unspecified role unspecified role prey prey 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 false Q5S007:p.Ser1443_Ser1444delinsAlaAla mutation disrupting mutation disrupting interaction mutation disrupting certain certain sequence position certain certain certain sequence position certain false Q5S007:p.Ser1444Ala mutation disrupting mutation disrupting interaction mutation disrupting certain certain sequence position certain certain certain sequence position certain false roc_domain sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain false required_phosphoser1444 opser O-phospho-L-serine (2S)-2-amino-3-(phosphonooxy)propanoic acid 2-amino-3-hydroxypropanoic acid 3-phosphate 2-azanyl-3-(phosphonooxy)propanoic acid ACT_SITE Phosphoserine intermediate MOD_RES Phosphoserine O-phospho-L-serine O-phosphonoserine O-phosphorylated L-serine O3-phosphoserine serine phosphate ester OPSer Phospho Phospho Seryl Phosphorylation phosphorylations certain certain sequence position certain certain certain sequence position certain false ecoli Escherichia coli (strain K12) n/a 1475310 unspecified role unspecified role bait bait n_terminus gst tag glutathione s tranferase tag glutathione S-tranferase tag glutathione S-tranferase tag gst tag n-term range n-terminal range n-term range n-term range n-terminal range n-term range false ecoli Escherichia coli (strain K12) direct interaction direct interaction false false false Figure 3D Binding of 14-3-3 to LRRK2-ROC was detected exclusively to PKA-phosphorylated but not to nonphosphorylated ROC fusion protein, and could be observed only when S1444 was present. lrrk2-1 1475309 n/a 1475311 putative self putative self putative self putative self Q5S007:p.Ser1444Ala mutation increasing rate mutation increasing interaction rate mutation increasing rate certain certain sequence position certain certain certain sequence position certain false Q5S007:p.Gly2019Ser mutation increasing rate mutation increasing interaction rate mutation increasing rate certain certain sequence position certain certain certain sequence position certain false c_terminus_strepii/flag_tag tandem tag tandem tag c-term range C-terminal range c-term range c-term range C-terminal range c-term range false Q5S007:p.Arg1441His mutation increasing rate mutation increasing interaction rate mutation increasing rate certain certain sequence position certain certain certain sequence position certain false spofr-sf_9 Spodoptera frugiperda insect cells Fall armyworm sf_9 insect cells Derived from pupal ovarian tissue of the fall armyworm Cloned from the parent cell line, IPLB-SF-21-AE (SF-21) derived from immature ovaries of fall armyworm Spodoptera frugiperda pupae cells are susceptible to Baculovirus infection phosphorylation phosphorylation reaction phosphorylation false true false Fig 6 B Q5S007 phosres