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/ klostermann-2002-3 The thylakoid membrane protein ALB3 associates with the cpSecY-translocase in Arabidopsis thaliana. arath Arabidopsis thaliana (Mouse-ear cress) Mouse-ear cress anti bait coip anti bait coimmunoprecipitation anti bait coip western blot western blot Immuno blot Immuno blot Klostermann E., Droste Gen Helling I., Carde JP., Schunemann D. Thylakoid membranes were prepared from Arabidopsis thaliana (ecotype Columbia) seedlings and solubilized with 1% digitonin prior to blue native gel electrophoresis. schuenemann@bio3.rwth.aachen.de Biochem. J. (0264-6021) 2002 klostermann-2002-1 The thylakoid membrane protein ALB3 associates with the cpSecY-translocase in Arabidopsis thaliana. arath Arabidopsis thaliana (Mouse-ear cress) Mouse-ear cress molecular sieving molecular sieving Gel Filtration Size Exclusion Chromatography Size Exclusion Chromatography Gel Filtration Sizing column western blot western blot Immuno blot Immuno blot Klostermann E., Droste Gen Helling I., Carde JP., Schunemann D. Thylakoid membranes were prepared from Arabidopsis thaliana (ecotype Columbia) seedlings and solubilized with 1% digitonin prior to blue native gel electrophoresis. schuenemann@bio3.rwth.aachen.de Biochem. J. (0264-6021) 2002 klostermann-2002-2 The thylakoid membrane protein ALB3 associates with the cpSecY-translocase in Arabidopsis thaliana. arath Arabidopsis thaliana (Mouse-ear cress) Mouse-ear cress bn-page blue native page bn-page western blot western blot Immuno blot Immuno blot Klostermann E., Droste Gen Helling I., Carde JP., Schunemann D. Thylakoid membranes were prepared from Arabidopsis thaliana (ecotype Columbia) seedlings and solubilized with 1% digitonin prior to blue native gel electrophoresis. schuenemann@bio3.rwth.aachen.de Biochem. J. (0264-6021) 2002 klostermann-2002-4 The thylakoid membrane protein ALB3 associates with the cpSecY-translocase in Arabidopsis thaliana. arath-leaf Arabidopsis thaliana leaf leaf leaf electron microscopy electron microscopy Electron crystallography Electron cryomicroscopy Electron crystallography Electron cryomicroscopy immunostaining immunostaining Klostermann E., Droste Gen Helling I., Carde JP., Schunemann D. Thylakoid membranes were prepared from Arabidopsis thaliana (ecotype Columbia) seedlings and solubilized with 1% digitonin prior to blue native gel electrophoresis. schuenemann@bio3.rwth.aachen.de Biochem. J. (0264-6021) 2002 klostermann-2002-5 The thylakoid membrane protein ALB3 associates with the cpSecY-translocase in Arabidopsis thaliana. arath Arabidopsis thaliana (Mouse-ear cress) Mouse-ear cress crosslink cross-linking study crosslink western blot western blot Immuno blot Immuno blot Klostermann E., Droste Gen Helling I., Carde JP., Schunemann D. Thylakoid membranes were prepared from Arabidopsis thaliana (ecotype Columbia) seedlings and solubilized with 1% digitonin prior to blue native gel electrophoresis. schuenemann@bio3.rwth.aachen.de Biochem. J. (0264-6021) 2002 scy1_arath Preprotein translocase subunit SCY1, chloroplastic SCY1 CpSecY SECY MSF3.9 At2g18710 protein protein arath Arabidopsis thaliana (Mouse-ear cress) Mouse-ear cress MITVSEVSSYSSSSSNFASLSRLNHKSSSRLRSSSLYKGSFFSVSTKTRRNTCKAKSWNLGLVINSRSSEASVFDPLGINPDETSGLSSIWESFVSLLSPSFESSSGNRRDKPSSGRGVAAAIEDSSIDFGDFFKGPLPGKFLKLLGFLALSRLGIYIPLGGVNREAFVGNLDQNSILSTLDTFSGGGIGRLGICSLGIVPFINAQIVFQLLAQVYPKLQDLQKKEGEAGRKKILQYTRYASVGFAIVQAIGQVFYLRPYVNDFSTEWVVSSVTLLTLGSVLTTYIGERISDLKLGNGTSLLIFTSIISYLPASFGRTTAEALQEGNYTGLGTIVVSFLLLVLGIVYVQEAERKIPLNYASRYTSKAGGLQKSAYLPFKVNSAGVMPIIFSTSSLALPATLARFTGISALKNVAFALTPGGSFYLPTNILLIAFFNYYYTFLQLDPDDVSEQLKRQGASIPLVRPGKSTALFIKTVLGRISVLGSAFLAVLAAGPAVVEQITHLTAFRGFAGTSVLILVGCATDTARKVQAEIISQKYKNIEFYELDKYDP alb3_arath Inner membrane protein ALBINO3, chloroplastic ALB3 F8N16.9 At2g28800 protein protein arath Arabidopsis thaliana (Mouse-ear cress) Mouse-ear cress MARVLVSSPSSFFGSPLIKPSSSLRHSGVGGGGTAQFLPYRSNNNKLFTTSTTVRFSLNEIPPFHGLDSSVDIGAIFTRAESLLYTIADAAVVGADSVVTTDSSAVQKSGGWFGFISDAMELVLKILKDGLSAVHVPYAYGFAIILLTIIVKAATYPLTKQQVESTLAMQNLQPKIKAIQQRYAGNQERIQLETSRLYKQAGVNPLAGCLPTLATIPVWIGLYQALSNVANEGLFTEGFFWIPSLGGPTSIAARQSGSGISWLFPFVDGHPPLGWYDTVAYLVLPVLLIASQYVSMEIMKPPQTDDPAQKNTLLVFKFLPLMIGYFALSVPSGLSIYWLTNNVLSTAQQVYLRKLGGAKPNMDENASKIISAGRAKRSIAQPDDAGERFRQLKEQEKRSKKNKAVAKDTVELVEESQSESEEGSDDEEEEAREGALASSTTSKPLPEVGQRRSKRSKRKRTV secy-alb3-3 763391 N/A 763396 unspecified role unspecified role prey prey N/A 763397 unspecified role unspecified role bait bait physical association physical association false false false Figure 3 alb3-secy 763391 N/A-x-x 763396 unspecified role unspecified role bait bait N/A-x-x 763397 unspecified role unspecified role prey prey physical association physical association false false false Figure 3 secy-alb3-1 763392 N/A 763397 unspecified role unspecified role neutral component neutral component N/A 763396 unspecified role unspecified role neutral component neutral component physical association physical association false false false Figure 1 Note that ALB3 and cpSecY eluted from the gel filtration column as higher-molecularmass species of approx. 200 kDa when thylakoids were solubilized using 1.5% digitonin. ALB3 and cpSecY eluted from the gel filtration column as higher-molecular mass species of approx. 180 kDa when the thylakoid membranes were solubilized under harsher conditions using dodecyl b-dmaltosideas detergent secy-alb3-2 763393 N/A-x-x 763396 unspecified role unspecified role neutral component neutral component N/A-x-x 763397 unspecified role unspecified role neutral component neutral component physical association physical association false false false Figure 2 Note that ALB3 and cpSecY were found to be components of a higher-molecular-mass complex of approx. 160 kDa, and no monomeric forms of ALB3 or cpSecY were detected. However, blue native gel electrophoresis revealed that ALB3 and cpSecY were also present in a variety of oligomeric complexes with molecular masses ranging from 120 to 700 kDa. secy-alb3-4 763394 N/A-x 763397 unspecified role unspecified role neutral component neutral component N/A-x 763396 unspecified role unspecified role neutral component neutral component colocalization colocalization false false false Figure 4 Note that almost all observed gold particles corresponding to cpSecY (10 nm) co-localized with the gold particles corresponding to ALB3 (5 nm), whereas a signi�cant number of free gold particles for ALB3 were detected. These observations provide further evidence that at least a portion of ALB3 interacts with the cpSecY complex. secy-alb3-5 763395 N/A-x 763396 unspecified role unspecified role neutral component neutral component N/A-x 763397 unspecified role unspecified role neutral component neutral component direct interaction direct interaction false false false Figure 5