DIP Database of Interacting Proteins 611 Young Drive East; Los Angeles CA 90095; USA http://dip.doe-mbi.ucla.edu/ dip@mbi.ucla.edu 611 Young Drive East; Los Angeles CA 90095; USA http://dip.doe-mbi.ucla.edu/ zegzouti-2006a-1 Phosphorylation and activation of PINOID by the phospholipid signaling kinase 3-phosphoinositide-dependent protein kinase 1 (PDK1) in Arabidopsis. in vitro In vitro protein kinase assay protein kinase assay weight autoradiogra molecular weight estimation by autoradiography weight autoradiogra Zegzouti H., Anthony RG., Jahchan N., Bogre L., Christensen SK. sioux@ucla.edu Proc. Natl. Acad. Sci. U.S.A. (0027-8424) 2006 IMEx Only protein-protein interactions imex curation zegzouti-2006a-2 Phosphorylation and activation of PINOID by the phospholipid signaling kinase 3-phosphoinositide-dependent protein kinase 1 (PDK1) in Arabidopsis. in vitro In vitro pull down pull down western blot western blot Immuno blot Immuno blot Zegzouti H., Anthony RG., Jahchan N., Bogre L., Christensen SK. sioux@ucla.edu Proc. Natl. Acad. Sci. U.S.A. (0027-8424) 2006 IMEx Only protein-protein interactions imex curation pid_arath Protein kinase PINOID Protein kinase ABRUPTUS T31E10.1 PID ABR At2g34650 protein protein arath Arabidopsis thaliana (Mouse-ear cress) Mouse-ear cress MLRESDGEMSLGTTNSPISSGTESCSSFSRLSFDAPPSTIPEEESFLSLKPHRSSDFAYAEIRRRKKQGLTFRDFRLMRRIGAGDIGTVYLCRLAGDEEESRSSYFAMKVVDKEALALKKKMHRAEMEKTILKMLDHPFLPTLYAEFEASHFSCIVMEYCSGGDLHSLRHRQPHRRFSLSSARFYAAEVLVALEYLHMLGIIYRDLKPENILVRSDGHIMLSDFDLSLCSDSIAAVESSSSSPENQQLRSPRRFTRLARLFQRVLRSKKVQTLEPTRLFVAEPVTARSGSFVGTHEYVAPEVASGGSHGNAVDWWAFGVFLYEMIYGKTPFVAPTNDVILRNIVKRQLSFPTDSPATMFELHARNLISGLLNKDPTKRLGSRRGAAEVKVHPFFKGLNFALIRTLTPPEIPSSVVKKPMKSATFSGRSSNKPAAFDYF pdpk1_arath 3-phosphoinositide-dependent protein kinase 1 PDPK1 PDK1 T32M21.110 At5g04510 protein protein arath Arabidopsis thaliana (Mouse-ear cress) Mouse-ear cress MLAMEKEFDSKLVLQGNSSNGANVSRSKSFSFKAPQENFTSHDFEFGKIYGVGSYSKVVRAKKKETGTVYALKIMDKKFITKENKTAYVKLERIVLDQLEHPGIIKLYFTFQDTSSLYMALESCEGGELFDQITRKGRLSEDEARFYTAEVVDALEYIHSMGLIHRDIKPENLLLTSDGHIKIADFGSVKPMQDSQITVLPNAASDDKACTFVGTAAYVPPEVLNSSPATFGNDLWALGCTLYQMLSGTSPFKDASEWLIFQRIIARDIKFPNHFSEAARDLIDRLLDTEPSRRPGAGSEGYVALKRHPFFNGVDWKNLRSQTPPKLAPDPASQTASPERDDTHGSPWNLTHIGDSLATQNEGHSAPPTSSESSGSITRLASIDSFDSRWQQFLEPGESVLMISAVKKLQKITSKKVQLILTNKPKLIYVDPSKLVVKGNIIWSDNSNDLNVVVTSPSHFKICTPKKVLSFEDAKQRASVWKKAIETLQNR pid-3 Kinase assay for Arabidopsis PID protein 672011 N/A 672013 putative self putative self putative self putative self O64682:p.[Ser288Glu;Ser290Glu] mutation increasing mutation increasing interaction mutation increasing certain certain sequence position certain certain certain sequence position certain false certain certain sequence position certain certain certain sequence position certain false 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 O64682:p.Asp205Ala mutation disrupting mutation disrupting interaction mutation disrupting certain certain sequence position certain certain certain sequence position certain false ecobb Escherichia coli (strain B / BL21) phosphorylation phosphorylation reaction phosphorylation false true false 1C 1D 3C autophosphorylation of PID protein. Increasing levels of Magnesium ions stimulated autophosphorylation of PID Calcium ions The authors concluded that "These data suggest that Ca ions may inhibit PID activity by competitive binding and displacement of the Mg2 ions". pdk1-pid-1 PDK1 phosphorylates the PID protein 672011 N/A 672013 enzyme target enzyme target substrate substrate neutral component neutral component 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 O64682:p.[Ser288Glu;Ser290Glu] mutation disrupting rate mutation disrupting interaction rate mutation disrupting rate certain certain sequence position certain certain certain sequence position certain false certain certain sequence position certain certain certain sequence position certain false O64682:p.[Phe435Val;Phe438Val] mutation disrupting rate mutation disrupting interaction rate mutation disrupting rate certain certain sequence position certain certain certain sequence position certain false certain certain sequence position certain certain certain sequence position certain false ecobb Escherichia coli (strain B / BL21) n/a 672014 enzyme enzyme neutral component neutral component 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 ecobb Escherichia coli (strain B / BL21) phosphorylation phosphorylation reaction phosphorylation false false false 3B, 3C, 4B, Supplementary information Figure 8 The authors concluded that "transphosphorylation by PDK1 activates PID through up-regulation of autophosphorylation activity". In addition that "PDK1 activates PID by phosphorylating one or more of the serines within the PID activation loop". phosphorylation of PID Time course experiments show that the increase in PDK1-dependent PID autophosphorylation is rapid, with peak levels of activation occurring after 30 min of coincubation pdk1-pid-2 PDK1 phosphorylates the PID protein 672011 N/A 672013 enzyme target enzyme target substrate substrate neutral component neutral component 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 arath-protoplast Arabidopsis thaliana protoplast Protoplast defined as a membrane-bound cell from which the outer wall has been partially or completely removed. n/a 672014 enzyme enzyme neutral component neutral component 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 arath-protoplast Arabidopsis thaliana protoplast Protoplast defined as a membrane-bound cell from which the outer wall has been partially or completely removed. phosphorylation phosphorylation reaction phosphorylation false false false 5 phosphorylation of PID pdk1-pid-3 Interaction between PDK1 and PID protein 672012 n/a 672014 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 ecobb Escherichia coli (strain B / BL21) N/A 672013 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 false ecobb Escherichia coli (strain B / BL21) physical association physical association false false false 3A