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/ Encoding and transducing the synaptic or extrasynaptic origin of NMDA receptor signals to the nucleus. Encoding and transducing the synaptic or extrasynaptic origin of NMDA receptor signals to the nucleus. Cell (0092-8674) 2013 imex curation Karpova A., Mikhaylova M., Bera S., Baer J., Reddy P.P., Behnisch T., Rankovic V., Spilker C., Bethge P., Sahin J., Kaushik R., Zuschratter W., Kahne T., Naumann M., Gundelfinger E.D., Kreutz M.R. Interactions involving the MBP-Jacob construct could not be captured as it was not clear if this was rat or murine in origin. Origin of Calpain was not given. Interactions involving Homer, MEK1/2 or dynein Abs were not captured as these could not be ascribed to a single gene product. kreutz@lin-magdeburg.de 30-Sept-2013: Contacted by IntAct-Help. 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 predetermined predetermined participant predetermined Karpova A., Mikhaylova M., Bera S., Baer J., Reddy P.P., Behnisch T., Rankovic V., Spilker C., Bethge P., Sahin J., Kaushik R., Zuschratter W., Kahne T., Naumann M., Gundelfinger E.D., Kreutz M.R. Described as a screen but looks more like a matrix, so I've done it as predetermined. Interactions involving the MBP-Jacob construct could not be captured as it was not clear if this was rat or murine in origin. Origin of Calpain was not given. Interactions involving Homer, MEK1/2 or dynein Abs were not captured as these could not be ascribed to a single gene product. kreutz@lin-magdeburg.de Accepted 2013-SEP-19 by MGT 30-Sept-2013: Contacted by IntAct-Help. Cell (0092-8674) 2013 Only protein-protein interactions imex curation Encoding and transducing the synaptic or extrasynaptic origin of NMDA receptor signals to the nucleus. Encoding and transducing the synaptic or extrasynaptic origin of NMDA receptor signals to the nucleus. Cell (0092-8674) 2013 imex curation Karpova A., Mikhaylova M., Bera S., Baer J., Reddy P.P., Behnisch T., Rankovic V., Spilker C., Bethge P., Sahin J., Kaushik R., Zuschratter W., Kahne T., Naumann M., Gundelfinger E.D., Kreutz M.R. Interactions involving the MBP-Jacob construct could not be captured as it was not clear if this was rat or murine in origin. Origin of Calpain was not given. Interactions involving Homer, MEK1/2 or dynein Abs were not captured as these could not be ascribed to a single gene product. kreutz@lin-magdeburg.de 30-Sept-2013: Contacted by IntAct-Help. Only protein-protein interactions in vitro In vitro spr surface plasmon resonance Optical biosensor BIAcore(r) BIAcore(r) Optical biosensor spr predetermined predetermined participant predetermined Karpova A., Mikhaylova M., Bera S., Baer J., Reddy P.P., Behnisch T., Rankovic V., Spilker C., Bethge P., Sahin J., Kaushik R., Zuschratter W., Kahne T., Naumann M., Gundelfinger E.D., Kreutz M.R. Interactions involving the MBP-Jacob construct could not be captured as it was not clear if this was rat or murine in origin. Origin of Calpain was not given. Interactions involving Homer, MEK1/2 or dynein Abs were not captured as these could not be ascribed to a single gene product. kreutz@lin-magdeburg.de Accepted 2013-SEP-19 by MGT 30-Sept-2013: Contacted by IntAct-Help. Cell (0092-8674) 2013 Only protein-protein interactions imex curation Encoding and transducing the synaptic or extrasynaptic origin of NMDA receptor signals to the nucleus. Encoding and transducing the synaptic or extrasynaptic origin of NMDA receptor signals to the nucleus. Cell (0092-8674) 2013 imex curation Karpova A., Mikhaylova M., Bera S., Baer J., Reddy P.P., Behnisch T., Rankovic V., Spilker C., Bethge P., Sahin J., Kaushik R., Zuschratter W., Kahne T., Naumann M., Gundelfinger E.D., Kreutz M.R. Interactions involving the MBP-Jacob construct could not be captured as it was not clear if this was rat or murine in origin. Origin of Calpain was not given. Interactions involving Homer, MEK1/2 or dynein Abs were not captured as these could not be ascribed to a single gene product. kreutz@lin-magdeburg.de 30-Sept-2013: Contacted by IntAct-Help. Only protein-protein interactions in vitro In vitro pull down pull down western blot western blot Immuno blot Immuno blot Karpova A., Mikhaylova M., Bera S., Baer J., Reddy P.P., Behnisch T., Rankovic V., Spilker C., Bethge P., Sahin J., Kaushik R., Zuschratter W., Kahne T., Naumann M., Gundelfinger E.D., Kreutz M.R. Interactions involving the MBP-Jacob construct could not be captured as it was not clear if this was rat or murine in origin. Origin of Calpain was not given. Interactions involving Homer, MEK1/2 or dynein Abs were not captured as these could not be ascribed to a single gene product. kreutz@lin-magdeburg.de Accepted 2013-SEP-19 by MGT 30-Sept-2013: Contacted by IntAct-Help. Cell (0092-8674) 2013 Only protein-protein interactions imex curation Encoding and transducing the synaptic or extrasynaptic origin of NMDA receptor signals to the nucleus. Encoding and transducing the synaptic or extrasynaptic origin of NMDA receptor signals to the nucleus. Cell (0092-8674) 2013 imex curation Karpova A., Mikhaylova M., Bera S., Baer J., Reddy P.P., Behnisch T., Rankovic V., Spilker C., Bethge P., Sahin J., Kaushik R., Zuschratter W., Kahne T., Naumann M., Gundelfinger E.D., Kreutz M.R. Interactions involving the MBP-Jacob construct could not be captured as it was not clear if this was rat or murine in origin. Origin of Calpain was not given. Interactions involving Homer, MEK1/2 or dynein Abs were not captured as these could not be ascribed to a single gene product. kreutz@lin-magdeburg.de 30-Sept-2013: Contacted by IntAct-Help. Only protein-protein interactions in vitro In vitro protein kinase assay protein kinase assay ms participant Identification by mass spectrometry ms participant Karpova A., Mikhaylova M., Bera S., Baer J., Reddy P.P., Behnisch T., Rankovic V., Spilker C., Bethge P., Sahin J., Kaushik R., Zuschratter W., Kahne T., Naumann M., Gundelfinger E.D., Kreutz M.R. Interactions involving the MBP-Jacob construct could not be captured as it was not clear if this was rat or murine in origin. Origin of Calpain was not given. Interactions involving Homer, MEK1/2 or dynein Abs were not captured as these could not be ascribed to a single gene product. kreutz@lin-magdeburg.de Accepted 2013-SEP-19 by MGT 30-Sept-2013: Contacted by IntAct-Help. Cell (0092-8674) 2013 Only protein-protein interactions imex curation Encoding and transducing the synaptic or extrasynaptic origin of NMDA receptor signals to the nucleus. Encoding and transducing the synaptic or extrasynaptic origin of NMDA receptor signals to the nucleus. Cell (0092-8674) 2013 imex curation Karpova A., Mikhaylova M., Bera S., Baer J., Reddy P.P., Behnisch T., Rankovic V., Spilker C., Bethge P., Sahin J., Kaushik R., Zuschratter W., Kahne T., Naumann M., Gundelfinger E.D., Kreutz M.R. Interactions involving the MBP-Jacob construct could not be captured as it was not clear if this was rat or murine in origin. Origin of Calpain was not given. Interactions involving Homer, MEK1/2 or dynein Abs were not captured as these could not be ascribed to a single gene product. kreutz@lin-magdeburg.de 30-Sept-2013: Contacted by IntAct-Help. Only protein-protein interactions in vitro In vitro pull down pull down anti tag western anti tag western blot anti tag western Karpova A., Mikhaylova M., Bera S., Baer J., Reddy P.P., Behnisch T., Rankovic V., Spilker C., Bethge P., Sahin J., Kaushik R., Zuschratter W., Kahne T., Naumann M., Gundelfinger E.D., Kreutz M.R. Interactions involving the MBP-Jacob construct could not be captured as it was not clear if this was rat or murine in origin. Origin of Calpain was not given. Interactions involving Homer, MEK1/2 or dynein Abs were not captured as these could not be ascribed to a single gene product. kreutz@lin-magdeburg.de Accepted 2013-SEP-19 by MGT 30-Sept-2013: Contacted by IntAct-Help. Cell (0092-8674) 2013 Only protein-protein interactions imex curation Encoding and transducing the synaptic or extrasynaptic origin of NMDA receptor signals to the nucleus. Encoding and transducing the synaptic or extrasynaptic origin of NMDA receptor signals to the nucleus. Cell (0092-8674) 2013 imex curation Karpova A., Mikhaylova M., Bera S., Baer J., Reddy P.P., Behnisch T., Rankovic V., Spilker C., Bethge P., Sahin J., Kaushik R., Zuschratter W., Kahne T., Naumann M., Gundelfinger E.D., Kreutz M.R. Interactions involving the MBP-Jacob construct could not be captured as it was not clear if this was rat or murine in origin. Origin of Calpain was not given. Interactions involving Homer, MEK1/2 or dynein Abs were not captured as these could not be ascribed to a single gene product. kreutz@lin-magdeburg.de 30-Sept-2013: Contacted by IntAct-Help. Only protein-protein interactions rat-brain Rattus norvegicus brain brain Brain [cerebrum] anti bait coip anti bait coimmunoprecipitation anti bait coip western blot western blot Immuno blot Immuno blot Karpova A., Mikhaylova M., Bera S., Baer J., Reddy P.P., Behnisch T., Rankovic V., Spilker C., Bethge P., Sahin J., Kaushik R., Zuschratter W., Kahne T., Naumann M., Gundelfinger E.D., Kreutz M.R. Immunoprecipitate from cytosolic nonvesicular fraction of rat brain. Interactions involving the MBP-Jacob construct could not be captured as it was not clear if this was rat or murine in origin. Origin of Calpain was not given. Interactions involving Homer, MEK1/2 or dynein Abs were not captured as these could not be ascribed to a single gene product. kreutz@lin-magdeburg.de Accepted 2013-SEP-19 by MGT 30-Sept-2013: Contacted by IntAct-Help. Cell (0092-8674) 2013 Only protein-protein interactions imex curation Encoding and transducing the synaptic or extrasynaptic origin of NMDA receptor signals to the nucleus. Encoding and transducing the synaptic or extrasynaptic origin of NMDA receptor signals to the nucleus. Cell (0092-8674) 2013 imex curation Karpova A., Mikhaylova M., Bera S., Baer J., Reddy P.P., Behnisch T., Rankovic V., Spilker C., Bethge P., Sahin J., Kaushik R., Zuschratter W., Kahne T., Naumann M., Gundelfinger E.D., Kreutz M.R. Interactions involving the MBP-Jacob construct could not be captured as it was not clear if this was rat or murine in origin. Origin of Calpain was not given. Interactions involving Homer, MEK1/2 or dynein Abs were not captured as these could not be ascribed to a single gene product. kreutz@lin-magdeburg.de 30-Sept-2013: Contacted by IntAct-Help. Only protein-protein interactions in vitro In vitro pull down pull down western blot western blot Immuno blot Immuno blot Karpova A., Mikhaylova M., Bera S., Baer J., Reddy P.P., Behnisch T., Rankovic V., Spilker C., Bethge P., Sahin J., Kaushik R., Zuschratter W., Kahne T., Naumann M., Gundelfinger E.D., Kreutz M.R. Interactions involving the MBP-Jacob construct could not be captured as it was not clear if this was rat or murine in origin. Origin of Calpain was not given. Interactions involving Homer, MEK1/2 or dynein Abs were not captured as these could not be ascribed to a single gene product. kreutz@lin-magdeburg.de Accepted 2013-SEP-19 by MGT 30-Sept-2013: Contacted by IntAct-Help. Cell (0092-8674) 2013 Only protein-protein interactions imex curation ima5_rat Importin subunit alpha-5 Kpna1 Importin alpha-5 Karyopherin subunit alpha-1 protein protein rat Rattus norvegicus (Rat) Rat MSTPGKENFRLKSYKNKSLNPDEMRRRREEEGLQLRKQKREEQLFKRRNVATAEEETEEEVMSDGGFHEAQINNMEMAPGGVITSDMTEMIFSNSPEQQLSATQKFRKLLSKEPNPPIDEVINTPGVVARFVEFLKRKENCTLQFESAWVLTNIASGNSLQTRIVIQAGAVPIFIELLSSEFEDVQEQAVWALGNIAGDSTMCRDYVLDCNILPPLLQLFSKQNRLTMTRNAVWALSNLCRGKSPPPEFAKVSPCLNVLSWLLFVSDTDVLADACWALSYLSDGPNDKIQAVIDAGVCRRLVELLMHNDYKVVSPALRAVGNIVTGDDIQTQVILNCSALQSLLHLLSSPKESIKKEACWTISNITAGNRAQIQTVIDANMFPALISILQTAEFRTRKEAAWAITNATSGGSAEQIKYLVELGCIKPLCDLLTVMDAKIVQVALNGLENILRLGEQEAKRNGSGINPYCALIEEAYGLDKIEFLQSHENQEIYQKAFDLIEHYFGTEDEDSSIAPQVDLSQQQYIFQQCEAPMEGSQL DIP protein P83953 original sequence version: 39 DIP protein P83953 original sequence version: 83 B5B66B860D96EC5B nsmf_mouse NMDA receptor synaptonuclear signaling and neuronal migration factor Nsmf Jac Nelf Juxtasynaptic attractor of caldendrin on dendritic boutons protein Nasal embryonic luteinizing hormone-releasing hormone factor protein protein mouse Mus musculus Mouse MGAAASRRRALRSEAMSSVAAKVRAARAFGEYLSQSHPENRNGADHLLADAYSGHDGSPEMQPAPQNKRRLSLVSNGRYEGSISDEAVSGKPAIEGPQPHVYTISREPALLPGSEAEAIELAVVKGRRQRERHPHHHSQPLRASPGSSREDISRPCQSWAGSRQGSKECPGCAQLVPGPSSRAFGLEQPPLPEAPGRHKKLERMYSVDGVSDDVPIRTWFPKENLFSFQTATTTMQAISVFRGYAERKRRKRENDSASVIQRNFRKHLRMVGSRRVKAQTFAERRERSFSRSWSDPTPMKADTSHDSRDSSDLQSSHCTLDEACEDLDWDTEKGLEAMACNTEGFLPPKVMLISSKVPKAEYIPTIIRRDDPSIIPILYDHEHATFEDILEEIEKKLNIYHKGAKIWKMLIFCQGGPGHLYLLKNKVATFAKVEKEEDMIHFWKRLSRLMSKVNPEPNVIHIMGCYILGNPNGEKLFQNLRTLMTPYKVTFESPLELSAQGKQMIETYFDFRLYRLWKSRQHSKLLDFDDVL DE83212A82B90987 ainx_rat Alpha-internexin Ina Inexa protein protein rat Rattus norvegicus (Rat) Rat MSFGSEHYLCSASSYRKVFGDGSRLSARLSGPGASGSFRSQSLSRSNVASTAACSSASSLGLGLAYRRLPASDGLDLSQAAARTNEYKIIRTNEKEQLQGLNDRFAVFIEKVHQLETQNRALEAELAALRQRHAEPSRVGELFQRELRELRAQLEEASSARAQALLERDGLAEEVQRLRARCEEESRGREGAERALKAQQRDVDGATLARLDLEKKVESLLDELAFVRQVHDEEVAELLATLQASSQAAAEVDVAVAKPDLTSALREIRAQYESLAAKNLQSAEEWYKSKFANLNEQAARSTEAIRASREEIHEYRRQLQARTIEIEGLRGANESLERQILELEERHSAEVAGYQDSIGQLESDLRNTKSEMARHLREYQDLLNVKMALDIEIAAYRKLLEGEETRFSTSGLSISGLNPLPNPSYLLPPRILSSTTSKVSSAGLSLKKEEEEEEEEEEGASKEVTKKTSKVGESFEETLEETVVSTKKTEKSTIEEITTSSSQKM 287E548F0DA4D818 mk03_human Mitogen-activated protein kinase 3 ERT2 MAP kinase isoform p44 Microtubule-associated protein 2 kinase Extracellular signal-regulated kinase 1 Insulin-stimulated MAP2 kinase p44-ERK1 MAPK3 ERK1 PRKM3 protein protein human Homo sapiens Human MAAAAAQGGGGGEPRRTEGVGPGVPGEVEMVKGQPFDVGPRYTQLQYIGEGAYGMVSSAYDHVRKTRVAIKKISPFEHQTYCQRTLREIQILLRFRHENVIGIRDILRASTLEAMRDVYIVQDLMETDLYKLLKSQQLSNDHICYFLYQILRGLKYIHSANVLHRDLKPSNLLINTTCDLKICDFGLARIADPEHDHTGFLTEYVATRWYRAPEIMLNSKGYTKSIDIWSVGCILAEMLSNRPIFPGKHYLDQLNHILGILGSPSQEDLNCIINMKARNYLQSLPSKTKVAWAKLFPKSDSKALDLLDRMLTFNPNKRITVEEALAHPYLEQYYDPTDEPVAEEPFTFAMELDDLPKERLKELIFQETARFQPGVLEAP DIP protein P27361 original sequence version: 171 DIP protein P27361 original sequence version: 101 DIP protein P27361 original sequence version: 124 DIP protein P27361 original sequence version: 129 DIP protein P27361 original sequence version: 143 E6020CE413EC41F7 nsmf_rat NMDA receptor synaptonuclear signaling and neuronal migration factor Nsmf Jac Nelf Juxtasynaptic attractor of caldendrin on dendritic boutons protein Nasal embryonic luteinizing hormone-releasing hormone factor protein protein rat Rattus norvegicus (Rat) Rat MGAAASRRRALRSEAMSSVAAKVRAARAFGEYLSQSHPENRNGADHLLADAYSGHEGSPEMQPAPHNKRRLSLVSNGRYEGSISDEAVSGKTATEGPQPRVYTISREPALLPGSEAEAIELAVVKGRRQRERHPHHHSQPLRASPGSSREDISRPCQSWAGSRQGSKECPGCAKLVPGPSPRAFGLEQPPLPEASGRHKKLERMYSVDGVSDDVPIRTWFPKENPFSFQTATTTMQAISVFRGYAERKRRKRENDSASVIQRNFRKHLRMVGSRRVKAQTFAERRERSFSRSWSDPTPMKADTSHDSRDSSDLQSSHCTLDEACEDLDWDTEKGLEATACDTEGFLPPKVMLISSKVPKAEYIPTIIRRDDPSIIPILYDHEHATFEDILEEIEKKLNIYHKGAKIWKMLIFCQGGPGHLYLLKNKVATFAKVEKEEDMIHFWKRLSRLMSKVNPEPNVIHIMGCYILGNPNGEKLFQNLRTLMTPYKVTFESPLELSAQGKQMIETYFDFRLYRLWKSRQHSKLLDFDDVL 01AEF4B6A09EB753 nsmf-map2k1-1 2 11 unspecified role unspecified role prey prey required nphosres N-phosphorylated residue NPhosRes undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined 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 ecoli Escherichia coli (strain K12) 12 unspecified role unspecified role bait bait n_terminus his tag his tag Hexa-His-tag 6-His-tag Histidine-tag Hexa-His-tag Histidine-tag 6-His-tag n-term range n-terminal range n-term range n-term range n-terminal range n-term range region sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain n_terminal sumo tag tag tag n-term range n-terminal range n-term range n-term range n-terminal range n-term range ecoli Escherichia coli (strain K12) direct interaction direct interaction An increase in response was observed on addition of ATP, which promote the phosphorylation of Jacob by ERK1 ATP Fig. 1F, 1G nsmf-map2k1-2 3 12 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) 11 unspecified role unspecified role prey prey required nphosres N-phosphorylated residue NPhosRes undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined ecoli Escherichia coli (strain K12) direct interaction direct interaction a GST-Jacob fusion protein interacts more efficiently with active versus inactive ERK1. Fig. 1H mapk3-nsmf-1 4 9 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 n_terminus his tag his tag Hexa-His-tag 6-His-tag Histidine-tag Hexa-His-tag Histidine-tag 6-His-tag n-term range n-terminal range n-term range n-term range n-terminal range n-term range n_terminal sumo tag tag tag n-term range n-terminal range n-term range n-term range n-terminal range n-term range ecoli Escherichia coli (strain K12) 11 enzyme enzyme unspecified role unspecified role required nphosres N-phosphorylated residue NPhosRes undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined ecoli Escherichia coli (strain K12) phosphorylation phosphorylation reaction phosphorylation Ser180 and 181 are phosphorylated by ERK1 Supp. Fig. 1B ina-nsmf 1 12 unspecified role unspecified role unspecified role unspecified role region sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain region sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain 10 unspecified role unspecified role unspecified role unspecified role region required to bind necessary binding region required to bind certain certain sequence position certain certain certain sequence position certain physical association physical association Supp. Fig. 4A ina-nsmf-3 1 10 unspecified role unspecified role unspecified role unspecified role region sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain 12 unspecified role unspecified role unspecified role unspecified role region sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain 44 42 physical association physical association Supp. Fig. 4A ina-nsmf-2 5 12 unspecified role unspecified role prey prey region myc tag myc tag EUKLISEED epitope tag EUKLISEED epitope tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined human-293 Homo sapiens transformed primary embryonal kidney cells Human 293 Transformed human primary embryonal kidney cells. The Cell type named 293 is synonymous with HEK-293. 10 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 ecoli Escherichia coli (strain K12) physical association physical association Supp. Fig. 4D nsmf-ina 7 12 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 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 ecoli Escherichia coli (strain K12) 10 unspecified role unspecified role prey prey rat-brain Rattus norvegicus brain brain Brain [cerebrum] physical association physical association Supp. Fig. 4D ina-kpna1 6 8 unspecified role unspecified role prey prey 12 unspecified role unspecified role prey prey 10 unspecified role unspecified role prey prey association association Fig. 4A Erk1/2 also present in the immunoprecipitate.