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Millipore/15-103 | PI3 Kinase/Akt/GSK3 Pathway Explorer Minipack/15-103/3 vials/Pk

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貨號: 15-103

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Description
CatalogueNumber	15-103
BrandFamily	Upstate
TradeName	Upstate
Description	PI3Kinase/Akt/GSK3PathwayExplorerMinipack
AlternateNames	ProteinkinaseB RAC-alphaserine/threonine-proteinkinase racproteinkinasealpha v-aktmurinethymomaviraloncogenehomolog1 murinethymomaviral(v-akt)oncogenehomolog-1 glycogensynthasekinase3alpha glycogensynthasekinase3beta
BackgroundInformation	PathwayExplorerAntibodyMiniPack: EachPathwayExplorerAntibodyMinipackcontainsthreerelatedantibodiesaspartofasignalingcascadeoracombinationoftotalandphosphorylatedformsofkeysignalingtargets.Eachofthethreeantibodiesare30%theoriginalpacksize.FullsizeversionsofeachofthePathwayExplorerantibodiesareavailableforsaleindividuallyunderthesamecatalognumberwiththeremovalof“SP”offofeachone(e.g.05-591SPcanbeorderedas05-591). PI3Kinase: PI3Kinase(Phosphatidylinositol3-Kinase)isresponsIBLeforphosphorylationofthe3positionoftheinositolringofPI(4,5)P2,togeneratePI(3,4,5)P3,apotentsecondmessengerrequiredforsurvivalsignaling,andinsulinaction.PI3Kinaseisaheterodimericcomplexcomposedofan85kDaregulatorysubunitanda110kDacatalyticsubunit.Tyrosinephosphorylationofgrowthfactorreceptorscreatesdockingsitesforbindingofp85(throughitsSH2domains)onthereceptors;p85bringswithitp110,whichisthenproximaltoitsphospho-lipidsubstrateonthemembrane.PI3KinaseisalsoactivatedbyRas,andbytheβ-γsubunitsofheterotrimericG-proteins.PI3Kinaseisinhibitablebywortmannin,ausefultoolforthestudyofthePI3Kinasesignalingpathway. Akt/PKB,PHDomain: Akt(proteinkinaseB),aserine/threoninekinase,hasemergedasacriticalenzymeinsignaltransductionpathwaysinvolvedincellproliferation,apoptosis,angiogenesis,anddiabetes.InmammalsthreeisoformsofAkt(α,β,γorAkt1,2,3)arereportedthatexhibitahighdegreeofhomology,butdifferslightlyinthelocalizationoftheirregulatoryphosphorylationsites.Aktαisthepredominantisoforminmosttissues,whereasthehighestexpressionofAktβisobservedintheinsulin-responsivetissues,andAktγisabundantinbraintissue.EachAktisoformiscomposedofthreefunctionallydistinctregions:anN-terminalpleckstrinhomology(PH)domainthatprovidesalipid-bindingmoduletodirectAkttoPIP3,acentralcatalyticdomain,andaC-terminalhydrophobicmotif. GSK3α/β: Glycogensynthasekinase-3(GSK-3),amultifunctionalserine/threoninekinase,isakeyregulatorofnumeroussignalingpathways.TwoisoformsofGSK-3arereportedinmammals:a51kDaGSK-3αanda47kDaGSK-3β.Thesetwoisoformsexhibitabout98%homologyintheirkinasedomains,butshareonlyabout36%identityinthelast76C-terminalaminoacidresidues.GSK-3αcontainsaglycine-richextensionatitsN-terminus.Aminor(~15%oftotal)splicevariantofGSK-3β,GSK-3β2,hasalsobeenidentified,whichcontainsa13-residueinsertwithinthekinasedomain.ItexhibitsreducedkinaseactivitytowardstheTauproteincomparedwith‘unspliced’GSK-3β.GSK-3β2islocalizedprimarilytoneuronalcellbodies,unlikeunsplicedGSK-3βthatisalsofoundinneuronalprocesses.GSK-3βplaysakeyinhibitoryroleintheWntsignalingpathway.Wntgenesencodealargefamilyofsecreted,cysteine-richproteinsthatareimportantindevelopmentandinmaintenanceofadulttissues.AbnormalitiesinWntsignalingarereportedtopromotebothhumandegenerativediseasesandcancer. *Seefullsizeversionsforcorrespondingreferences.

ProductInformation
Components	06-195SPAnti-PI3Kinase,p85 05-591SPAnti-Akt/PKB,PHDomain,cloneSKB1 05-412SPAnti-GSK3α/β,clone4G-1E
Control	05-412SP:IncludedPositiveAntigenControl:Catalog#12-303,Jurkatlysate.Add2.5μLof2-mercaptoethanolper100μLoflysateandboilfor5minutestoreducethepreparation.Use20μgperlaneforminigels. 06-195SP:Jurkatcelllysate. IncludedPositiveAntigenControl:Catalog#12-303,Jurkatcelllysate.Add2.5μLof2-mercaptoethanolper100μLoflysateandboilfor5minutestoreducethepreparation.Load20μgofreducedlysateperlaneforminigels.
Presentation	3individualtubescontainingeitherAnti-PI3Kinase,p85;Anti-Akt/PKB,PHDomain,cloneSKB1;orAnti-GSK3α/β,clone4G-1E
Properties	Eachvialis30%thesizeoftheparentcatalognumber

StorageandShippingInformation

Applications
Application	ThisAntibodypackcontainsAnti-PI3KinaseAntibody,Anti-Akt/PKBAntibody,Anti-GSK3α/βAntibodyvalidatedforuseinEnzymeAssay,ICC,IF,Flow,IP&WB.
KeyApplications	EnzymeAssays Immunocytochemistry Immunofluorescence FlowCytometry Immunoprecipitation WesternBlotting
ApplicationsNotRecommended	Immunoprecipitation
ApplicationNotes	05-412SP:ImmunofluorescenceStaining: L6cellsweregrown,fixed,andpermeABIlized.; FlowCytometry: NCI/H292cellswerestainedwithAnti-GSK3α/β,clone4G-1EconjugatedtoAlexaFluor?555;Immunoprecipitation:Notrecommendedfor05-412SP. 06-195SP:Immunoprecipitation:5μLofapreviouslotimmunoprecipitatedPI3kinasefromhumanJurkatcelllysateasconfirmedbysubsequentwesternblot.Theantibodywillprecipitatethe85kDasubunit. 05-591SP:ImmunofluroescenceAnalysis: A431cellswerefixed,permeabilized,andstainedwithanti-Akt,cloneSKB1(2μg/mL)conjugatedtoAlexaFluor?488; FlowCytometry:NIH/3T3cells(detachedusingtrypsin-EDTA)persamplewerestainedwith0.5μgofanti-Akt/PKB,PHDomain,cloneSKB1(filledhistogram)or1μgofmouseIgG1(openhistogram)followedbyagoatanti-mouseIgG-PEconjugate. ImmunoprecipitationKinase:4μgofapreviouslotimmunoprecipitatedactiveAktfrom700μgofL6cellsstimulatedwith15nMIGF-1for2-5minutes. ImmunoprecipitationKinaseAssayData: ThekinaseactivityofimmunoprecipitatedAkt/PKBwasdeterminedasdescribedaboveusingAkt/SGKSubstratePeptideasasubstrate.Activekinasewasdetectedbycomparingtheincorporationof[32P]intoAkt/SGKSubstratePeptideascatalyzedbyimmunoprecipitates(1mg)ofL6celllysatesstimulatedwith15nMIGF-1.Thetestresultsareshowninthetabletotheleft.

BIOLOGicalInformation
Immunogen	05-412SP:18residuesyntheticpeptide(C-KQLLHGEPNVSYICSRY)correspondingtoaminoacids203-219derivedfromaregionofthecatalyticdomain(betweensuBDomainVIIandVIII)oftheDrosophilaGSK3/shaggyenzyme,coupledtoovalbumin.Clone4G-1E.;05-591SP:GST-fusionproteincorrespondingtoresidues1-149ofhumanAkt1.;06-195SP:GSTfusionproteinscontaining1)theN-terminalSH2domainofratPI3kinaseand2)thefulllength85kDasubunitofratPI3kinase.
Epitope	Varies.SeeindividualDataSheets
Clone	05-412SP:4G-1E;05-591SP:SKB1
Concentration	PleaserefertotheCertificateofAnalysisforthelot-specificconcentration.
SpeciesReactivity	Human Mouse Rat Vertebrates Monkey Yeast(S.pombe) Dictyostelium
SpeciesReactivityNote	05-412SP:Human,Mouse,Rat,S.pombe,andDictyostelium.Becauseoftheextremelyhighconservationofthisimmunogenamongtheknownenzymesofmanydifferentspecies,thisantibodyispredictedtocross-reactwithnearlyallspecieshomologues. 05-591SP:Human,mouseandrat;otherspeciescross-reactivityunknown.
EntrezGeneNumber	NP_852556 NP_001014431 NP_476714
EntrezGeneSummary	AKT1:Theserine-threonineproteinkinaseencodedbytheAKT1geneiscatalyticallyinactiveinserum-starvedprimaryandimmortalizedfibroblasts.AKT1andtherelatedAKT2areactivatedbyplatelet-derivedgrowthfactor.Theactivationisrapidandspecific,anditisabrogatedbymutationsinthepleckstrinhomologydomainofAKT1.Itwasshownthattheactivationoccursthroughphosphatidylinositol3-kinase.InthedevelopingnervoussystemAKTisacriticalmediatorofgrowthfactor-inducedneuronalsurvival.Survivalfactorscansuppressapoptosisinatranscription-independentmannerbyactivatingtheserine/threoninekinaseAKT1,whichthenphosphorylatesandinactivatescomponentsoftheapoptoticmachinery.Multiplealternativelysplicedtranscriptvariantshavebeenfoundforthisgene.[providedbyRefSeq]. P13KINASE:Phosphatidylinositol3-kinasephosphorylatestheinositolringofphosphatidylinositolatthe3-primeposition.Theenzymecomprisesa110kDcatalyticsubunitandaregulatorysubunitofeither85,55,or50kD.Thisgeneencodesthe85kDregulatorysubunit.Phosphatidylinositol3-kinaseplaysanimportantroleinthemetabolicactionsofinsulin,andamutationinthisgenehasbeenassociatedwithinsulinresistance.Alternativesplicingofthisgeneresultsinthreetranscriptvariantsencodingdifferentisoforms.[providedbyRefSeq].
GeneSymbol	AKT Akt1 GSK3 GlycogenSynthaseKinase3 PKB PKB-ALPHA PRKBA PI3Kinase PI3K phosphatidylinositol3-kinaseregulatorysubunit phosphatidylinositol3kinase PIK3R1
UniProtNumber	P31749 P49840 P27986
UniProtSummary	PI3Kinase,p85: FUNCTION:Bindstoactivated(phosphorylated)protein-Tyrkinases,throughitsSH2domain,andactsasanadapter,mediatingtheassociationofthep110catalyticunittotheplasmamembrane.Necessaryfortheinsulin-stimulatedincreaseinglucoseuptakeandglycogensynthesisininsulin-sensitivetissues. Subunitstructure:Heterodimerofap110(catalytic)andap85(regulatory)subunits.InteractswithphosphorylatedTOM1L1.InteractswithphosphorylatedLIME1uponTCRand/orBCRactivation.InteractswithSOCS7.InteractswithRUFY3Bysimilarity.InteractswithphosphorylatedLAT,LAX1andTRAT1uponTCRactivation.InteractswithCBLB.InteractswithHIV-1NeftoactivatetheNefassociatedp21-activatedkinase(PAK).ThisinteractiondependsontheC-terminusofbothproteinsandleadstoincreasedproductionofHIV.InteractswithHCVNS5A.TheSH2domainsinteractwiththeYTHMmotifofphosphorylatedINSRinvitro.Alsointeractswithtyrosine-phosphorylatedIGF1Rinvitro.InteractswithCD28andCD3ZuponT-cellactivation.InteractswithIRS1andphosphorylatedIRS4,aswellaswithNISCHandHCST. TISSUESPECIFICITY:Isoform2isexpressedinskeletalmuscleandbrain,andatlowerlevelsinkidneyandcardiacmuscle.Isoform2andisoform4arepresentinskeletalmuscle(atproteinlevel). DOMAIN:TheSH3domainmediatesthebindingtoCBLB,andtoHIV-1Nef. Post-translationalmodification:PolyubiquitinatedinT-cellsbyCBLB;whichdoesnotpromoteproteasomaldegradationbutimpairsassociationwithCD28andCD3ZuponT-cellactivation. AKT: FUNCTION:Generalproteinkinasecapableofphosphorylatingseveralknownproteins.PhosphorylatesTBC1D4.Signalsdownstreamofphosphatidylinositol3-kinase(PI(3)K)tomediatetheeffectsofvariousgrowthfactorssuchasplatelet-derivedgrowthfactor(PDGF),epidermalgrowthfactor(EGF),insulinandinsulin-likegrowthfactorI(IGF-I).Playsaroleinglucosetransportbymediatinginsulin-inducedtranslocationoftheGLUT4glucosetransportertothecellsurface.MediatestheantiapoptoticeffectsofIGF-I.Mediatesinsulin-stimulatedproteinsynthesis,partlybyplayingaroleinbothinsulin-inducedphosphorylationof4E-BP1andininsulin-inducedactivationofp70S6kinase.Promotesglycogensynthesisbymediatingtheinsulin-inducedactivationofglycogensynthase. CATALYTICACTIVITY:ATP+aprotein=ADP+aphosphoprotein. ENZYMEREGULATION:Threespecificsites,oneinthekinasedomain(Thr-308)andthetwootheronesintheC-terminalregulatoryregion(Ser-473andTyr-474),needtobephosphorylatedforitsfullactivation. SUBUNIT:InteractswithCENTG1isoform2(PIKE-A)inthepresenceofguaninenucleotides.TheC-terminusinteractswithCCDC88A/GRDNandTHEM4.InteractswithAKTIP.Interacts(viaPHdomain)withMTCP1,TCL1AANDTCL1B.InteractswithCDKN1B;theinteractionphosphorylatesCDKN1Bpromoting14-3-3bindingandcell-cycleprogression. SUBCELLULARLOCATION:Cytoplasm.Nucleus.Cellmembrane.Note=Nucleusafteractivationbyintegrin-linkedproteinkinase1(ILK1).NucleartranslocationisenhancedbyinteractionwithTCL1A. TISSUESPECIFICITY:Inallhumancelltypessofaranalyzed. DOMAIN:BindingofthePHdomaintothephosphatidylinositol3-kinasealpha(PI(3)K)resultsinitstargetingtotheplasmamembrane. DOMAIN:TheAGC-kinaseC-terminalmediatesinteractionwithTHEM4. PTM:PhosphorylationonThr-308,Ser-473andTyr-474isrequiredforfullactivity.Ser-473phosphorylationbytheRictor-mTorcomplexfavorsThr-308phosphorylationbyPDPK1.Ser-473phosphorylationisenhancedbyinteractionwithCENTG1isoform2(PIKE-A).Ser-473phosphorylationisenhancedinfocalcorticaldysplasiaswithTaylor-typeballooncells. GSK3: FUNCTION:ParticipatesintheWntsignalingpathway.Implicatedinthehormonalcontrolofseveralregulatoryproteinsincludingglycogensynthase,MYBandthetranscriptionfactorJUN.PhosphorylatesJUNatsitesproximaltoitsDNA-bindingdomain,therebyreducingitsaffinityforDNA.PhosphorylatesMUC1inbreastcancercells,anddecreasestheinteractionofMUC1withCTNNB1/beta-catenin. CATALYTICACTIVITY:ATP+[tauprotein]=ADP+[tauprotein]phosphate. ENZYMEREGULATION:InhibitedwhenphosphorylatedbyAKT1. SUBUNIT:MonomerBysimilarity.InteractswithCABYR,MUC1,NINandPRUNE. TISSUESPECIFICITY:Expressedintestis,thymus,prostateandovaryandweaklyexpressedinlung,brainandkidney. POST-TRANSLATIONALMODIFICATION:PhosphorylatedbyAKT1andILK1. SEQUENCESIMILARITIES:Belongstotheproteinkinasesuperfamily.CMGCSer/Thrproteinkinasefamily.GSK-3subfamily.Contains1proteinkinasedomain.
MolecularWeight	05-412SP:51kDa(α)or46kDa(β);05-591SP:60kDa;06-195SP:85kDa

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