考研资料：北京大学细胞生物学第六章基质与内膜下课件

B.SignalHypothesis

--G.Blobel&D.Sabatini,1971.AmodelfortheSignalMechanismofCotranslationalImportEvidenceThatProteinSynthesizedonRibosomesAttachedtoERMembranesPassDirectlyintotheERLumen(D.Sabatini)Milstein:IgGMilsteinetal:StudyingthesynthesisoflightchainofIgG(incell-freesystems,20AalongeratN-terminalendthantheauthenticlightchain)AddingERmembranestothissystemleadstotheproductionofanIgGlightchainofthecorrectsize.

ASchematicmodelforthesynthesisofasecretoryproteinonamembrane-boundribosomeoftheroughERSignal-recognitionparticle,SRP:Sixdifferentpolypeptidescomplexedwitha300-nucleotide(7S)moleculeofRNA.ERsignalsequence:Typically15-30aminoacids:Consistofthreedomains:apositivelychargedN-terminalregion,acentralhydrophobicregion,andapolarregionadjoiningthesitewherecleavagefromthematureproteinwilltakeplace.AsignalsequenceonnascentseretoryproteinstargetsthemtotheERandisthencleavedoffSRPreceptor(GTPbindingprotein)SRPhavethreemainactivesites:OnethatrecognizesandbindstoERsignalsequence;Onethatinteractswiththeribosometoblockfurthertranslation;OnethatbindstotheERmembrane(dockingprotein))ThesortingsignalofVSVglycoproteins:Asp-X-Gln或DXEFigure6-43.Thesortingofproteinsdestinedfortheapicalandbasolateralplasmamembranesofepithelialcells.

WhenculturedMDCKcellsareinfectedsimultaneouslywithVSVandinfluenzavirus,theVSVglycoproteinisfoundonlyonthebasolateralmembrane,whereastheHAglycoproteinoftheinfluenzavirusisfoundonlyontheapicalmembrane.Liketheseviralproteins,somecellularproteinsaresorteddirectlytotheapicalmembraneandotherstothebasolateralmembraneviaspecifictransportvesiclesthatbudfromthetrans-Golginetwork.Incertainotherpolarizedcells,someapicalandbasolateralproteinsaretransportedtogethertothebasolateralsurface;theapicalproteinsthenmoveselectively,byendocytosisandtranscytosis,totheapicalmembrane.[K.Simonsetal.,Cell

62:207;K.Mostovetal.,JCB.

116:577]Start-transferSequence&Stop-transferSequenceC.

AModelforthePostranslationalImportofPolypeptidesintotheMit.Post-translationalmodificationandqualitycontrolintherERDisulfidebondsareformedandrearrangedintheERlumenOnlyinERlumenistherearedoxenvironmentforoxidationof–SHgroups.PDI:proteindisulfideisomerase,foundinabundanceintheERlumenCorrectfoldingofnewlymadeproteinsisfacilitatedbyseveralERproteinsProteinswithoutanysignalsequencearecytosolresidualproteins6.

TypesofVesicleTransportandTheirFunctionsA.Thethreedifferenttypesofcoatedvesicles.Differentcoatproteinsselectdifferentcargoandshapethetransportvesiclesthatmediatethevariousstepsinthebiosynthetic-secretoryandendocyticpathways.COPII-coatedvesiclesmovematerialsfromtheERtotheGolgi.TheassemblyofaCOPII-coatedvesicles.Sar—GTPbindingprotein:Sar-GTPbindstotheER;Sar-GDPdissociatesfromtheERAntibodiesisabletoblockthebuddingofvesiclefromERbuthavenoeffectonvesicletransportfromoneGolgicompartmenttoanotherinmammaliancell.COPI-coatedvasiclestransportingEscapedERresidentProteinsBacktotheER.TheassemblyofaCOPI-coatismediatedbyADP-ribosylationfactor(ARF),GTPbindingprotein,whichisrequiredforvesicletransferbetweencisternae.COPIcoatedvesiclesmayselectspecificcargo.

ERisanopenprison.

SolubleERproteinbearRetrievingsignal—KDEL(Lys-Asp-Glu-Leu)inmammalandHDELinyeast,whereasERmembraneproteinsbearthesignalKKXX.

TheKDELreceptorpresentinvesiculartubularclustersandtheGolgiapparatus..(3)COPI-coatedvesiclewerefirstidentifiedbytreatmentofGTPanalogues---COPI-coatedvesicleaccumulatedwithinthecellandcouldbeisolatedbycentrifugation.AmodelfortheretrievalofERresidentproteins.The

KDELreceptorcapturesthesolubleERresidentproteinsandcarriestheminCOPI-coatedtransportvesiclesbacktotheER.NeutralpH:dissociatefromtheKDEL;lowpH:bindingtheKDEL

Clathrin-coatedvesicle:TransportingCargofromtheTGNtoendosomes,Lysosomes,andplantvacuolesandalsomovematerialsfromthePMtocytoplasmiccompartmentsalongtheendocyticpathway.TheTGNofGolgiistheSourseofClathrin-coatedvesicle.(2)Clathrin-coatscontain:

proteinclathrin-----whichformsastructuralscaffold,adaptors----multisubunit,whichformsaninnershell.

Theformationofclathrin-coatedpitsintheTGNB.TheSNAREHypothesisforTransportVesicleTargetingandFusionSpecificityinvesicledockingandfusionisthoughttobeattainedthroughspecificinteractionsbetweenspecificv-SNAREproteinsonthevesiclemembranesandt-SNAREproteinsonthemembranesofthetargetcompartment.SNAREsareaproteinfamily.Thereareatleast20differentSNAREsinananimalcell.TheroleofSNAREsinguidingvesiculartransport.ThestructionofpairedSNAREs(thefour-helixbundle).TheSNAREsresponsiblefordockingsynapticvesiclesattheplasmamembraneofnerveterminalsconsistofthreeproteins.Thebasicmolecularcomponentsineukaryoticcellsincludev-SNAREs(v-SNAPreceptors)ontransportvesicles,t-SNAREs(t-SNAPreceptors)ontargetmembranes,RabGTPase,N