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Thursday9/82016 MikeMueckler [email protected] IntracellularTarge?ngofNascentPolypep?des MitochondriaaretheSitesof Oxida?veATPProduc?on Sugars Figure 14-10 Molecular Biology of the Cell (© Garland Science 2008) Triglycerides MitochondrialBiogenesis • Mitochondriacontaintheirowngenomeand proteinsynthe3cmachinery(tRNAs,mRNAs, ribosomes,ini3a3onandelonga3onfactors,etc.) • Mitochondriaarecomprisedofhundredsof dis3nctproteins,onlyahandfulofwhichare encodedinthemitochondrialgenome(variesby species) • Mostmitochondrialproteinsareencodedin nuclearDNA,synthesizedinthecytosol,and importedpost-transla3onallyintotheorganelle MitochondriaPossess4Subcompartments UseofinvitroSystemstoElucidate MitochondrialImportMechanisms ProteinsareIncorporatedIntoMitochondria ViaSeveralDifferentRoutes Figure 12-23 Molecular Biology of the Cell (© Garland Science 2008) Targe?ngtotheMatrixRequiresanNTerminalImportSequence N-terminalImportSequencesFormAmphipathicα HelicesthatInteractwiththeTom20/22Receptor HydrophobiccleW Figure 12-22 Molecular Biology of the Cell (© Garland Science 2008) ProteinImportintotheMatrix RequiresPassageThrough TwoSeparateMembraneTranslocons ProteinsTraversetheTOMandTIM TransloconsinanUnfoldedState Transloca?oninto theMatrixOccursat ZonesofAdhesion ProteinImportintotheMatrixRequiresATP HydrolysisandanIntactProtonGradient AcrosstheInnerMembrane Figure 12-26 Molecular Biology of the Cell (© Garland Science 2008) Targe?ngtotheInnerMembraneOccursVia3Dis?nctRoutes Stop-Transfer-Mediated Oxa1-Mediated Single-Pass Proteins Cytochromeoxidase subunitCoxVa Tom70/Tim22/54-Mediated Mul?-PassProteins ADP/ATPAn?porter ATPSynthaseSubunit9 Targe?ngtotheIntermembranous SpaceOccursViaTwoDis?nctPathways IMSpaceProtease CytochromeB2 DirectDelivery CytochromecHemeLyase Targe?ngtotheOuterMembrane ViatheSAMProteinComplex (Sor3ngand Assembly Machinery) (β-Barrell) Figure 12-27 Molecular Biology of the Cell (© Garland Science 2008) Nuclear Transport • Bidirec?onal • SingleLargePoreComplex Spans2lipidbilayers • NuclearPoresmuchlarger thanothertranslocons Figure 12-8 Molecular Biology of the Cell (© Garland Science 2008) EMofTransverseSec?onShowinga Side-ViewthroughtwoNPCs Figure 12-9c Molecular Biology of the Cell (© Garland Science 2008) ScanningEMofNPCsasViewedfrom theNucleoplasm Figure 12-9b Molecular Biology of the Cell (© Garland Science 2008) StructureofaNuclearPoreComplex Figure 12-9a Molecular Biology of the Cell (© Garland Science 2008) GatedDiffusionBarrierModelof NuclearTransport Meshworkofdisordered proteindomains containingFGrepeats Figure 12-10 Molecular Biology of the Cell (© Garland Science 2008) NuclearImportSignalsareHighly DiverseinSequence • Bindtodis?nctnuclearimportreceptors • Canbeanywhereintheproteinsequence butprobablyresideonsurfacepatches • Somearenotyetiden?fied Figure 12-11 Molecular Biology of the Cell (© Garland Science 2008) GoldPar?clesCoatedwithPep?des ContainingaNLSTraverseNPCs Proteinsdonothavetobe unfoldedbeforethey traversethenuclearpore Figure 12-12 Molecular Biology of the Cell (© Garland Science 2008) NuclearImportandExportSequencesare RecognizedbyDifferentMembersofthe SameReceptorFamily(Keryopherins) Figure 12-13 Molecular Biology of the Cell (© Garland Science 2008) Direc?onalityisConferredonNuclearTransportbya GradientofRan-GDP/GTPAcrosstheNuclearEnvelope Figure 12-14 Molecular Biology of the Cell (© Garland Science 2008) NuclearImportandExportOperateViaReciprocal UseoftheRan-GDP/GTPConcentra?onGradient Figure 12-15 Molecular Biology of the Cell (© Garland Science 2008)