Mueckler Review slides - Washington University in St. Louis Sessions/2017 Review... ·...

MCBTest1– MuecklerReview

9-30-17

Func%onsofCellularMembranes1.   Plasmamembraneactsasaselec%velypermeablebarriertothe

environment•  Uptakeofnutrients•  Wastedisposal•  Maintainsintracellularionicmilieu2.   Plasmamembranefacilitatescommunica%on•  Withtheenvironment•  Withothercells3.Intracellularmembranesallowcompartmentaliza%onandsepara%onofdifferentchemicalreac%onpathways•  Increasedefficiencythroughproximity•  PreventfuHlecyclingthroughseparaHon•  ProteinsecreHon

FaJyAcidSaturaHon

Figure 10-11a Molecular Biology of the Cell (© Garland Science 2008)

(Hydrophobic)

(Hydrophilic)

BilayersareThermodynamicallyStableStructuresFormedfromAmphathicLipids

Figure 10-8 Molecular Biology of the Cell (© Garland Science 2008)

TheForma%onofCell-LikeSphericalWater-FilledBilayersisEnerge%callyFavorable

Figure 10-11b Molecular Biology of the Cell (© Garland Science 2008)

PhosphoLipidMovementswithinBilayers(µM/sec)

(1012-1013/sec) (108-109/sec)

Figure 12-58 Molecular Biology of the Cell (© Garland Science 2008)

A“Scramblase”EnzymeCatalyzesSymmetric

GrowthofBothLeafletsintheER

Figure 12-58 Molecular Biology of the Cell (© Garland Science 2008)

A“Flippase”EnzymepromotesLipid

AsymmetryinthePlasmaMembrane

Figure 10-17a Molecular Biology of the Cell (© Garland Science 2008)

PhospholipidsareInvolvedinSignalTransduc%on

PI-4,5P PI-3,4,5P

1.Ac%va%onofLipidKinases

2.PhospholipasesProduceSignalingMoleculesviatheDegrada%onof

Phospholipids

CholesterolBiosynthesisOccursintheCytosolandattheERMembraneThroughIsoprenoidIntermediates

(Rate-LimiHngStepinER)

3WaysinwhichLipidsMaybeTransferredBetweenDifferentIntracellularCompartments

VesicleFusionDirectProtein-Mediated

TransferSolubleLipid

BindingProteins

Figure 10-19 Molecular Biology of the Cell (© Garland Science 2008)

The3BasicCategoriesofMembraneProtein

Integral Lipid-Anchored

Peripheral(canalsointeractviaPLheadgroups)

Single-Pass Mul%-pass

Fa_yacylanchor

GPIAnchor

TransmembraneHelix

LinkerDomain

β-Strands

Figure 3-5 Molecular Biology of the Cell (© Garland Science 2008)

MembraneDomainsare“Inside-Out”

Right-SideOutSolubleProtein

Figure 10-31 Molecular Biology of the Cell (© Garland Science 2008)

Func%onalCharacteriza%onofIntegralMembraneProteinsRequiresSolubiliza%onandSubsequentRecons%tu%onintoaLipidBilayer

Figure 10-39 Molecular Biology of the Cell (© Garland Science 2008)

4WaysthatProteinMobilityisRestrictedinBiologicalMembranes

IntramembraneProtein-ProteinInteracHons

InteracHonwiththecytoskeleton

InteracHonwiththeextracellularmaxtrix

IntercellularProtein-ProteinInteracHons

Figure 12-41a Molecular Biology of the Cell (© Garland Science 2008)

RibosomalSubunitsareShared

BetweenFreeandMembrane-Bound

Polysomes

Targe&nginforma&onresides

intheNascentpolypep&dechain

Signal-MediatedTarge%ngtotheRER

Proper%esofSecretorySignalSequences

HydrophobicCoreN MatureProtein

8-12Residues

15-30Residues

++

• LocatedatN-terminus• 15-30Residuesinlength• Hydrophobiccoreof8-12residues• OZenbasicresiduesatN-terminus(Arg,Lys)• Nosequencesimilarity

cleavage

InVitroTransla%on/Transloca%onSystem

•  mRNA•  Roughmicrosomes•  Ribosomes•  tRNAs•  SolubletranslaHonfactors•  LowMWcomponents•  Energy(ATP,creaHne-P,creaHnekinase)

ReHculocyteorwheatgermlysate

InVitroTransla%on/Transloca%onSystem

mRNA+

TranslaHonComponents

+Aminoacid*

Protein* SDSPAGE

InVitroTransla%onofProlac%nmRNA

ProlacHnisapolypepHdehormone(MW~22kd)secretedbyanteriorpituitary

12345678MW(kd)

2522

Lanes:1.  PurifiedprolacHn2.  NoRM3.  RM4.  NoRM/digestwith

Protease5.  RM/digestwith

Protease6.  RM/detergenttreatand

addProtease7.  ProlacHnmRNAminus

SS+RM/digestwithProtease

8.  SS-globinmRNA+RM/digestwithProtease

18

SDSGel

Figure 12-48 Molecular Biology of the Cell (© Garland Science 2008)

Mul%passTopologiesareGeneratedbyMul%pleInternalSignal/AnchorSequences

TypeIVa

+–

+–

+–

+–

TheChargeDifferenceRuleforMul%spanningMembraneProteins

NH2 COOH

NH2

COOH

+ + –

+ +

NH2 COOH

NH2

COOH

+ +

+ +cytoplasm

cytoplasm

––

–

–

–

–

Figure 12-53 Molecular Biology of the Cell (© Garland Science 2008)

OligosaccharideProcessingintheRERisUsedforQualityControl

DisulfideBridgesareFormedintheRERbyProteinDisulfideIsomerase(PDI)

Figure 12-23 Molecular Biology of the Cell (© Garland Science 2008)

ProteinsareIncorporatedIntoMitochondriaViaSeveralDifferentRoutes

Figure 12-26 Molecular Biology of the Cell (© Garland Science 2008)

ProteinImportintotheMatrixRequiresATPHydrolysisandanIntactProtonGradient

AcrosstheInnerMembrane

Targe%ngtotheInnerMembraneOccursVia3Dis%nctRoutes

Oxa1-MediatedStop-Transfer-Mediated Tom70/Tim22/54-Mediated

Mul%-PassProteinsSingle-PassProteins

CytochromeoxidasesubunitCoxVa

ATPSynthaseSubunit9

ADP/ATPAn%porter

CytochromeB2 CytochromecHemeLyase

Targe%ngtotheIntermembranousSpaceOccursViaTwoDis%nctPathways

DirectDeliveryIMSpaceProtease

Figure 12-27 Molecular Biology of the Cell (© Garland Science 2008)

Targe%ngtotheOuterMembraneViatheSAMProteinComplex

(SorHngandAssemblyMachinery)

(β-Barrell)

Figure 12-8 Molecular Biology of the Cell (© Garland Science 2008)

NuclearTransport

• Bidirec%onal• SingleLargePoreComplexSpans2lipidbilayers• NuclearPoresmuchlargerthanothertranslocons

Figure 12-13 Molecular Biology of the Cell (© Garland Science 2008)

NuclearImportandExportSequencesareRecognizedbyDifferentMembersoftheSameReceptorFamily(Keryopherins)

Figure 12-15 Molecular Biology of the Cell (© Garland Science 2008)

NuclearImportandExportOperateViaReciprocalUseoftheRan-GDP/GTPConcentra%onGradient