19|OxidativePhosphorylationandPhotophosphorylation
2013 W. H. Freeman and Company
CHAPTER19OxidativePhosphorylationand
Photophosphorylation
Electrontransportchaininmitochondria Captureoflightenergyinphotosynthesis Buildinguptheprotonmotiveforce SynthesisofATPinmitochondriaandchloroplasts
Keytopics:
EnergyfromreducedfuelsisusedtosynthesizeATPinanimals
Carbohydrates,lipids,andaminoacidsarethemainreducedfuels forthecell
ElectronsfromreducedfuelsaretransferredtoreducedcofactorsNADH orFADH2
Inoxidativephosphorylation,energyfromNADH andFADH2 areusedtomakeATP
EnergyFlowinCellularRespiration
OxidativePhosphorylation
ElectronsfromthereducedcofactorsNADH andFADH2arepassedtoproteinsintherespiratorychain
Ineukaryotes,oxygen istheultimateelectronacceptorfortheseelectrons
EnergyofoxidationisusedtophosphorylateADP
OxidativePhosphorylation
Photophosphorylation Inphotosyntheticorganismslight causeschargeseparationbetweenapairchlorophyllmolecules
Energyoftheoxidizedandreducedchlorophyllmolecules isusedtodrivesynthesisofATP
Wateristhesourceofelectrons thatarepassedviaachainofproteintransporterstotheultimateelectronacceptor,NADP+
Oxygenisthebyproductofwateroxidation
Photophosphorylation
ChemiosmoticTheory
ADP+Pi ATP isHighlyThermodynamicallyUnfavorable Howdowemakeitpossible? PhosphorylationofADPisnotaresultofadirectreactionbetweenADPandsomehighenergyphosphatecarrier
EnergyneededtophosphorylateADPisprovidedbytheflowofprotonsdowntheelectrochemicalgradient
Theenergyreleasedbyelectrontransportisusedtotransportprotonsagainsttheelectrochemicalgradient
Chemiosmoticenergycouplingrequiresmembranes
TheprotongradientneededforATPsynthesiscanbestablyestablishedacrossamembranethatisimpermeabletoions Plasmamembraneinbacteria Innermembraneinmitochondria Thylakoidmembraneinchloroplasts
Membranemustcontainproteinsthatcouplethedownhillflowofelectrons intheelectrontransferchainwiththeuphillflowofprotonsacrossthemembrane
Membranemustcontainaproteinthatcouplesthedownhillflowofprotons tothephosphorylationofADP
ChemiosmoticTheory
StructureofaMitochondrionDoublemembraneleadstofourdistinctcompartments:1.OuterMembrane:
Relativelyporousmembraneallowspassageofmetabolites
2.IntermembraneSpace(IMS): similarenvironmenttocytosol higherprotonconcentration(lowerpH)
3.InnerMembrane Relativelyimpermeable,withprotongradientacrossit Locationofelectrontransportchaincomplexes ConvolutionscalledCristaeservetoincreasethesurfacearea
4.Matrix Locationofthecitricacidcycleandpartsoflipidandaminoacidmetabolism
Lowerprotonconcentration(higherpH)
StructureofaMitochondrion
Electrontransportchaincomplexescontainaseriesofelectroncarriers
Eachcomplexcontainsmultipleredoxcentersconsistingof: FlavinMononucleotide(FMN)orFlavinAdenineDinucleotide(FAD) InitialelectronacceptorsforComplexIandComplexII Cancarrytwoelectronsbytransferringoneatatime
Cytochromesa,borc
Ironsulfurcluster
Cytochromes Oneelectroncarriers Ironcoordinatingporphoryin ringderivatives a,borcdifferbyringadditions
IronSulfurClusters Oneelectroncarriers Coordinatingbycysteines intheprotein Containingequalnumberofironandsulfuratoms
CoenzymeQorUbiquinone
Ubiquinone isalipidsolubleconjugateddicarbonylcompoundthatreadilyacceptselectrons
Uponacceptingtwoelectrons,itpicksuptwoprotonstogiveanalcohol,ubiquinol
Ubiquinolcanfreelydiffuseinthemembrane,carryingelectrons withprotonsfromonesideofthemembranetoanotherside
CoenzymeQisamobileelectroncarriertransportingelectronsfromComplexesIandIItoComplexIII
CoenzymeQorUbiquinone
FreeEnergyofElectronTransport
ReductionPotential(E)Eo=Eo(e acceptor) Eo(e donor)
Go=nFEoFornegativeGneedpositiveE
E(acceptor) >E(donor)
Electronsaretransferredfromlower(morenegative)tohigher(morepositive)reductionpotential.
FreeEnergyreleasedisusedtopumpproton,storingthisenergyastheelectrochemicalgradient
FlowofElectronsfromBiologicalFuelsintotheElectronTransportChain
NADH:ubiquinoneoxidoreductase,a.k.a.ComplexI
Oneofthelargestmacromolecularassembliesinthemammaliancell
Over40differentpolypeptidechains,encodedbybothnuclearandmitochondrialgenes
NADHbindingsiteinthematrixside Noncovalentlyboundflavinmononucleotide(FMN)acceptstwoelectronsfromNADH
Severalironsulfurcenterspassoneelectronatatimetoward the ubiquinonebindingsite
ComplexI
NADH:Ubiquinoneoxidoreducaseisaprotonpump
TransferoftwoelectronsfromNADHtoubiquinoneisaccompaniedbyatransferofprotonsfromthematrix(N)totheintermembranespace(P)
Experimentssuggestthataboutfourprotons aretransportedperoneNADH
NADH+Q+5H+N =NAD+ +QH2 +4H+P
ReducedcoenzymeQpicksuptwoprotons Protonsaretransportedbyprotonwires
Aseriesofaminoacidsthatundergoprotonationanddeprotonationtogetanettransferofaprotonfromonesideofamembranetoanother
SuccinateDehydrogenase,a.k.a.ComplexII
FADacceptstwoelectronsfromsuccinate Electronsarepassed,oneatatime,viaironsulfurcenterstoubiquinone,whichbecomesreducedQH2
Doesnottransportprotons
ComplexII
Ubiquinone:Cytochromec Oxidoreductase,a.k.a.ComplexIII
UsestwoelectronsfromQH2 toreducetwomoleculesofcytochromec
Additionallycontainsironsulfurclusters,cytochromebs,andcytochromecs
TheQcycleresultsinfouradditionalprotonsbeingtransportedtotheIMS
ComplexIII
TheQCycle
Experimentally,fourprotons aretransportedacrossthemembranepertwoelectronsthatreachCytC
Twoofthefourprotons comefromQH2 TheQcycleprovidesagoodmodelthatexplainshowtwoadditionalprotonsarepickedupfromthematrix
Two moleculesofQH2 becomeoxidized,releasingprotonsintotheIMS
Onemoleculebecomesrereduced, thusaNettransferoffourprotonsperreducedCoenzymeQ
TheQCycle
TheQCycle:Cycle1
TheQCycle:Cycle2
Thesecondmobileelectroncarrier Asolublehemecontainingprotein in
theintermembranespace Hemeironcanbeeitherferrous
(Fe3+,oxidized)orferric(Fe2+,reduced)
Cytochromec carriesasingleelectronfromthecytochromebc1complextocytochromeoxidase
Cytochromec
Cytochromec absorbsvisiblelight
IntenseSoretbandnear400nmabsorbsbluelightandgivescytochromec anintenseredcolor
Cytochromesarenamedbythepositionoftheirlongestwavelength()peak
CytochromeOxidase,a.k.a.ComplexIV
Mammaliancytochromeoxidaseisamembraneproteinwith13subunits
Containstwohemegroups:aanda3 Containscopperions
CuA:twoionsthatacceptelectronsfromCytc CuB:bondedtohemea3 formingabinuclearcenterthattransfersfourelectronstooxygen
ComplexIV
CytochromeoxidasepasseselectronstoO2
Fourelectronsareusedtoreduce oneoxygenmoleculeintotwowatermolecules
Fourprotonsarepickedupfromthematrix inthisprocess Fouradditionalprotonsarepassedfromthematrixtotheintermembranespace
ElectronflowthroughComplexIV
Multiplecomplexesassociatetogethertoformarespirasome
SummaryoftheElectronFlowintheRespiratoryChain
SummaryofElectronTransport
ComplexI ComplexIV1NADH +11H+(N) +O2 >NAD+ +10H+(P) +H2O
ComplexII ComplexIVFADH2 +6H+(N) +O2 >FAD+6H+(P) +H2O
DifferenceinnumberofprotonstransportedreflectsdifferencesinATPsynthesized.
Reactiveoxygenspeciescandamagebiologicalmacromolecules
ProtonMotiveForce
Theproteinsintheelectrontransportchaincreatedtheelectrochemicalprotongradient byoneofthreemeans: activelytransportprotonsacrossthemembrane
ComplexIandComplexIV
Chemicallyremoveprotonsfromthematrix ReductionofCoQandreductionofoxygen
Releaseprotonsintotheintermembranespace OxidationofQH2
ProtonMotiveForce
ChemiosmoticModelforATPSynthesis Electrontransportsetsupaprotonmotiveforce EnergyofprotonmotiveforcedrivessynthesisofATP
Consequently,electrontransportiscoupledtoATPsynthesis
Asdescribed,ATPsynthesisrequireselectrontransportButelectrontransportalsorequiresATPsynthesis
MitochondrialATPSynthaseComplex
Containstwofunctionalunits: F1
Solublecomplexinthematrix IndividuallycatalyzesthehydrolysisofATP
F0 Integralmembranecomplex TransportsprotonsfromIMStomatrix,dissipatingtheprotongradient
EnergytransferredtoF1 tocatalyzephosphorylationofADP
MitochondrialATPSynthaseComplex
TheF1 catalyzesADP+Pi ATP Hexamerarrangedinthree dimers Dimerscanexistinthreedifferentconformations:
Open:empty Loose:bindingADPandPi Tight:catalyzesATPformationandbindsproduct
BindingChangeModel
CouplingProtonTranslocationtoATPSynthesis
ProtontranslocationcausesarotationoftheF0 subunitandthecentralshaft
Thiscausesaconformationalchange withinallthethree pairs
TheconformationalchangeinoneofthethreepairspromotescondensationofADPandPi intoATP
EvidenceofRotation
TransportofADPandPi intotheMatrix
MalateAspartateShuttle
Glycerol3PhosphateShuttle
RegulationofOxidativePhosphorylation
Primarilyregulatedbysubstrateavailability NADHandADP/Pi DuetocouplingbothsubstratesrequiredforelectrontransportandATPsynthesis
InhibitorofF1 (IF1) PreventshydrolysisofATPduringlowoxygen OnlyactiveatlowerpH,encounteredwhenelectrontransportitstalled(i.e.,lowoxygen)
InhibitionofOxPhosleadstoaccumulationofNADH CausesfeedbackinhibitioncascadeuptoPFK1inglycoysis
RegulationofOxidativePhosphorylation
LightEnergyisConvertedtoATPinPlantChloroplasts
VariousPigmentsHarvesttheLightEnergy
Photopigmentsabsorbdifferentwavelengthsoflight
Theenergyistransferredtothephotosyntheticreactioncenter
OrganizationofLightAbsorbingMoleculesinChloroplasts
FromLightEnergytoChargeSeparation
Chloroplastscouplethetwotypesofphotosystemsfoundinbacteria
PhotosystemIIevolvesoxygen
WaterSplitting
PhotosystemIresultsinreducedNADPH
StructureofPhotosystemI
Cytochromeb6f ComplexlinksPSIIandIandtranslocatesprotonsintothelumen
Organizationofphosyntheicmachineryinthethylakoidmembrane
Locationofphosyntheicmachineryinthethylakoidmembrane
Lightinducedredoxreactionscauseacidificationoflumen
TheprotonmotiveforceacrossthethylakoidmembranedrivesthesynthesisofATP.
FlowofProtons:Mitochondria,Chloroplasts,Bacteria
Accordingtoendosymbiotictheory, mitochondriaandchloroplastsarosefromentrappedbacteria
Bacterialcytosol becamemitochondrialmatrix andchloroplaststroma
Chapter19:Summary
Thereducedcofactorspasselectronsintotheelectrontransportchaininmitochondria
Theenergyofsunlightcreateschargeseparationinthephotosyntheticreactioncomplex
Stepwiseelectrontransportisaccompaniedbythedirectionaltransportofprotonsacrossthemembraneagainsttheirconcentrationgradient
TheenergyintheelectrochemicalprotongradientdrivessynthesisofATPbycouplingtheflowofprotonsviaATPsynthasetoconformationalchangesthatfavorformationofATPintheactivesite
Inthischapter,welearned: