TheCitricAcidCycleOxidizesTwoCarbonUnits

TheconversionofpyruvateintoacetylCoAbythepyruvate
dehydrogenasecomplexisthelinkbetweenglycolysisand
cellularrespirationbecauseacetylCoAisthefuelforthecitric
acidcycle.
Thecitricacidcyclebeginswiththecondensationofafour
carbonunit,oxaloacetate,andatwocarbonunit,theacetylgroup
ofacetylCoA.
OxaloacetatereactswithacetylCoAandH

2Otoyieldcitrate
andCoA.

Thisreaction,whichisanaldolcondensationfollowedbya
hydrolysis,iscatalyzedbycitratesynthase.
OxaloacetatefirstcondenseswithacetylCoAtoformcitryl
CoA,amoleculethatisenergyrichbecauseitcontainsthe
thioesterbondthatoriginatedinacetylCoA.
ThehydrolysisofcitrylCoAthioestertocitrateandCoA
drivestheoverallreactionfarinthedirectionofthe
synthesisofcitrate.
Itisveryimportantthatsidereactions,notablythehydrolysisof
acetylCoAtoacetateandCoA,beminimized.Letusbriefly
considerhowthecitratesynthasepreventsthewasteful
hydrolysisofacetylCoA.
Xraycrystallographicstudiesofcitratesynthaseandits
complexeswithseveralsubstratesandinhibitorsrevealedthatthe
enzymeundergoeslargeconformationalchangesinthecourseof
catalysis.
Citratesynthaseexhibitssequential,orderedkinetics:

oxaloacetatebindsfirst,followedbyacetylCoA.Thereasonfor
theorderedbindingisthatoxaloacetateinducesamajor
structuralrearrangementleadingtothecreationofabindingsite
foracetylCoA.Thebindingofoxaloacetateconvertstheopen
formoftheenzymeintoaclosedform
Smalldomainrotates19degreesrelativetothelargedomain.
o Thesestructuralchangescreateabindingsiteforacetyl
CoA.
Citratesynthasecatalyzesthecondensationreactionbybringing
thesubstratesintocloseproximity,orientingthem,andpolarizing
certainbonds
ThedonationandremovalofprotonstransformsacetylCoAinto
anenolintermediate.
1. Theenolattacksoxaloacetatetoformacarboncarbon
doublebondlinkingacetylCoAandoxaloacetate.
2. ThenewlyformedcitrylCoAinducesadditionalstructural
changesintheenzyme,causingtheactivesitetobecome
completelyenclosed.
3. TheenzymecleavesthecitrylCoAthioesterbyhydrolysis.
4. CoAleavestheenzyme,followedbycitrate,andthe
enzymereturnstotheinitialopenconformation.
CitratesynthaseiswellsuitedtohydrolyzecitrylCoAbutnot
acetylCoA
1. First,acetylCoAdoesnotbindtotheenzymeuntil
oxaloacetateisboundandreadyforcondensation.
2. Second,thecatalyticresiduescrucialforthehydrolysisof
thethioesterlinkagearenotappropriatelypositioneduntil
citrylCoAisformed.
o

Citrateisisomerizedintoisocitrate

Thehydroxylgroupisnotproperlylocatedinthecitratemolecule
fortheoxidativedecarboxylationsthatfollow.Thus,citrateis
isomerizedintoisocitratetoenablethesixcarbonunittoundergo
oxidativedecarboxylation.

Theisomerizationofcitrateisaccomplishedbyadehydration
stepfollowedbyahydrationstep.Theresultisaninterchange
ofanHandanOH.

Aconitaseisanironsulfurprotein
o Itsfourironatomsarecomplexedtofourinorganicsulfides
andthreecysteinesulfuratoms,leavingoneironatom

availabletobindcitratethroughoneofitsCOO groupsand
anOHgroup
o ThisFeSclusterparticipatesindehydratingand
rehydratingtheboundsubstrate.

Isocitrateisoxidizedanddecarboxylatedtoalphaketoglutarate

Firstoffouroxidationreductionreactionsinthecitricacidcycle.
Theoxidativedecarboxylationofisocitrateiscatalyzedby
isocitratedehydrogenase.

Theintermediateinthisreactionisoxalosuccinate,anunstableb
ketoacid.Whileboundtotheenzyme,itlosesCO2toforma
ketoglutarate.

Therateofformationofaketoglutarateisimportantin
determiningtheoverallrateofthecycle.
Thisoxidationgeneratesthefirsthightransferpotentialelectron
carrier,NADH,inthecycle.

SuccinylcoenzymeAisformedbytheoxidativedecarboxylationof
alphaketoglutarate

Theconversionofisocitrateintoaketoglutarateisfollowedbya
secondoxidativedecarboxylationreaction,theformationof
succinylCoAfromaketoglutarate.

Thisreactioniscatalyzedbytheaketoglutarate
dehydrogenasecomplex,anorganizedassemblyofthreekindsof
enzymesthatishomologoustothepyruvatedehydrogenase
complex.

Bothreactionsincludethedecarboxylationofanaketoacidand
thesubsequentformationofathioesterlinkagewithCoAthathas
ahightransferpotential.

Acompoundwithhighphosphoryltransferpotentialisgenerated
fromsuccinylcoenzymeA

Inthecitratesynthasereaction,thecleavageofthethioesterbond
powersthesynthesisofthesixcarboncitratefromthefour
carbonoxaloacetateandthetwocarbonfragment.
ThecleavageofthethioesterbondofsuccinylCoAiscoupledto
thephosphorylationofapurinenucleosidediphosphate,usually
ADP.Thisreaction,whichisreadilyreversible,iscatalyzedby
succinylCoAsynthetase(succinatethiokinase).

Thisreactionistheonlystepinthecitricacidcyclethatdirectly
yieldsacompoundwithhighphosphoryltransferpotential.
o Intissuesthatperformlargeamountsofcellularrespiration,
suchasskeletalandheartmuscle,theADPrequiring
isozymepredominates.
o Intissuesthatperformmanyanabolicreactions,suchasthe
liver,theGDPrequiringenzymeiscommon.
o TheGDPrequiringenzymeisbelievedtoworkinreverse
ofthedirectionobservedintheTCAcycle;thatis,GTPis
usedtopowerthesynthesisofsuccinylCoA,whichisa
precursorforhemesynthesis.

Mechanism:SuccinylcoenzymeAsynthetasetransformstypesof
biochemicalenergy

Themechanismofthisreactionisaclearexampleofanenergy
transformation:energyinherentinthethioestermoleculeis
transformedintophosphorylgrouptransferpotential

Oxaloacetateisregeneratedbytheoxidationofsuccinate

Reactionsoffourcarboncompoundsconstitutethefinalstageof
thecitricacidcycle:theregenerationofoxaloacetate.

Amethylenegroup(CH2)isconvertedintoacarbonylgroup
(CPO)inthreesteps:anoxidation,ahydration,andasecond
oxidationreaction.Oxaloacetateistherebyregeneratedfor
anotherroundofthecycle,andmoreenergyisextractedinthe
formofFADH2andNADH.

Succinateisoxidizedtofumaratebysuccinatedehydrogenase.
+
ThehydrogenacceptorisFADratherthanNAD ,whichisused
intheotherthreeoxidationreactionsinthecycle.FADisthe
hydrogenacceptorinthisreactionbecausethefreeenergy
+
changeisinsufficienttoreduceNAD .FADisnearlyalwaysthe
electronacceptorinoxidationsthatremovetwohydrogenatoms
fromasubstrate

Succinatedehydrogenase,likeaconitase,isanironsulfur
protein.
Infact,succinatedehydrogenaseisdirectlyassociatedwiththe
electrontransportchain,thelinkbetweenthecitricacidcycle
andATPformation.FADH2producedbytheoxidationof
succinatedoesnotdissociatefromtheenzyme,incontrastwith
NADHproducedinotheroxidationreductionreactions.Rather,
twoelectronsaretransferredfromFADH2directlytoironsulfur
clustersoftheenzyme,whichinturnpassestheelectronsto
coenzymeQ(CoQ).CoenzymeQ,animportantmemberofthe
electrontransportchain,passeselectronstotheultimateacceptor,
molecularoxygen,
ThenextstepisthehydrationoffumaratetoformLmalate.
+
FumarasecatalyzesastereospecifictransadditionofH and

OH .TheOH groupaddstoonlyonesideofthedoublebondof
fumarate;hence,onlytheLisomerofmalateisformed.

Finally,malateisoxidizedtoformoxaloacetate.Thisreactionis
+
catalyzedbymalatedehydrogenase,andNAD isagainthe
hydrogenacceptor.

Theoxidationofmalateisdrivenbytheuseoftheproducts
oxaloacetatebycitratesynthaseandNADHbytheelectron
transportchain

Thecitricacidcycleproduceshightransferpotentialelectrons,
ATP,andCO2

Thenetreactionofthecitricacidcycleis
o

1.

2.

3.

4.

Twocarbonatomsenterthecycleinthecondensationofan
acetylunit(fromacetylCoA)withoxaloacetate.Twocarbon
atomsleavethecycleintheformofCO2inthesuccessive
decarboxylationscatalyzedbyisocitratedehydrogenaseanda
ketoglutaratedehydrogenase.
Fourpairsofhydrogenatomsleavethecycleinfouroxidation
+
reactions.TwoNAD moleculesarereducedintheoxidative
decarboxylationsofisocitrateandaketoglutarate,oneFAD
moleculeisreducedintheoxidationofsuccinate,andone
+
NAD moleculeisreducedintheoxidationofmalate.Recall
+
alsothatoneNAD moleculeisreducedintheoxidative
decarboxylationofpyruvatetoformacetylCoA
Onecompoundwithhighphosphoryltransferpotential,
usuallyATP,isgeneratedfromthecleavageofthethioester
linkageinsuccinylCoA.
Twowatermoleculesareconsumed:oneinthesynthesisof

citratebythehydrolysisofcitrylCoAandtheotherinthe
hydrationoffumarate.
Isotopelabelingstudiesrevealedthatthetwocarbonatoms
thatentereachcyclearenottheonesthatleave.Thetwo
carbonatomsthatenterthecycleastheacetylgroupare
retainedduringtheinitialtwodecarboxylationreactionsand
thenremainincorporatedinthefourcarbonacidsofthecycle.
Notethatsuccinateisasymmetricmolecule.Consequently,
thetwocarbonatomsthatenterthecyclecanoccupyanyof
thecarbonpositionsinthesubsequentmetabolismofthefour
carbonacids.Thetwocarbonsthatenterthecycleasthe
acetylgroupwillbereleasedasCO2insubsequenttrips
throughthecycle.
TheelectrontransportchainoxidizestheNADHandFADH2
formedinthecitricacidcycle.Thetransferofelectronsfrom
thesecarrierstoO2,theultimateelectronacceptor,leadstothe
generationofaprotongradientacrosstheinnermitochondrial
membrane.Thisprotonmotiveforcethenpowersthe
generationofATP;
Consequently,ninehightransferpotentialphosphorylgroups
aregeneratedwhentheelectrontransportchainoxidizes3
NADHmoleculesand1FADH2molecule,andonehigh
transferpotentialphosphorylgroupisdirectlyformedinone
roundofthecitricacidcycle.Thus,oneacetylunitgenerates
approximately10moleculesofATP.
Recallthatmolecularoxygendoesnotparticipatedirectlyin
thecitricacidcycle.However,thecycleoperatesonlyunder
+
aerobicconditionsbecauseNAD andFADcanbe
regeneratedinthemitochondriononlybythetransferof
electronstomolecularoxygen.Glycolysishasbothanaerobic
andananaerobicmode,whereasthecitricacidcycleis
strictlyaerobic

Lipids

Membranesareasdiverseinstructureastheyarein
function.However,theydohaveincommonanumberof
importantattributes:
1. Membranesaresheetlikestructures,onlytwomolecules
thick,whichformclosedboundariesbetweendifferent
compartments.Thethicknessofmostmembranesis
between60(6nm)and100(10nm).
2. Membranesconsistmainlyoflipidsandproteins.The
massratiooflipidstoproteinsrangesfrom1:4to4:1.
Membranesalsocontaincarbohydratesthatarelinkedto
lipidsandproteins.
3. Membranelipidsaresmallmoleculesthathaveboth
hydrophilicandhydrophobicmoieties.Theselipids
spontaneouslyformclosedbimolecularsheetsin
aqueousmedia.Theselipidbilayersarebarrierstothe
flowofpolarmolecules.
4. Specificproteinsmediatedistinctivefunctionsof
membranes.Proteinsserveaspumps,channels,
receptors,energytransducers,andenzymes.Membrane
proteinsareembeddedinlipidbilayers,whichcreate
suitableenvironmentsfortheiraction.
5. Membranesarenoncovalentassemblies.Theconstituent
proteinandlipidmoleculesareheldtogetherbymany
noncovalentinteractions,whichactcooperatively.
6. Membranesareasymmetric.Thetwofacesofbiological
membranesalwaysdifferfromeachother.
7. Membranesarefluidstructures.Lipidmoleculesdiffuse
rapidlyintheplaneofthemembrane,asdoproteins,
unlesstheyareanchoredbyspecificinteractions.In
contrast,lipidmoleculesandproteinsdonotreadily
rotateacrossthemembrane.Membranescanberegarded
astwodimensionalsolutionsoforientedproteinsand
lipids.
8. Mostcellmembranesareelectricallypolarized,such
thattheinsideisnegative[typically260millivolts

(mV)].Membranepotentialplaysakeyroleintransport,
energyconversion,andexcitability
Thepropertiesoffattyacidsandoflipidsderivedfromthem
aremarkedlydependentonchainlengthanddegreeof
saturation.
Unsaturatedfattyacidshavelowermeltingpointsthando
saturatedfattyacidsofthesamelength.
Chainlengthalsoaffectsthemeltingpoint,
Thus,shortchainlengthandunsaturationenhancethefluidity
offattyacidsandoftheirderivatives.
Lipidsarewaterinsolublebiomoleculesthatarehighly
solubleinorganicsolventssuchaschloroform.
Lipidshaveavarietyofbiologicalroles:
o Theyserveasfuelmolecules,
o Highlyconcentratedenergystores,
o Signalmolecules
o Messengersinsignaltransductionpathways,and
componentsofmembranes.
Thethreemajorkindsofmembranelipidsarephospholipids,
glycolipids,andcholesterol.

Phospholipidsarethemajorclassofmembranelipids

Aphospholipidmoleculeisconstructedfromfourcomponents:
oneormorefattyacids,aplatformtowhichthefattyacidsare
attached,aphosphate,andanalcoholattachedtothephosphate.
Thefattyacidcomponentsprovideahydrophobicbarrier,
whereastheremainderofthemoleculehashydrophilicproperties
thatenableinteractionwiththeaqueousenvironment.
Theplatformonwhichphospholipidsarebuiltmaybeglycerol,a
threecarbonalcohol,orsphingosine,amorecomplexalcohol.
Phospholipidsderivedfromglycerolarecalled
phosphoglycerides.
o Aphosphoglycerideconsistsofaglycerolbackbonesto
whichareattachedtwofattyacidchainsanda
phosphorylatedalcohol.

Inphosphoglycerides,thehydroxylgroupsatC1andC2of
glycerolareesterifiedtothecarboxylgroupsofthetwofattyacid
chains.
TheC3hydroxylgroupoftheglycerolbackboneisesterifiedto
phosphoricacid.
Whennofurtheradditionsaremade,theresultingcompoundis
phosphatidate(diacylglycerol3phosphate),thesimplest
phosphoglyceride.
Onlysmallamountsofphosphatidatearepresentinmembranes.
However,themoleculeisakeyintermediateinthebiosynthesis
oftheotherphosphoglycerides.
Themajorphosphoglyceridesarederivedfromphosphatidateby
theformationofanesterbondbetweenthephosphategroupof
phosphatidateandthehydroxylgroupofoneofseveralalcohols.
Thecommonalcoholmoietiesofphosphoglyceridesarethe
aminoacidserine,ethanolamine,choline,glycerol,andinositol

Thesecondmajorclassofmembranelipids,glycolipids,are
sugarcontaininglipids.
Theaminogroupofthesphingosinebackboneisacylatedbya
fattyacid,asinsphingomyelin.Glycolipidsdifferrom
sphingomyelinintheidentityoftheunitthatislinkedtothe
primaryhydroxylgroupofthesphingosinebackbone.In
glycolipids,oneormoresugars(ratherthanphosphorylcholine)
areattachedtothisgroup.
Cholesterol,thethirdmajortypeofmembranelipid,hasa
structurethatisquitedifferentfromthatofphospholipids.Itisa
steroid,builtfromfourlinkedhydrocarbonrings.

Ahydrocarbontailislinkedtothesteroidatoneend,anda
hydroxylgroupisattachedattheotherend.Inmembranes,the
orientationofthemoleculeisparalleltothefattyacidchainsof
thephospholipids,andthehydroxylgroupinteractswiththe
nearbyphospholipidheadgroups.
However,theselipidspossessacriticalcommonstructural
theme:membranelipidsareamphipathicmolecules(amphiphilic
molecules).Amembranelipidcontainsbothahydrophilicanda
hydrophobicmoiety.
Thetwohydrophobicfattyacidchainsareapproximatelyparallel
toeachother,whereasthehydrophilicphosphorylcholinemoiety
pointsintheoppositedirection.
Thehydrophilicunit,alsocalledthepolarheadgroup,is
representedbyacircle,andthehydrocarbontailsaredepictedby
straightorwavylines
Membraneformationisaconsequenceoftheamphipathicnature
ofthemolecules.Theirpolarheadgroupsfavorcontactwith
water,whereastheirhydrocarbontailsinteractwithoneanother
inpreferencetowater.
Onewayistoformaglobularstructurecalledamicelle.
Thepolarheadgroupsformtheoutsidesurfaceofthemicelle,
whichissurroundedbywater,andthehydrocarbontailsare
sequesteredinside,interactingwithoneanother
Alipidbilayerisalsocalledabimolecularsheet.
Thehydrophobictailsofeachindividualsheetinteractwithone
another,formingahydrophobicinteriorthatactsasa
permeabilitybarrier.
Thehydrophilicheadgroupsinteractwiththeaqueousmedium
oneachsideofthebilayer.

Thefavoredstructureformostphospholipidsandglycolipidsin
aqueousmediaisabimolecularsheetratherthanamicelle.
Thereasonisthatthetwofattyacidchainsofaphospholipidora
glycolipidaretoobulkytofitintotheinteriorofamicelle.
Theformationofbilayersinsteadofmicellesbyphospholipidsis
ofcriticalbiologicalimportance.Amicelleisalimitedstructure,
usuallylessthan200(20nm)indiameter.
Incontrast,abimolecularsheetcanextendtomacroscopic
7
6
dimensions,asmuchasamillimeter(10 ,or10 nm)ormore.
Lipidbilayersformspontaneouslybyaselfassemblyprocess.In
otherwords,thestructureofabimolecularsheetisinherentinthe
structureoftheconstituentlipidmolecules.Thegrowthoflipid
bilayersfromphospholipidsisrapidandspontaneousinwater.
Hydrophobicinteractionsarethemajordrivingforceforthe
formationoflipidbilayers.
Watermoleculesarereleasedfromthehydrocarbontailsof
membranelipidsasthesetailsbecomesequesteredinthe
nonpolarinteriorofthebilayer.
Furthermore,vanderWaalsattractiveforcesbetweenthe
hydrocarbontailsfavorclosepackingofthetails.
Finally,thereareelectrostaticandhydrogenbondingattractions
betweenthepolarheadgroupsandwatermolecules.Thus,lipid
bilayersarestabilizedbythefullarrayofforcesthatmediate
molecularinteractionsinbiologicalsystems.
Becauselipidbilayersareheldtogetherbymanyreinforcing,
noncovalentinteractions(predominantlyhydrophobic),theyare
cooperativestructures.
Thesehydrophobicinteractionshavethreesignificantbiological
consequences:
1. (1)lipidbilayershaveaninherenttendencytobeextensive;
2. (2)lipidbilayerswilltendtocloseonthemselvessothat
therearenoedgeswithexposedhydrocarbonchains,andso
theyformcompartments;and
3. (3)lipidbilayersareselfsealingbecauseaholeinabilayer
isenergeticallyunfavorable.

Lipidbilayersarehighlyimpermeabletoionsandmostpolar
molecules

Lipidbilayermembraneshaveaverylowpermeabilityfor
ionsandmostpolarmolecules.
Thepermeabilityofsmallmoleculesiscorrelatedwiththeir
solubilityinanonpolarsolventrelativetotheirsolubilityin
water.
Thisrelationsuggeststhatasmallmoleculemighttraversea
lipidbilayermembraneinthefollowingway:
a. First,itshedsitssolvationshellofwater;
b. Then,itisdissolvedinthehydrocarboncoreofthe
membrane;and,
c. Finally,itdiffusesthroughthiscoretotheotherside
ofthemembrane,whereitbecomesresolvatedby
water.

ThermodynamicsofMembraneTransport

Sodiumionspassthroughspecificchannelsinthehydrophobic
barrierformedbymembraneproteins.
Thismeansofcrossingthemembraneiscalledfacilitated
diffusionbecausethediffusionacrossthemembraneisfacilitated
bythechannel.
Itisalsocalledpassivetransportbecausetheenergydrivingthe
ionmovementoriginatesfromtheiongradientitself,withoutany
contributionbythetransportsystem.
Proteintransportersembeddedinthemembranearecapableof
usinganenergysourcetomovethemoleculeupaconcentration
gradient.
Becauseaninputofenergyfromanothersourceisrequired,this
meansofcrossingthemembraneiscalledactivetransport.
Anunequaldistributionofmoleculesisanenergyrichcondition
becausefreeenergyisminimizedwhenallconcentrationsare
equal.
Thefreeenergychangeintransportingthisspeciesfromside1,

whereitispresentataconcentrationofc1,toside2,whereitis
presentatconcentrationc2,is
o

G=RT ln ( c 2/c 1)
3

whereRisthegasconstant(8.315310 kJmol deg ,or

3
1
1
1.987310 kcalmol deg )andTisthetemperaturein
kelvins.
Forachargedspecies,theunequaldistributionacrossthe
membranegeneratesanelectricalpotentialthatalsomustbe
consideredbecausetheionswillberepelledbythelikecharges.
Electrochemicalpotentialormembranepotential.
o

G=RT ln ( c 2/c 1)+ ZF V

InwhichZistheelectricalchargeofthetransported
species,DVisthepotentialinvoltsacrossthemembrane,
1
1
andFistheFaradayconstant(96.5kJV mol ,or23.1
1
1
kcalV mol ).
Atransportprocessmustbeactivewhen Gispositive,
Itcanbepassivewhen Gisnegative.
o

OxidativePhosphorylation

Webeginourstudyofoxidativephosphorylationbyexamining
theoxidationreductionreactionsthatallowtheflowofelectrons
fromNADHandFADH2tooxygen.
Theelectronflowtakesplaceinfourlargeproteincomplexesthat
areembeddedintheinnermitochondrialmembrane,together
calledtherespiratorychainortheelectrontransportchain.

TheresultingunequaldistributionofprotonsgeneratesapH
gradientandatransmembraneelectricalpotentialthatcreatesa
protonmotiveforce.ATPissynthesizedwhenprotonsflowback
tothemitochondrialmatrixthroughanenzymecomplex.

Thus,theoxidationoffuelsandthephosphorylationofADPare
coupledbyaprotongradientacrosstheinnermitochondrial
membrane.
Collectively,thegenerationofhightransferpotentialelectrons
bythecitricacidcycle,theirflowthroughtherespiratorychain,
andtheaccompanyingsynthesisofATPiscalledrespirationor
cellularrespiration.
Theprimaryfunctionofthecitricacidcyclewasidentifiedasthe
generationofNADHandFADH2bytheoxidationofacetylCoA.
Inoxidativephosphorylation,electronsfromNADHandFADH2
areusedtoreducemolecularoxygentowater.
o Electrontransportchain.

Theelectrontransferpotentialofanelectronismeasuredasredox
potential

Inoxidativephosphorylation,theelectrontransferpotentialof
NADHorFADH2isconvertedintothephosphoryltransfer
potentialofATP.
Themeasureofphosphoryltransferpotentialisalreadyfamiliar
tous:itisgivenby G ' forthehydrolysisoftheactivated
phosphorylcompound.Thecorrespondingexpressionforthe
electrontransferpotentialisE0,thereductionpotential(also
calledtheredoxpotentialoroxidationreductionpotential).
Thus,astrongreducingagent(suchasNADH)ispoisedto
donateelectronsandhasanegativereductionpotential,whereas
astrongoxidizingagent(suchasO2)isreadytoacceptelectrons
andhasapositivereductionpotential.

A1.14voltpotentialdifferencebetweenNADHandmolecularoxygen

driveselectrontransportthroughthechainandfavorstheformationof
aprotongradient

Thedrivingforceofoxidativephosphorylationistheelectron
transferpotentialofNADHorFADH2relativetothatofO2.
Thereleasedenergyisinitiallyusedtogenerateaprotongradient
thatisthenusedforthesynthesisofATPandthetransportof
metabolitesacrossthemitochondrialmembrane.
ElectronsaretransferredfromNADHtoO2throughachainof
threelargeproteincomplexescalledNADHQoxidoreductase,
Qcytochromecoxidoreductase,andcytochromecoxidase
Electronflowwithinthesetransmembranecomplexesleadstothe
transportofprotonsacrosstheinnermitochondrialmembrane.
Afourthlargeproteincomplex,calledsuccinateQreductase,
containsthesuccinatedehydrogenasethatgeneratesFADH2in
thecitricacidcycle.
ElectronsfromthisFADH2entertheelectrontransportchainat
Qcytochromeoxidoreductase.
SuccinateQreductase,incontrastwiththeothercomplexes,does
notpumpprotons.
NADHQoxidoreductase,succinateQreductase,Qcytochrome
coxidoreductase,andcytochromecoxidasearealsocalled
ComplexI,II,III,andIV,respectively.

Twospecialelectroncarriersferrytheelectronsfromone
complextothenext.
ThefirstiscoenzymeQ(Q),alsoknownasubiquinonebecauseit
isaubiquitousquinoneinbiologicalsystems.
o Ubiquinoneisahydrophobicquinonethatdiffusesrapidly
withintheinnermitochondrialmembrane.
ElectronsarecarriedfromNADHQoxidoreductasetoQ
cytochromecoxidoreductase,thesecondcomplexofthechain,
bythereducedformofQ.
ElectronsfromtheFADH2generatedbythecitricacidcycleare
transferredfirsttoubiquinoneandthentotheQcytochromec
oxidoreductasecomplex.
IncontrastwithQ,thesecondspecialelectroncarrierisaprotein.
Cytochromec,asmallsolubleprotein,shuttleselectronsfromQ
cytochromecoxidoreductasetocytochromecoxidase,thefinal
componentinthechainandtheonethatcatalyzesthereduction
ofO2.

ThehighpotentialelectronsofNADHentertherespiratorychainat
NADHQoxidoreductase

TheelectronsofNADHenterthechainatNADHQ
oxidoreductase(alsocalledComplexIandNADH
dehydrogenase).
NADHQoxidoreductaseisLshaped,withahorizontalarm
lyinginthemembraneandaverticalarmthatprojectsintothe
matrix.
Thereactioncatalyzedbythisenzymeappearstobe
o

TheinitialstepisthebindingofNADHandthetransferofits
twohighpotentialelectronstotheflavinmononucleotide(FMN)
prostheticgroupofthiscomplextogivethereducedform,
FMNH2
TheelectronacceptorofFMN,theisoalloxazinering,isidentical
withthatofFAD.
ElectronsarethentransferredfromFMNH2toaseriesofiron
sulfurclusters,thesecondtypeofprostheticgroupinNADHQ
oxidoreductase.

FeSclustersinironsulfurproteins(alsocallednonhemeiron
proteins)playacriticalroleinawiderangeofreduction
reactionsinbiologicalsystems.

NADHQoxidoreductasecontainsboth2Fe2Sand4Fe4S
2+
clusters.IronionsintheseFeScomplexescyclebetweenFe
3+
(reduced)andFe (oxidized)states.
o Unlikequinonesandflavins,ironsulfurclustersgenerally
undergooxidationreductionreactionswithoutreleasingor
bindingprotons.
NADHtransfersitstwoelectronstoFMN.
o TheseelectronsflowthroughaseriesofFeScentersand
thentocoenzymeQ.
o TheflowoftwoelectronsfromNADHtocoenzymeQ
throughNADHQoxidoreductaseleadstothepumpingof
fourhydrogenionsoutofthematrixofthemitochondrion.
o Inacceptingtwoelectrons,Qtakesuptwoprotonsfrom
thematrixasitisreducedtoQH2.

UbiquinolistheentrypointforelectronsfromFADH2offlavoproteins

FADH2enterstheelectrontransportchainatthesecondprotein
complexofthechain.
Succinatedehydrogenase,acitricacidcycleenzyme,ispartof
thesuccinateQreductasecomplex(ComplexII),aintegral
membraneproteinoftheinnermitochondrialmembrane.
FADH2doesnotleavethecomplex.
o Rather,itselectronsaretransferredtoFeScentersandthen
finallytoQtoformQH2,whichthenisreadytotransfer
electronsfurtherdowntheelectrontransportchain.
o ThesuccinateQreductasecomplex,incontrastwith
NADHQoxidoreductase,doesnotpumpprotonsfromone
sideofthemembranetotheother.

Electronsflowfromubiquinoltocytochromec
throughQcytochrome

c
oxidoreductase

TheelectronsfromQH2arepassedontocytochromecbythe
secondofthethreeprotonpumpsintherespiratorychain,Q

cytochromecoxidoreductase(alsoknownasComplexIIIandas
cytochromereductase).
ThefunctionofQcytochromecoxidoreductaseistocatalyzethe
transferofelectronsfromQH2tooxidizedcytochromec(Cytc),
awatersolubleprotein,andconcomitantlypumpprotonsoutof
themitochondrialmatrix.
Theflowofapairofelectronsthroughthiscomplexleadstothe
+
effectivenettransportof2H tothecytoplasmicside,halfthe
yieldobtainedwithNADHQreductasebecauseofasmaller
thermodynamicdrivingforce.
Qcytochromecoxidoreductaseitselfcontainstwotypesof
cytochromes,namedbandc1.
Acytochromeisanelectrontransferringproteinthatcontainsa
hemeprostheticgroup.Theironionofacytochromealternates
betweenareducedferrous(12)stateandanoxidizedferric(13)
stateduringelectrontransport.

TheQcyclefunnelselectronsfromatwoelectroncarriertoaone
electroncarrierandpumpsprotons

QH2passestwoelectronstoQcytochromecoxidoreductase,but
theacceptorofelectronsinthiscomplex,cytochromec,can
acceptonlyoneelectron.
ThemechanismforthecouplingofelectrontransferfromQto
cytochromectotransmembraneprotontransportisknownasthe
Qcycle
TwoQH2moleculesbindtothecomplexconsecutively,each
+

givinguptwoelectronsandtwoH .Theseprotonsarereleased
tothecytoplasmicsideofthemembrane.
1. ThefirstQH2toexittheQpoolbindstothefirstQbinding
site(Qo),anditstwoelectronstravelthroughthecomplexto
differentdestinations.
a. Oneelectronflows,first,totheRieske2Fe2Scluster;

then,tocytochromec1;
c. and,finally,toamoleculeofoxidizedcytochromec,
convertingitintoitsreducedform.
i. Thereducedcytochromecmoleculeisfreetodiffuse
awayfromtheenzymetocontinuedownthe
respiratorychain.
Thesecondelectronpassesthroughtwohemegroupsof
cytochromebtoanoxidizedubiquinoneinasecondQbinding
site(Qi).
ThenowfullyoxidizedQleavesthefirstQsite,freetoreenter
theQpool.
AsecondmoleculeofQH2bindstotheQositeofQ
cytochromecoxidoreductaseandreactsinthesamewayas
thefirst.
Theremovalofthesetwoprotonsfromthematrixcontributes
totheformationoftheprotongradient.
b.

2.

3.

InoneQcycle,twoQH2moleculesareoxidizedtoformtwo
Qmolecules,andthenoneQmoleculeisreducedtoQH2.
Thecytochromebcomponentofthereductaseisinessencea
recyclingdevicethatenablesbothelectronsofQH2tobeused
effectively.

Cytochromec
oxidasecatalyzesthereductionofmolecularoxygento
water

Thelastofthethreeprotonpumpingassembliesofthe
respiratorychainiscytochromecoxidase(ComplexIV).
Cytochromecoxidasecatalyzesthetransferofelectronsfrom
thereducedformofcytochromectomolecularoxygen,the
finalacceptor.
FourelectronsarefunneledtoO2tocompletelyreduceitto

1.

H2O,and,concomitantly,protonsarepumpedfromthematrix
tothecytoplasmicsideoftheinnermitochondrialmembrane.
Asmuchofthisfreeenergyaspossiblemustbecapturedin
theformofaprotongradientforsubsequentuseinATP
synthesis.
CytochromecoxidasecontainstwohemeAgroupsandthree
copperions,arrangedastwocoppercenters,designatedAand
B.Onecenter,CuA/CuA,containstwocopperionslinkedby
twobridgingcysteineresidues.Thiscenterinitiallyaccepts
electronsfromreducedcytochromec.Theremainingcopper
ion,CuB,iscoordinatedbythreehistidineresidues,oneof
whichismodifiedbycovalentlinkagetoatyrosineresidue.
+
ThecoppercentersalternatebetweenthereducedCu
2+
(cuprous)formandtheoxidizedCu (cupric)formasthey
acceptanddonateelectrons.
TherearetwohemeAmolecules,calledhemeaandhemea3,
incytochromecoxidase.HemeAdiffersfromthehemein
cytochromecandc1inthreeways:
a. (1)aformylgroupreplacesamethylgroup,
b. (2)aC17hydrocarbonchainreplacesoneofthevinyl
groups,and
c. (3)thehemeisnotcovalentlyattachedtotheprotein.
Hemeaandhemea3havedistinctredoxpotentialsbecause
theyarelocatedindifferentenvironmentswithincytochrome
coxidase.AnelectronflowsfromcytochromectoCuA/CuA,
tohemeatohemea3toCuB,andfinallytoO2.
Fourmoleculesofcytochromecbindconsecutivelytothe
enzymeandtransferanelectrontoreduceonemoleculeofO2
toH2O.
Electronsfromtwomoleculesofreducedcytochromecflow
downanelectrontransferpathwaywithincytochromec
oxidase,onestoppingatCuBandtheotherathemea3.With
bothcentersinthereducedstate,theytogethercannowbind

2.

3.

4.

anoxygenmolecule.
Asmolecularoxygenbinds,itabstractsanelectronfromeach
ofthenearbyionsintheactivecentertoformaperoxide
2
(O2 )bridgebetweenthem.
Twomoremoleculesofcytochromecbindandrelease
electronsthattraveltotheactivecenter.Theadditionofan
+
electronaswellasH toeachoxygenatomreducesthetwo
2+
ionoxygengroupstoCuB OHandFe3+OH.
+

ReactionwithtwomoreH ionsallowsthereleaseoftwo
moleculesofH2Oandresetstheenzymetoitsinitial,fully
oxidizedform.
a.
b.

5.

Thefourprotonsinthisreactioncomeexclusivelyfrom
thematrix.Thus,theconsumptionofthesefourprotons
contributesdirectlytotheprotongradient.
Cytochromecoxidaseusesthisenergytopumpfour
additionalprotonsfromthematrixtothecytoplasmicsideof
themembraneinthecourseofeachreactioncycleforatotal
ofeightprotonsremovedfromthematrix

o HighenergyelectronsintheformofNADHandFADH2are
generatedbythecitricacidcycle.Theseelectronsflowthroughthe
respiratorychain,whichpowersprotonpumpingandresultsinthe

reductionofO2.