生物氧化 Biological Oxidation课件

BiologicalOxidation生物氧化Biologicaloxidationistheprocessinwhichsubstances(carbohydrate,Lipid,AAs)areoxidizedinlivingorganism,ultimatelyformingCO2

、H2Oandreleasingenergy.(cellularrespiratory)CarbohydratelipidsProteinsCO2+H2OO2energyADP+PiATPHeatenergyConceptofBiologicaloxidation

Similaritiesofbiologicaloxidation(invivo)

andexternaloxidation(invitro)

Oxidationtypes—Oxygenation、Dehydrogenation、Electronlost；

FollowthethegeneralruleofREDOXreactionTheoxygenconsumedbyoxidation、finalproduct(CO2andH2O)andreleaseenergyarethesameinvivoandinvitrooxidation.Reactionenvironmentmild(37℃,pH7.4)Enzymaticreactiongradually;

Energygraduallyreleaseandeasytocapture.ATPproductionefficiency.Addwatertoindirectgainoxygenbydehydrogenationreaction;TakeoffhydrogencombineswithoxygenorganicproduceH2O;DecarboxylationproduceCO2

DifferencesofbiologicaloxidationandexternaloxidationBiologicaloxidationExternaloxidation

SuddenreleaseenergyThecarbonandhydrogendirectmaterialcombinedgeneratingCO2andH2Ooxygen

ThegeneralprocessofbiologicaloxidationSection1

TheOxidativeRespiratoryChainisComposedbyElectron-carryingProteinComplexes

TheOxidativerespiratorychainalsoknowaselectron-transferchain(ETC),isasequenceoftransferringelectronsfromsubstratetomolecularoxygeninareobiccellsbyelectron-carryingproteinsembeddedintheinnermitochondrialmembrane.Thisprocessincludesaseriesofoxidative-reductionreactionsandreleasessufficientenergytodrivethesynthesisofATP.Oxidative

RespiratoryChainDefinitionhydrogencarrierandelectroncarrier

（2H2H++2e）Composition

指线粒体内膜中按一定顺序排列的一系列具有电子传递功能的酶复合体，可通过连锁的氧化还原将代谢物脱下的电子最终传递给氧生成水。这一系列酶和辅酶称为氧化呼吸链(respiratorychain)又称电子传递链(electrontransferchain)。氧化呼吸链概念Enzymecomplexisanaturalformofmitochondrialinnermembraneofrespiratorychain,eachcomponentcontainedspecificelectrontransferability.TheenergyreleasingofthisprocessdrivenH+outofmitochondriamembraneandchangeintotrans-membraneH+gradient,andthenbeusedforthebiosynthesisofATP.（一）OxidativerespiratorychainiscomposedoffourkindsofcomplexwithelectrontransferabilityComplexEnzymeNameMass(kD)NumberofsubunitsProstheticGroupsCombiningSiteComplexINADH-CoQReductase85039FMN,Fe-SNADH（matrixside）CoQ（lipidscore）ComplexIISuccinate-CoQReductase1404FAD,Fe-SSuccinate（matrixside）CoQ（lipidscore）ComplexIIICoQ-CytcReductase25011HemesbL,bH,c1Fe-SCytc(intermembrancespace)Cyt-c131HemecCytc1，CytaComplexIVCytcOxidase16213Hemesa,a3;CuA,CuBCytc(intermembrancespace)Theubiquinoneisnotincludedintheabovefourcomplexmitochondrialrespiratorychaincomplexes

ⅣCytcoxNADH+H+NAD+1/2O2+2H+H2OcytosolmatrixMitochondrialinnermembrane

ⅠQH2Q

Ⅱfumaric

acid

succinate4H+4H+Ⅲ2H+4H+CytcoxCytcredCytcred2H+2H+Thepositionofelectron-transportchaincomplexinthemitochondrialmembraneOrderofRespiratoryChainComplexesThecomponents&functionoftheeachelectron-carriercomplexesinthesetworespiratorychainNAD+andNADP+R=H:NAD+;R=H2PO3:NADP+

NAD+（NADP+）和NADH（NADPH）Mutualtransformation

REDOXreactionchangesoccurringinpentavalentnitrogenandtrivalentnitrogen，belongtodualelectrontransportbody

FMNandFAD

异咯嗪核黄素核糖醇FMNstructurecontainsriboflavin,functiongroupisisoalloxazinering,unstableintermediateproductisFMNH·,

showthreekindsofmoleculestate

inreversibleREDOXreaction,belongtosingle,dualelectrontransportbody.•FMNFMNH•FMNH2Theprostheticgroupofiron-sulfurproteinisFe-Scenterwhichcontaining

equivalenceironatomsandsulfuratoms.Transferelectronby

Fe2+Fe3++eBelongtosingleelectrontransferbody

ⓈInorganicsulfur

iron-sulfurproteins

SS

inorganicsulfurCys-sTheubiquinone(泛醌，CoQ)isakindofliposolublequinone(醌类)compoundwithlongermultipleisoprenesidechain

（humanCoQ10）,REDOXreactioncangenerateintermediateproductsemiquinoneradical.

Theubiquinone(CoQ)issingle,dualelectrontransportbody

,transferreductionequivalentandelectronbyrecruitingandshuttlinginvariouscomplex,playsacentralroleinthemobileelectrontransferandprotonmovingcoupling.

Cytochrome(Cyt)TheCytochromes(Cyt)

arekindsofsingleelectrontransferproteinscontaininghemeprostheticgroup,classifiedbydifferentabsorptionspectra.

Fe2+

Fe3++e

ComplexⅠalsocalledtheNADH-CoQreductaseComplexⅠelectrontransferorder

:

NADH→FMN→Fe-S→CoQ→Fe-S→CoQComplex

Ⅰhaveprotonpumpfunction:everytransfertwoelectroncompanywithfourH+pumpedfrommitochondrialinnermembranetointermembrancespace

1、ComplexⅠ:transferelectronofNADH+H+toubiquinoneFunctionofComplexINADH+H+

NAD+

FMNFMNH2ReducedFe-S

OxidizedFe-S

QQH2complexII

issuccinatedehydrogenaseofTAC,

alsocalledsuccinate-ubiquinonereductase.Electrontransfer

Order：

Succinate→FAD→Fe-S→CoQcomplexIIwithoutprotonpumpfunction.2、ComplexII:Transfereletronsfromsuccinatetoubiquinonesuccinatefumarate

FADFADH22Fe2+-S2Fe3+-SQQH23、ComplexIII:TransferelectronfromReducedubiquinonetoCytcComplexIIIalsocalledubiquinol-cytochromecreductase,orcytochromeb,c1complex,containstwohemes-b562andb566,andcytochromec1,andtheRieskeprotein(可移动铁硫蛋白).TheubiquinonerecruitreductionequivalentandelectronfromcomplexⅠ,Ⅱ,andtransferthemtocomplexⅢ.Electrontransfer

Order：

CoQH2→(CytbL→CytbH)→Fe-S→Cytc1→CytcElectrontransferofComplexIIIby“Qcycle”.Fourprotonsaretranslocatedacrossthemembraneforeverytwoelectronsbetransferredtocytc，ComplexⅢalsohaveprotonpumpfunction

.Cytcisthe

onlyonewater-solubleglobularproteinofrespiratory

chain,notincludedincomplexandtransferelectrontocomplexⅣ.ComplexⅢQH2→→Cytcb562;b566;Fe-S;c1ComplexⅣalsocalled

cytochromecoxidaseElectrontransfer

Order：

Cytc→CuA→Cyta→Cyta3–CuB→O2Subunit1:Cyta,Cyta3–CuB

Subunit2：CuA×2

Subunit3:pumpedtwoprotonsacrossthemembrane.

Subunit1and2formbinuclearcenterandtransferelectrontoO2

4、ComplexIV:TransfereletronfromCytctoO2PathwaysofElectronTransferThroughComplexIVEachoftheintermediatesformedinthereductionofO2remainstightlyboundtothebinuclearcenteruntilwaterisproducedwhichcanlargelypreventedfromcelldamage.2H+2H2OThepositionofelectron-transportchaincomplexinthemitochondrialmembraneStandardOxidation-ReductionpotentialApartandrestructuring

Specificinhibitorsblocking

Slowlysupplyoxygentorespiratorychain

（二）Thecomponentsofrespiratorychainarragementfromlowtohighaccordingoxidation-reductionpotential.Standardoxidation-reductionpotential

ofelectroncarriersinrespiratorychainRedoxcoupleE0(V)RedoxcoupleE0(V)NAD+/NADH+H+－0.32Cytc1Fe3+/Fe2+0.22FMN/FMNH2－0.219CytcFe3+/Fe2+0.254FAD/FADH2－0.219CytaFe3+/Fe2+0.29CytbL(bH)Fe3+/Fe2+0.05(0.10)Cyta3Fe3+/Fe2+0.35Q10/Q10H20.061/2O2/H2O0.816E0(V)：标准氧化还原电位E0(V)高的组分对电子亲和力强1、NADHRespiratoryChainNADH→ComplexⅠ→CoQ→ComplexⅢ→Cytc→ComplexⅣ→O22、SuccinateRespiratoryChainSuccinate→ComplexⅡ→CoQ→ComplexⅢ→Cytc→ComplexⅣ→O2Section2OxidativePhosphorylation:RespiratorychainreleaseenergycouplingwithADPphosphorylationtogenerateATPOxidativephosphorylation:ElectrontransfercouplingADPphosphorylationtogenerateATPSubstratelevelphosphorylation:Dehydrogenationreaction,generatingthehigh-energybondcouplingADP(GDP)generateATP(GTP)1.ThecouplingsiteofoxidativephosphorylationiscomplexⅠ,Ⅲ,ⅣThemethodsforestimateofthecouplingsites:P/OratiosFree-EnergyChanges

:⊿Gº=-nF⊿EºSubstateCompositionofrespirarychainP/OratioNumberofATPNAD+→ComplexⅠ→CoQ→ComplexⅢ2.52.5→Cytc→ComlexⅣ→O2ComplexⅡ→CoQ→ComplexⅢ1.71.5→Cytc→ComplexⅣ→O2Cytc→ComplexⅣ→O20.881(1)P/Oratio

isameasureofthenumberofATPmoleculesformedduringthetransferoftwoelectronsthroughallorpartoftheelectrontransportchain.P/OratiosofsomesubstratesPyruvatesuccinate

ascorbicacidFAD+→ATPATPATP氧化磷酸化偶联部位NADHFMN(Fe-S)琥珀酸FAD(Fe-S)CoQCytb→Cytc1→CytcCytaa3O2(2)Free-EnergyChangepH7.0StandardFreeEnergyChange(△G0)StandardReductionPotentialChange(△E0)：△G0=-nF△E0n:electronicnumber;F:Faradayconstants(96.5kJ/mol·V)∆E0E0∆G0pyruvatesuccinateVitCCouplingsitesofOxidativePhosphorylationADP+PiATP(needenergy30.5kJ/mol)Atotalof10H+areejectedfromthemitochondrialmatrixper2e-transferredfromNADHtooxygenviatherespiratorychain.

ComplexⅠ(4H+)、Ⅲ(4H+)andⅣ(2H+)haveprotonspumpfunction.2.ProtonconcentrationgradientservesastheenergyreservoirfordrivingATPformationChemiosmotichypothesisThechemiosmotichypothesisistheconceptthataprotonconcentrationgradientservesastheenergyreservoirfordrivingATPformationandwasoriginallyformulatedbyPeterMitchellintheearly1960s.Oxidativephosphorylationdependentonclosedmitochondrialmembrane

completely；Mitochondrialmembraneisnotpermeation

toH+,OH-

、K+,

Cl-ions；Electronictransmissionchaincandriveprotonsremovedfrommitochondria,formingadeterminationofthetransmembraneelectrochemicalgradient

；IncreasetheacidityoflateralmitochondrialmembranecanleadtoATPsynthesis,howeveraddsomesubstancewhichincreasepermeationofprotonscanreduceendomembraneprotonsgradientandreduceATPsynthesis.

Chemicalpenetrationhypothesishasbeenwidelyexperimentalsupportmatrix

Mt.innermembrane

++++----H+O2H2OH+e-ADP+PiATPChemiosmotichypothesissimpleschematic:3.ThesynthesisofATPusingenergyoftheprotongradientasprotonsflowthroughtheATPsynthaseF1:

Hydrophilicpartα3β3γδεsubunitscomplex,OSCP,IF1subunitboundtotheinnermitochondrialmembrane，catalyzesthesynthesisofATP.

F0：Hydrophobicpartab2c9~12subunits

Embeddedinthemitochondrialmembrane,formingtransmembraneprotonchannel.ATPSynthaseStructureofATPSynthaseATPsynthase,arotatingmolecularmotorStator(定子):a,bsubunitsofF0,alongwiththeα-andβ-subunitsandtheδ-subunitRotor(转子):c-subunitinthemembraneareattachedtotheshaftcontainingtheγandεofF1Rotortoturn:Protonsflowthroughtheα-andc-subunitsofF0

ATPsynthesis:conformationalchangesintheβ-subunitsF0domainofATPsynthase

Centralcylinder

csubunits

Externalpart

asubunit

RedlineprotonpathThebindingchangemodel(结合变构模型)forATPsynthesisbyATPsynthase

βsubunitconformation:Open(O):noactivity,LowaffinitywithATPLoose(L):noactivity,bindingwithADP+PiTense(T):ATPsynthesisactivity,bindingwithATP4protonsareneededfor1ATPsynthesisNADHRespiratoryChain10/4=2.5ATPFADRespiratoryChain

6/4=1.5ATP4.ATPplaysacentralroleinenergygeneration、utilization、transferandstoragehighenergyphosphatebondTheStandardFreeEnergyChange(△G0)releasedbyhydrolysisreactionmorethan25kJ/mol,ordinaryrepresentationPhighenergyphosphatecompound

Thecompoundscontaininghigh-energy

phosphatebonds化合物△G0kJ/mol(kcal/mol)磷酸烯醇式丙酮酸－61.9(－14.8)氨基甲酰磷酸－51.4(－12.3)1，3-二磷酸甘油酸－49.3(－11.8)磷酸肌酸－43.1(－10.3)ATP→ADP＋Pi－30.5(－7.3)乙酰辅酶A－31.5(－7.5)ADP→AMP＋Pi－27.6(－6.6)焦磷酸－27.6(－6.6)1-磷酸葡萄糖－20.9(－5.0)Examplesofhigh-energycompoundsATP+UDPADP+UTPATP+CDPADP+CTPATP+GDPADP+GTP

ADP+ADPATP+AMPEffectofnucleoside-diphosphatekinaseEffectofadenylatekinaseATP:theperfectenergycurrencyforcellCreatinephosphate(CP)isthestorageformofhigh-energy

compoundsCPisusedinnerveandmuscleforstorageof~Pbonds.

ProductionandapplicationofATPSection3Internalandexternalfactorsinfluenceoxidativephosphorylationfunction1.ADPisamajoradjusmentfactoroftheoxidativephosphorylationrate.ATP/ADPAnabolismCatabolism

Allostericregulation2.

ThyroidhormonestimulatestheoxygenconsumptionandheatproductionandincreaseThyroidhormoneinduceNa+,K+-ATPenzymesynthesis,ATPutilizationisincreased.

respiratorycontrolratio(呼吸控制率，RCR)(1)RespiratoryChaininhibitorblockedtheprocessofelectrontransferComplexIinhibitor:

rotenone(鱼藤酮)、piericidinA

(粉蝶霉素A)amobarbital(异戊巴比妥)etc.Blockingtransmissionelectrontotheubiquinone

.3.OxidativephosphorylationinhibitorComplexIIinhibitor:carboxin（萎锈灵）ComplexIIIinhibitor:

antimycinA(抗霉素A)BlockingtransmissionelectronfromCytbHtotheQNsite)myxothiazol(粘噻唑)

BlockingQPsite

ComplexIVinhibitor:

CN－、N3－

bindtooxidized-Cyta3

CObindtoreduced-Cyta3

BlockingtransmissionelectronfromCytatoCuB-Cyta3.

NADHFMN(Fe-S)SuccinateacidFAD(Fe-S)CoQCytb→Cytc→CytcCytaa3O2rotenonepiericidinA

amobarbital

×antimycinAmyxothiazol×CO、CN-、N3-andH2S×Electrontransferandbindingsitesforspecificinhibitors

carboxin×(2)UncouplerdestroytransmembraneelectrochemicalprotongradientTheenergyconservedasapropongradientwasreleasedasheat,inhibitATPsynthesis.ProtonsflowintomitochondriamatrixnotthroughATPsyntheasepathway.Forexample:dinitrophenol(二硝基苯酚，DNP）uncouplingprotein（解偶联蛋白UCP1）棕色脂肪MechanismofUncoupler

（Brownadiposetissuemitochondria

）ⅢⅠⅡF0F1ⅣCytcQcytosolmatrixUncouplerHeatengeryH+H+ADP+PiATP(3)ATPsynthaseinhibitorsuppresselectrontransferandATPsynthesisForexample:

Oligomycin(寡霉素)canbindtoF0,Dicyclohexylcarbodiimide(二环己基碳二亚胺，DCCP)covalentlybindtocunitsofF0,

inhibit

H+backflowwhichincreasedprotonelectrongradientandtheninduceelectrontransferinhibition.OligomycinoligomycinModelofATPSynthaseCanpreventprotonsbackflowfromF0protonschannelandrestrainATPproduction.pyruvateADPsuccinateMt.rotenoneoligomycinantimycinA(CN-)timeDNPInfluenceofdifferentsubstratesandinhibitorsonquantityconsumingoxygen4.MitochondrialDNAmutationscanaffectthebodyoxidativephosphorylationfunctionMt.DNAencodes37gene7unitsofcomplexⅠ1unitofcomplexⅢ3unitofcomplexⅣ2unitofATPsyntheaseMt.DNAmutationcausedegenerativediseasessuchasParkinsondisease,Alzheimer’sdiseaseetc.5.Mitochondrialmembraneselectivetransportvarioussubstance

Mitochondrialoutermembranehashighlypermeability,theinnermembraneisquiteimpermeabletomostofmoleculesandions.Permeableto：

Pyr,succinate,α-ketoglutarate,malate,citrite,Gluetc.Impermeableto：

H+,NADH,NADPH,OAA(草酰乙酸)etc.

(dependentonvarioustransporter)transportersentermitochondriaoutmitochondriaATP-ADPtranslocaseADP3-ATP4-PhosphatestransporterH2PO4-+H+dicarboxylicacidtransporterHPO42-Malateα-Ketoglutarate

transporterMalateα-KetoglutarateGlutamate-AspartatetransporterGlutamateAspartatemonocarboxylicacid

transporterPyruvate

OH-tricarboxylicacid

transporterMalateCitratebasicaminoacid

transporterOrnithineCitrullineCarnitinetransporterFattyacylcarnitine

CarnitineSomeofthetransportersinmitochondrialmembrane（1）CytosolicNADHentermitochondrialbyshuttlesystem

α-GlycerophosphateshuttleMalate-Asparateshuttle

Shuttlesystem：-Glyce