生物氧化 Biological Oxidation課件

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

指線粒體內(nèi)膜中按一定順序排列的一系列具有電子傳遞功能的酶復(fù)合體，可通過(guò)連鎖的氧化還原將代謝物脫下的電子最終傳遞給氧生成水。這一系列酶和輔酶稱為氧化呼吸鏈(respiratorychain)又稱電子傳遞鏈(electrontransferchain)。氧化呼吸鏈概念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

異咯嗪核黃素核糖醇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(醌類)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(可移動(dòng)鐵硫蛋白).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)：標(biāo)準(zhǔn)氧化還原電位E0(V)高的組分對(duì)電子親和力強(qiáng)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氧化磷酸化偶聯(lián)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(轉(zhuǎn)子):c-subunitinthemembraneareattachedtotheshaftcontainingtheγandεofF1Rotortoturn:Protonsflowthroughtheα-andc-subunitsofF0

ATPsynthesis:conformationalchangesintheβ-subunitsF0domainofATPsynthase

Centralcylinder

csubunits

Externalpart

asubunit

RedlineprotonpathThebindingchangemodel(結(jié)合變構(gòu)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)乙酰輔酶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(魚(yú)藤酮)、piericidinA

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

.3.OxidativephosphorylationinhibitorComplexIIinhibitor:carboxin（萎銹靈）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（解偶聯(lián)蛋白UCP1）棕色脂肪MechanismofUncoupler

（Brownadiposetissuemitochondria

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

Oligomycin(寡霉素)canbindtoF0,Dicyclohexylcarbodiimide(二環(huán)己基碳二亞胺，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