生物化學(xué)英文版課件：chapter 13 Overview of Metabolism and Principles of Bioenergetics

Chapter13

OverviewonmetabolismandPrinciplesofBioenergetics

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BiochemistryLecture(Nov.15,2012)ATPThestudyofMetabolismandbioenergeticsiskeytounderstandlifeAhummingbirdcanstoreenoughfueltofly900kilometerswithoutrest!Biosynthesis&BiodegradationBioenergeticsDefiningMetabolismTheword“metabolism”means"change",or"overthrow".

Biochemically,itmeans:theentirehighlyintegratedandregulatednetworkofchemicaltransformationsoccurringinalivingorganism,throughwhichcellsgrowandreproduce,maintaintheirstructures,andrespondtotheirenvironment.SanctoriusSanctorius(1561-1636)

The“weighingchair"LouisPasteur(1822-1895)Yeastfermentationcatalyzedby“ferments”.FriedrichW?hler(1800-1882)

synthesisofureaEduardBuchner(1860-1917)Cell-freefermentationHansKrebs(1900-1981)Ureacycle&citricacidcycleMetabolismisarbitrarilydividedintotwocategories:TheYing&YangofMetabolism

Catabolism

(biodegradation):Reactionsinvolvingthebreakingdownoforganicsubstrates,typicallybyoxidativebreakdown,toprovidechemicallyavailableenergy(e.g.ATP)and/ortogeneratemetabolicintermediatesusedinsubsequentanabolic

reactions.Anabolism

(biosynthesis):Theprocessesthatresultinthesynthesisofcellularcomponentsfromprecursorsoflowermolecularweight(oftenviaendergonicreactionsandthusbeingenergyconsuming).Alivingcellcarriesoutthousandsofreactionssimultaneously,witheachreactionsequencecontrolledsothatunwantedaccumulationordeficienciesofintermediatesandproductsdonotoccur!!!Organismsareclassifiedasautotrophsandheterotrophsbasedontheirmetabolicfeatures(energyandcarbon)Autotrophs(“self-feeding”)—Deriveenergyfromsunlightorinorganicsubstances,andusingCO2assolecarbonsourcetoproducecomplexorganiccompounds;Includinggreenplants,algae,andcertainbacteria;Beingthe“producers”inthefoodchain.Heterotrophs(“feedingonothers”)—Deriveenergyandcarbonfromoxidationoforganiccompounds(madebyautotrophs);Includingallanimalsandmostbacteriaandfungi;Beingthe“consumers”inthefoodchain.“Producers”“Consumers”Metabolisminvariouslivingorganismsleadtotherecyclingofcarbon,oxygeninthebiosphere.

Autotrophs&heterophsareinterdependentoneachotherinthebiosphere.Metabolismleadstothecycling

ofnitrogeninthebiosphere

Therecyclingofmatterisdrivenbytheflowofenergyinonedirectionthroughthebiosphere,i.e.,beingconstantlytransformedintounusableformssuchasheat.GeneralFeaturesofMetabolism

Occursinspecificcellularlocationsasaseriesofenzyme-catalyzedpathways.Highlycoupledandinterconnected(“EveryroadleadstoRome”).Highlyregulated

toachievethebesteconomy(“Balancedsupplyanddemand”).Thenumberofreactionsislarge,however,thenumberoftypesofreactionsisrelativelysmall(whathappensinanimalrespirationhappensinplantphotosynthesis).Wellconserved

duringevolution(“whathappensinbacteriahappensinhumanbeing”).carbohydrateAminoacidsCoenzymes(vitamines)Aminoacidshormonesnucleotideslipids22ndeditiondesignedbyDr.DonaldE.NicholsonEnergyproductionThenetworkforallthemetabolicpathways

Thebasicroadmapofcentralmetabolicpathways:occurringinthreestagesDegradative&biosyntheticpathwaysarealwaysdistinct:forthermodynamicsandregulationreasons.PolymersMonomersUltimatedegradation(乙酰輔酶A)Metabolicpathwayscouldbeconvergent,divergentorcyclic.AnabolicreactionsareHighlydivergent!CatabolicreactionsareHighlyconvergent!MostbiochemicalreactionsfallintofivegeneralcategoriesMakingorbreakingofC-Cbonds;Intramolecularrearrangements(isomerizationandelimination);Occurviafree-radicalintermediates;Grouptransfer;Oxidation-reduction.Thebiochemicalreactionshavebeenproductsofevolutionaryselectionbasedontheirrelevanceforthelifeprocess,aswellastheirrates(afterbeingcatalyzedbyproperenzymes),i.e.,notalltheorganicreactionsyoulearnedinorganicchemistryoccurinlivingcells.ThousandsofbiochemicalreactionsmightoccurinthehumanComputationalpredictioninhumans:atotalof1653metabolicenzymes,only622ofwhichwereassignedrolesin135predictedmetabolicpathways.Uncharacterizedpathways?References:Romeroetal.,(2004)“Computationalpredictionofhumanmetabolicpathwaysfromthecompletehumangenome”,GenomeBiology,6:R2.2.Smith,E.andMorowitzj,H.J.(2004)“Universalityinintermediarymetabolism”,PNAS,101:13168–13173.3.Caetano-Anolles,G.,Kim,H.S.andMittenthal,J.E.(2007)“"Theoriginofmodernmetabolicnetworksinferredfromphylogenomicanalysisofproteinarchitecture".PNAS,104:9358–63.

IssuesforcurrentandfutureinvestigationonmetabolismObservationofmetabolicprocessesinintactlivingorganisms(e.g.,inthebrainsundervariousstates)Continuetounveilnewpathwaysandnewregulationstrategiesofmetabolism.Studiesonenzymes.Metabolismdifferencesamongvariousorganismsorvariousstatesofthesameorganism(fordiagnosingandtreatingsuchdiseasesascancer,infectionsofbacteriaorviruses,obesity,etc;tounderstandaging).Appropriateandinappropriatenutrition.Biotechnologicalapplicationofknowledgelearnedfrommetabolicstudiesinmedicine,agricultureandindustry.HowoneshouldlearnaboutmetabolismCompareandrelate(interconnect)thechemicalreactions(Whereareyouinthemetabolismnetwork?)Trytocontemplateonthewaysthelivingorganismsusedtoachieveabalancedanddynamicsteadystate(Howcouldthemultilayeredregulationcooperatesoeffectively?).Understandtheclassicalexperimentsandthoughtsthatledtotherevelationoftheknowledgedescribed(Whyhe/shewontheNobelPrize?).Beawareofthenatureofthedata(Couldtheinvitroobservationsbeextendedtowhathappensinvivo?).Understandtheaspectsthatneedfurtherstudies(HowcouldIwinaNobelPrize?).BioenergeticsThequantitativestudy(mainlyusingtheprinciplesofchemicalthermodynamics)ofenergytransductionsinlivingcellsandthephysical-chemicalnatureunderlyingtheseprocesses.

BioenergeticsbeganwithearlyquantitativestudiesonanimalrespirationLavoisierusedacalorimetertoestimateheatproduced(watermeltedfromice)bytheguineapig'smetabolism:Animalrespiration(transportofO2fromairtotissues,andCO2inoppositedirection)

isnothingbutslowcombustionofcarbonandhydrogen,likethatofacandleburning(1789).Keyissues:WhereisO2convertedtoCO2andH2Oinanimals?Whatcontributesthecarbonandhydrogen?

A.Lavoisier(1743-1794)Measuringheatproduced;O2takenin;H2Oproduced;CO2produced.Usingguineapig.Ascientificunderstandingofanimalrespiration(biologicaloxidation)Locationofbiologicaloxidation:

Lung→blood→alltissues→allcellsRevelationsofthemolecularmechanism:

HowO2participates(productionofCO2andH2O);Whatenzymes(cytochromes,dehydrogenases,etc)participate;Whataretherolesofironandlightabsorbingredoxcomponents(hemegroups);Whatsubcellularlocations(mitochondria);Howtostudyquantitativelyfortheenergytransformation(athermodynamicsapproach);etc.TheEnergyconceptwasestablishedbyphysicists(19thcentury)Anabstractnumericalphysicalquantity(indirectlyobserved)ormathematicalprinciplethatindicatetheabilityofasystemtodowork.Itisnotadescriptionofamechanismoranythingconcrete.Energycanneitherbecreatednordestroyed:Itcanonlybetransformedfromoneformtoanother(Helmholtz,1847;proposedinthecontextofhisstudiesonmusclemetabolism).TheGibbsfreeenergy--theenergythatcanbeconvertedintoworkatauniformtemperatureandpressurethroughoutasystem(Gibbs,1876).J.P.Joule(1818-1889)H.vonHelmholtz(1821-1894)J.W.Gibbs(1839-1903)TheGibbsfreeenergyconceptwasappliedtostudychemicalreactionsGibbsdevelopedthechemicalthermodynamics:relatingfreeenergychangewithequilibriumconstant.

G=

G'o+RTlnQ

(Q=[products]/[reactants])

G'o=-RTlnK'eq

(K'eq:equilibriumconstant)Theactualfreeenergychange(

G)

determineswhetherareactionoccursfavorably(orspontaneously).Thestandardfreeenergychange

inbiochemistry(

G'o)isaconstant(measuredunderastandardsetofconditions).

Gforareactioncanbelarger,smaller,orthesameas

G'o,dependingontheconcentrationsofthereactantsandproducts(Q:massactionratio).J.W.Gibbs(1839-1903)

AsmallchangeinstandardfreeenergyleadstoalargechangeinequilibriumconstantLivingorganismshavetoconsumeenergyTogenerateandmaintainitshighlyorderedstructure(biosynthesis).Togeneratemotion(mechanicalwork).Togenerateconcentrationandelectricalgradientsacrosscellmembranes(activetransport).Togenerateheatandlightincertainorganisms.The“energyindustry”(production,storageanduse)iscentraltotheeconomyofthecellsociety!

Livingorganismsconsumefreeenergy

Livingcellsaregenerallyheldatconstanttemperatureandpressure:chemicalenergy(Gibbsfreeenergy,

G-“availableenergy”)hastobeusedbylivingorganisms.Livingorganismsrequireacontinualinputoffreeenergy.Biologicalenergytransformationobeythetwobasiclawsofthermodynamics.Freeenergychangeinoxidation-reductionreactionscanbecalculatedbymeasuringthereductionpotentialReductionpotential(involtsormillivolts)measuresthetendencyofachemicalspeciestoacquireelectronsandtherebybereduced.Themorepositivethepotential,thegreaterthespecies'affinityforelectronsandtendencytobereduced.Standardreductionpotential(E'o)isdefinedrelativetoareferenceelectrode.e-e-E'o=0.00VE'o=0.00VpH7NegativeE'o

pH7positiveE'o

pH0pH0Standardreductionpotentialsofbiologicallyimportanthalf-reactionshavebeensystematicallymeasured.Theactualreductionpotential(E)ofeachhalf-reactioncanbecalculatedaccordingtotheNernstequation

Theactualreductionpotential(E')dependsonthestandardreductionpotential(E'o

),electronstransferredpermolecule(n),temperature(T),ratioof[reducedform]/[oxidizedform]:[reducedform][oxidizedform]E'E'oWaltherNernst(1864-1941)

G

ofaredoxreactioncanbedirectlycalculatedfromthevalueof

E

(=E

oftheelectronacceptor–E

oftheelectrondonor):

Gcanbecalculatedfrom

EusingtheNerstEquationWhen

Eispositive,

Gisnegative.Thethermodynamicsconceptsappliedinbiochemicalstudies(since1930s)BorsookandSchott(1931)Theroleoftheenzymeinthesuccinate-enzyme-fumarateequilibrium,J.Biol.Chem.92:535-557.Borsook,H.&Schott,H.F.(1931)Thefreeenergy,heat,andentropyofformationofl-malicacid.J.Biol.Chem.92:559-567.

Reductionpotentials,equilibriumconstants,heatcapacitiesmeasured,freeenergy,entropycalculated.Thethermodynamicconceptswereappliedinstudyingthesynthesisofproteins(1930s)BorsookandHuffman,(1938).Somethermodynamicalconsiderationsofaminoacids,peptides,andrelatedsubstances,in"ChemistryoftheAminoAcidsandProteins"(C.L.A.Schmist,editor)C.C.Thomas,Springfield,Ill.ItwasoriginallythoughtthesynthesisshouldoccurbyMassActioninreversal,ascatalyzedbyproteases,glycogenphosphorylases,andpolynucleotidephosphorylases.Itwaslaterrealizedthatthefreeenergychangeinhydrolysiswassolargethatonecouldnotgetsynthesisbyanyfeasibledegreeofconcentrationofaminoacids.Energyisneededtobeputintothesystem,viaacoupledreaction.Amountoffreeenergyexpendedtoformthepeptidebondsorphosphodiesterbondsinvivoisfarhigherthantheirstandardfreeenergyofformationinvitro,tobuyspecificityofthebondsformed!Adirectsourceofenergyformusclecontractionwassearched!Non-lacticmusclecontractionsattheexpenseofthedephosphorylationofcreatinephosphate(1930)

Lundsgaard,E.,Biochem.Z.217,162;227,51(1930).Frogmusclespoisonedwithiodoacetate(unabletosplitglucosetolacticacid)arecapableofcarryingoutcontractions！Aparallelbreakdownofcreatinephosphateobservedinthepresenceofiodoacetate.LundsgaardLacticaciddoesnotserveasanenergysourceformusclecontraction!CreatinephosphateArapidlymobilizablereserveofhigh-energyphosphatesinskeletalmuscleandthebrain.ATPdiscoveredinmuscle(1929)Lohmann,(1928)UederdasWorkommenundUmsatzvonPyrophosphatinderZelle,Biochem.Z,202:466-493;203:164-207.FiskeandSubbarow(1929)PhosphorusCompoundsofMuscleandLiver,Science,70:381–382.Langen&Hucho(2008)KarlLohmannandtheDiscoveryofATP,AngewandteChemie,47:1824-1827.

FromFiskeandSubbarowpaper.MyosinwasfoundtobeanATPaseEngelhardtWA,LiubimovaMN.(1939)Myosineandadenosinetriphosphatase,Nature,144:688

AcidificationtopHbelow4rapidlydestroysthisactivity;completelylostafter10minat37oC,butthepresentofATPstabilizesit.Engelhardt(1941):ThefreeenergycouplingandATPenergycurrencytheoriesproposed(1941)Lipmann,F.(1941).“MetabolicGenerationandUtilizationofPhosphateBondEnergy”.AdvancesinEnzymologyandRelatedSubjects,1:99-162.Kalckar,H,(1941).“TheNatureofEnergeticCouplinginBiologicalSynthesis”Chem.Rev.28:71-178.

“Energy-richphosphate”(likeATP)proposedtodriveenergy-requiringbiologicalprocesses(e.g,Musclecontraction,transportofionsandothermoleculesacrossmembranes,chemicalreactionforthebiosynthesisofproteinsandnucleicacids).Thebiologicaloxidoreduction(respiration)iscompulsorilycoupledtophosphorylation.

LipmannKalckarATPistheuniversalcurrencyforbiologicalenergyThiswasfirstperceivedbyFritzLipmannandHermanKalckarin1941whenstudyingglycolysis.Hydrolysisofthetwophosphoanhydride(磷酸酐鍵)bondsinATPgeneratemorestableproductsreleasinglargeamountoffreeenergy(

G'o

is-30.5kJ/mol;Gpincellsis-50to-65kJ/mol).TheATPmoleculeiskineticallystableatpH7andenzymecatalysisisneededforitshydrolysis.ATPactuallyexistsasasumofvariousspeciesincells(e.g.,ATP4-andMgATP2-).FritzLipmann(1899-1986)

Fig.2.Thetwo-dimensionalstickmodeloftheadenosinephosphatefamilyofmolecules,showingtheatomandbondarrangement.Thecommerceofthecell,metabolism,usesATPasthecommonenergycurrency.GTP,CTP,UTPhassimilarbondingwhytheywerenotchosenbynature?Thehumanbodyonaveragecontainsonly250gramsofATPbutturnsoveritsownbodyweightequivalentinATPeachday!ATPprovidesenergyusuallythroughgrouptransfer(proteincouldalsobesuchacceptors)GlnsynthetaseK’eq=10-

G'o/1.36

Thereactionisthusacceleratedbyabout105fold!ATPmightbeconsideredasa“coenzyme”inthissense.Conceptofcoupledreactionsformulated(1900)

WilhelmOstwald,(1900)Z.Physik.Chem.34:248Thefreeenergyreleasedbyanexergonicprocesscanbeusedtodriveanendergonicprocessthatwouldnotgobyitself.“Wasteno(free)energy;useitwell".NucleophilicattacksNotphosphateATPusuallyprovidesenergybygrouptransferofphosphorylgroups(磷?；?-PO32-),notphosphategroups(-OPO32-),

formingcovalentintermediates,notbysimplehydrolysis.Forenergytobesupplied,thetwoprocesseshavetobecoupled!ATPsuppliesenergyforallkindsofcellularprocessesATPhasanintermediatephosphorylgrouptransferpotential,thusADPcanacceptandATPcandonatephosphorylgroups(formingtheATP-ADPcycleandactingasanenergycurrency)WhydoesATPhaveahighphosphoryltransferpotential?G0’dependsonthedifferenceinfreeenergiesofproductsandreactants,therefore,bothmustbeconsidered

;Thereisnosuchthingas“highenergybond”.Fourfactorsareimportant:1.Reliefofchargerepulsion;2.Resonancestabilization;3.Ionization;4.Stabilizationduetohydration.ATPisnotalong-termstorageformoffreeenergyinlivingcells,butphosphocreatineisonesuchphosphorylreservoir,orso-calledphosphagen(alsoinorganicpolyphosphate).Biologicalenergywasfoundtobeproducedviaoxidation-reductionreactions(i.e.,electrontransferring)

MetabolicfuelsCO22e-

2e-2e-2e-H2OO2ATPFreeenergyOxidationMetabolicfuelsareoxidizedtoCO2,withelectronstransferredfirsttouniversalcarriers(e.g.,NAD+andFAD),andeventuallytoO2.EnergyisreleasedduringsuchredoxreactionsandeventuallyconservedinATP.(NADH,FADH2)TransmembraneProtongradientFunctionalgroupsinorganiccompoundspresentinoneoffourgeneraloxidationstates,equivalenttoalkane,alcohol,ketone,orcarboxylicacid

Inaerobicorganisms,theultimateelectronacceptorintheoxidationofcarbonisO2,andtheoxidationproductisCO2Nicotinamideadeninedinucleotide(NAD+)wasfoundtobeacommoncofactorforhydrogen-transferringenzymes(1906)

Harden&Young(1906)."TheAlcoholicFermentofYeast-Juice".Proc.Roy.Soc.78:369–375.