生物化学课件

AnOverviewofMetabolism代谢的定义生物代谢是指生物活体与外界环境不断进行的物质（包括气体、液体和固体）交换过程。合成代谢一般是指将简单的小分子物质转变成复杂的大分子物质的过程。分解代谢则是将复杂的大分子物质转变成小分子物质的过程。糖、脂和蛋白质的合成代谢途径各不相同，但是它们的分解代谢途径则有共同之处，即糖、脂和蛋白质经过一系列分解反应后都生成了酮酸并进入三羧酸循环，最后被氧化成CO2和H2O。◆◆新陈代谢的类型：新陈代谢

合成代谢（同化作用）分解代谢（异化作用）由小分子合成大分子需要能量将大分子分解成小分子释放能量1.TheRolesofmetabolismMetabolismisahighlycoordinatedanddirectedcellactivity,inwhichmanymultienzymesystemscooperatetoaccomplishfourfunctions:(l)toobtainchemicalenergybycapturingsolarenergyorbydegradingenergy-richnutrientsfromtheenvironment;(2)toconvertnutrientmoleculesintothecell'sowncharacteristicmolecules,includingmacromolecularprecursors;(3)topolymerizemonomericprecursorsintoproteins,nucleicacids,lipids,polysaccharides,andothercellcomponents;(4)tosynthesizeanddegradebiomoleculesrequiredinspecializedcellularfunctions.TheenergysupplyanddemandinHeterotrophs:

theATP-ADPcycle2.TheGeneralFeaturesofmetabolismOccursinlinear,branchedorcircularpathways;Highlyinterconnected(“EveryroadleadstoRome”).Highlyregulatedtoachievethebesteconomy(“Balancedsupplyanddemand”).Thenumberofreactionsislarge(over1000)andthenumberoftypesofreactionsisrelativelysmall.（见课本p5~15）Wellconservedduringevolution:reflectingtheunityofthelifephenomena(“whathappensinbacteriahappensinhumanbeing”).DegradationisconvergentandenergyreleasingSynthesisisdivergentandenergyconsumingThecitricacidcycle乙酰辅酶A草酰乙酸柠檬酸异戊烯焦磷酸3.Thingsthatwillbecovered1).Generalprinciplesforbioenergetics.

2).Oxidativedegradationoffuels(glycolysis,b-oxidation,ureacycle,a-ketoacidoxidation, citricacidcycle),generatingNADH,FADH2,ATP,andCO2.

3).OxidationofNADHandFADH2byO2andgenerationofATPandH2O(respiratorychains,ATPsynthase).

4).Photosynthsis(photophosphorylationandcarbonfixation).

5).Regulationofmetabolism.4.Understanding

Metabolism:aretrospect4.1

Sugardegradation(glycolysis)andsynthesis1897,EduardBuchner,Cell-freefermentation(NobelPrizein1907).OttoFritzMeyerhof,conversionofglucosetolacticacidinmuscleandbacktoglucoseinliver(NobelPrizein1922).SirArthurHardenandHansvonEuler-Chelpin,involvementofenzymes,coenzymes,andphosphorylatedintermediates(NobelPrizein1929).CarlFerdinandCoriandGertyTheresaCori,formationofglucose-1-phosphatefromglycogenandpyrophosphatebytheactionofphosphorylase(NobelPrizein1947).LuisF.Leloir,UDP-glucoseistheprecursorforglycogensynthesis(NobelPrizein1970).Thewholeglycolysispathway(conversionofglucosetopyruvate)wasrevealedby1940.4.2Completeoxidationoffuels:fromtwo-carbonunitstoCO2AlbertvonSzent-Gyorgyi,plantacids,fumaricacid(反丁烯二酸),malicacid(苹果酸)werenotconsumed,butactascatalystsforthecellularcombustionprocess(NobelPrizein1937).FritzAlbertLipmann,roleofco-enzymeAandATP(NobelPrizein1953);SirHansAdolfKrebs,Acetyl-CoAandcitricacidcycle(柠檬酸循环)forcompleteoxidationtwo-carbonunits(NobelPrizein1953).4.3SynthesisofATPusingenergyreleasedfromfueloxidationOttoHeinrichWarburg,involvementofiron-containingcytochromes(细胞色素)

incellularrespiration(NobelPrizein1931).PeterD.Mitchell,proposedthechemiosmotictheory（化学渗透学说）torelateelectronflowtoATPsynthesisinallorganisms(NobelPrizein1978).PaulD.Boyer,JohnE.Walker,enzymaticmechanismforATPsynthesis(NobelPrizein1997).4.4LipiddegradationandsynthesisKonradBlochandFeodorLynen,pathwaysforcholesterolandfattyacidsynthesis(NobelPrizein1964).MichaelS.BrownandJosephL.Goldstein,regulationofcholesterolbiosynthesis(NobelPrizein1985).4.5RegulationofMetabolismEarlW.Sutherland,Jr.,cAMPasthesecondmessagerforhormonestoregulatecellmetabolism(NobelPrizein1971).AlfredG.GilmanandMartinRodbell,involvementofmembraneG-proteinsinsignaltransductionofhormones(NobelPrizein1994).EdmondH.FischerandEdwinG.Krebs,regulationofenzymaticactivitybyreversiblephosphorylation(NobelPrizein1992).4.6PhotosynthesisHansFischer,constitutionofchlorophyll(叶绿素)

anditssimilaritytoheme(NobelPrizein1930).MelvinCalvin,CalvincycleforCO2assimilation(NobelPrizein1961).JohannDeisenhofer,RobertHuber,andHartmutMichel,3-DstructureofaPhotosyntheticreactioncenterfromapurplebacterium(NobelPrizein1988).4.7DNA,RNAandProteinsynthesisSeveroOchoaandArthurKornberg,enzymaticsynthesisofRNAandDNA(NobelPrizein1959).MarshallW.Nirenberg,HarGobindKhorana,interpretationofthegeneticcodesinproteinsynthesis(NoblePrizein1968).FrancoisJacob,AndreLwoff,andJacquesMonod,Mechanismstoswitchgenesoneandoffinprokaryotes(NobelPrizein1965).DavidBaltimore,RenatoDulbecco,andHowardMartinTemin,enzymaticRNA-dependentDNAsynthesisintumorviruses(NobelPrizein1975).BarbaraMcClintock,mobilegeneticelementsortransposons(NobelPrizein1983).SusumuTonegawa,geneticprincipleforthegenerationofantibodydiversity(NobelPrizein1987).SidneyAltmanandThomasR.Cech,RNAcatalyzedRNAprocessing(Ribozyme，核酶)(NobelPrizein1989).RichardJ.RobertsandPhilipA.Sharp,eukaryoticgenesaresplitandhavetobespliced(剪接）aftertranscription(NoblePrizein1993).GunterBlobel,intrinsicsignalsgovernproteinlocalization(NobelPrizein1999).HowtostudyBiochemistryIICompareandrelatethechemicalreactions(thesubstrates,theproductsandthetypeofconversion)enzymes,coenzymes,physiologicalroles,waysofregulationinvolved,etc.(Thismustbesimilar/relatedtothat!)Understandtheclassicalexperimentsandthoughtsthatledtotherevelationoftheknowledgedescribed(whywasoneawardedtheNobelPrize?).Beawarewiththedegreeofspeculativenessoncertainmodels(nothingis100%certaininscience).Understandtheaspectsthatneedfurtherstudies(howcouldIwinaNobelPrize?)Chapter20PrinciplesofBioenergeticsBioenergetics(生物能学）thequantitativestudyofenergytransductionsinlivingcellsandthechemicalnatureunderlyingtheseprocesses.一、BioenergeticsandThermodynamics1.Cellsneedenergytodoalltheirwork形成和保持细胞高度有序的结构(biosynthesisofmacromolecules).推动所有类型的运动(mechanicalworkandtransport).建立跨膜的离子浓度和电荷梯度.保持体温.在某些生物中产生光.The“energyindustry”(production,storageanduse)iscentraltotheeconomyofthecellsociety!2.

ChemicalenergyisthefavorableenergyformforcellstodoalltheirworkAntoineLavoisier`sinsightonanimalrespirationinthe18thcentury:itisnothingbutaslowcombustionofcarbonandhydrogen(thesamenatureasalightingcandle).(拉瓦锡1743－1794)Livingcellsaregenerallyheldatconstanttemperatureandpressure:chemicalenergy(freeenergy)hastobeusedbylivingorganisms,nothermalenergy,neithermechanicalenergyisavailabletodoworkincells.Biologicalenergytransformationobeythetwobasiclawsofthermodynamicsrevealedbyphysicistsandchemistsinthe19thcentury:energycanneitherbecreatednorbedestroyed(butconserved);energyconversionisnever100%efficient(somewillalwaysbewastedinincreasingthedisorderor“entropy”oftheuniverse).Thefreeenergyconceptofthermodynamicismoreimportanttobiochemiststhantochemists(whocanalwaysincreasethetemperatureandpressuretomakeareactiontooccur!).3.Thefreeenergy(G)conceptofbiochemicalreactionsFreeenergy(G):

在恒定的温度和压力条件下，一个生物化学反应或过程可以用来做功的能量的多少。注意：自由能强调的是反应始态与终态之间的能量变化，任何体系的绝对自由能是无法测定的。Freeenergychange(

G):

即产物与反应物之间的自由能差异。Gibbs发现:在恒定的温度和压力条件下，任何自发过程的自由能都是减少的(productsshouldhavelessfreeenergythanreactantsforareactiontooccurspontaneously,i.e.,

Ghasanegativevalue).（注意：自发性与化学反应的速率没有任何关系

）（4）生物化学中的标准自由变化(

G'o):

valueofthechangeinfreeenergyunderconditionsof298K(25oC),1atmpressure,pH7.0(chemistsusepH0,i.e.,theconcentrationofH+theyuseis1M,not10-7Masbiochemistsusehere)andinitialconcentrationsof1Mforallreactantsandproducts(exceptH+).（5）Theactualfreeenergychang(G)dependson

G'o,temperature,ratioofproductandreactantconcentrations(Q):

G=

G'o+RTlnQ（6）Enzymesonlyspeedupthermodynamicallyfavorablereactions(havinganegative

G)!（7）

Go'与K'

eq

的关系(theprimeagainindicatesitsbiochemicaltransformation):atequilibrium,

G=0,Q=K`eq,thus

Go'

=-RTlnK'eq

（8）当两个反应偶联时，

Gand

G‘o

的值具有加和性（additive）(i.e.,sharingcommonintermediates),因此，一个热力学允许的反应可以驱动一个热力学不利的反应.这样的现象在生物化学中经常遇到。Glucose+Phosphate→Glucose-6-phosphate+H2O

Go‘=+13.8KJ/molATP+H2O→ADP+Pi

Go‘=-30.5KJ/mol二、PhosphateGroupTransfersandATPATP水解的标准自由能（

Go‘

）：[ATP]=[ADP]=[Pi]=1mol/L,[H+]=10-7mol/L

Go‘=-30.5KJ/mol因此我们称ATP为高能磷酸化合物，而发生断裂的焦磷酸键称为高能键。（注意：我们所说的高能与化学中的高能键是完全不同的两个概念）2.在细胞中，ATP水解的实际自由能变化（

G）与

Go‘差别很大：(

G'o

is-30.5kJ/mol;Gincellsis-50to-65kJ/mol)Forexample,inhumanerythrocytestheconcentrationsofATP,ADP,andPiare2.25,0.25,and1.65mM,respectively,thepHis7.0andthetemperatureis25°C.ΔG=ΔG°'+RTln[ADP][Pi][ATP]=-30,500J/mol+(8.315J/mol•K)(298K)ln[(2.50•10-4)(1.65•10-3)/(2.25•10-3)]=-30,500+2,480ln(1.83•10-4)

=-51.8(kJ/mol)3.TheATPmoleculeiskineticallystableatpH7(i.e.,ithasahighactivationenergy,G‡forhydrolysis)andenzymecatalysisisneededforitshydrolysis.4.ATPisnotalong-termstorageformoffreeenergyinlivingcells,beingconsumedwithinaminutefollowingitsformation.Arestinghumanconsumesabout40kgofATPin24hours!5.其他高能磷酸化物和硫酯类化合物：6.ATPhasanintermediatephosphorylgrouptransferpotential,thusADPcanacceptandATPcandonatephosphorylgroups.ATPprovidesenergybygrouptransfer(donatingaPi,PPiorAMPtoformcovalentintermediates),notbysimplehydrolysis.ATPhasanintermediatephosphorylgrouptransferpotentialATPcantransferaPi,PPiorAMPtoareactant7.Inthelab,aslittleasafewpicomoles(10-12mol)ofATPcanbemeasuredusingfireflyluciferinandluciferase(荧光素酶),usingspectroscopicmethods.三、生物主要是通过氧化还原反应的电子转移获得生物能1.Whenelectronsflowfromalowaffinitycarrier(reductant,e.g.,glucose)toahighaffinitycarrier(oxidant,e.g.,O2),eitherinanelectricbatteryorinalivingcell,energyisreleasedandworkcanbedone.Oxidationofenergy-richbiologicalfuelsoftenmeansdehydrogenation(catalyzedbydehydrogenases,脱氢酶)fromcarbonshavingvariousoxidationstates.Inthelivingcells,electronsaretransferreddirectlyaselectrons(betweenmetalions),ashydrogenatoms(H++e-),orasahydrideion(:H-orH++2e-).生物细胞中氧化还原反应电子转移方式：以电子的形式传递：呼吸链中细胞色素之间的氧化还原反应。2.以氢原子的形式传递：以FMN或FAD为氢受体的脱氢反应。3.以氢负离子（或H++2e-）形式传递：以NADN+或NADP+为受体的脱氢反应。2.Reductionpotentials(E)measureaffinityforelectrons:Theaffinityforelectronsofacompound(initsoxidizedform)isindicatedbyitsreductionpotential(E).Standardreductionpotential(Eo')ofeachoxidant(aconstant)ismeasuredbyconnectingahalf-cellhavingtheoxidizedandreducedspeciesoftheredoxpaireachat1M,or1atmforgases,pH7toareferencehalf-cellhaving1MH+and1atmH2,whoseEo'isarbitrarilyassignedas0.00V.ApositivevalueofEo'indicatesatendencytoacquireelectronfromthereferencehalfcell(with1MH+/1atmH2).Thestandardreductionpotential(Eo')ofaconjugateredoxpairismeasuredbyconnectingthesamplehalf-celltotheH+/H2referencehalf-cell.pH7pH0WaltherNernstderivedanequationthatrelatesstandardreductionpotential(E0)toreductionpotential(E)atanyconcentrationofoxidizedandreducedspeciesinthecell:在生物化学中用Eo'取代EoEo'3.生物化学反应的自由能变化与电池电动势：Theenergymadeavailabletodoworkbythisspontaneouselectronflow(thefree-energychangefortheoxidation-reductionreaction)isproportionaltoΔE:ΔG=-nFΔEorΔGº'=-nFΔEº'Acetaldehyde+NADH+H+

→ethanol+NAD+TherelevanthalfreactionsandtheirEovaluesare:(1)Acetaldehyde+2H++2e-

→ethanol

Eº‘=-0.197V(2)NAD++2H++2e-

→NADH+H+

Eº'=-0.320VFortheoverallreaction,ΔEº'=-0.197V-(-0.320V)=0.123V,andnis2.Therefore,ΔGº'=-nFΔEº'=-2(96.5kJ/V•mol)(0.123V)=-23.7kJ/mol.5.Afewuniversalcarrierscollectelectronsfromthestepwiseoxidationofvarioussubstrates.Cellularoxidationofanutrientoccursviastepwisereactions(pathways)forefficientenergytransduction.NAD+,NADP+,FAD,andFMNareuniversalreversibleelectroncarriers(ascoenzymesofvariousenzymes).NADandNADParedinucleotidesabletoaccept/donateahydrideion(with2e-)foreachroundofreduction/oxidation.NAD(asNAD+)usuallyactsinoxidationsandNADP(asNADPH)inreductions.NADandNADPcaneasilydiffuseoutoftheenzymes,butFMNandFADaretightlyboundtotheenzymes(thusbeingcalledprostheticgroups,andthecomplexproteinsbeingcalledflavoproteins).NADHandFADH2willbefurtheroxidizedviatherespiratorychainforATPproduction.:H-QuinonoidNAD(NicotinamideAdenineDinucleotide)andNADP(NicotinamideAdenineDinucleotidePhosphate)Nicotinamide(derivedfromniacin)Benzenoid(insolution)IsoalloxazineFMN(Flavinmononucleotide)andFAD(FlavinAdenineDinucleotide)Eº'ofFAD/FMNoftendiffersindifferentflavoproteins,whichareoftencomplexandcontainotherinorganicionstohelpelectrontransfer异咯嗪环(derivedfromriboflavin)

ATP的水解和镁离子的作用Carbonshavevariousoxidationstates,withhydrocarbonbeingthemostreducedandCO2beingthemostoxidizaed.

首先，在你们这个年龄应该充满理想和抱负，对知识、科学和人生有着无限追求。所以，我觉得考研的主要目的是为了进一步提高自己的知识层次，培养从事科学研究的能力，为实现更大和更远的目标奠定基础。如果这样，你就应该选一个自己喜欢的学科专业，立志为其不懈努力，奋斗不止。

其次，可能你对学科专业的认识还很模糊，难以确定自己喜欢什么，并且近期目标就是考上研究生，那么，就根据自己的专业知识能力和英语水平，选择比较有把握考上的单位。因为三年以后如何变化和发展是很难预料的。也许，在这三年中，你对科学的认识更加深刻，业务知识和科研能力都取得了飞跃进展，到时考博和出国深造都有可能。往后的发展机遇可能更多更好。最后，如果你的目的就是为了研究生毕业后容易就业或找一份好的工作，那么，我认为生物化学和分子生物学专业最好。一是目前以及在未来一段时间内，这方面的专业人员还比较短缺，例如，近年来，我们学院来了许多研究生和博士生，但没有一个生物化学和分子生物学专业的，而我们学院急需这方面的师资和学术骨干。并且据我了解，全国普通高校都存在类似问题。二是随着生物技术的发展和应用，我国生物技术产业将迅速发展，而生物化学和分子生物学是生物技术的理论基础和技术支撑，所以未来的就业机会很多。三是生物化学和分子生物学专业比较容易转行，如你所说的健康和营养问题都离不开生物化学的理论和技术。既是到一些公司搞营销，也属生物化学专业选择面最广。

以上所述，仅是我个人的看法，还要根据你自己的情况而定。

糖酵解GlycolysisandtheCatabolismofHexosesAnoverviewonD-glucosemetabolism大多数机体的能源物质，彻底氧化：

G'o=–2840kJ/mole通过磷酸戊糖途径可产生NADPH和ribose-5-P。可以以多糖(glycogenorstarch)或转化为脂肪进行长期储存。几乎所有的生物分子都可由葡萄糖合成（aminoacids,nucleotides,fattyacids,coenzymesandothermetabolicintermediates.）MetabolismofglucoseunderAerobicandAnaerobicConditions:AnoverviewonD-glucosemetabolismTheDevelopmentofBiochemistryandtheDelineationofGlycolysisWentHandbyHand1897,EduardBuchner(Germany),accidentalobservation:sucrose(asapreservative)wasrapidlyfermentedintoalcoholbycell-freeyeastextract.Theacceptedviewthatfermentationisinextricablytiedtolivingcells(i.e.,thevitalisticdogma,活力论)wasshakenandBiochemistrywasborn:Metabolismbecamechemistry!1900s,ArthurHardenandWilliamYoungPiisneededforyeastjuicetofermentglucose,ahexosediphosphate(fructose1,6-bisphosphate)wasisolated.1900s,ArthurHardenandWilliamYoung(GreatBritain)separatedtheyeastjuiceintotwofractions:oneheat-labile,non－dialyzablezymase(enzymes)andtheotherheat-stable,dialyzablecozymase(metalions,ATP,ADP,NAD+).1910s-1930s,GustavEmbdenandOttoMeyerhof(Germany),studiedmuscleanditsextracts:Reconstructedallthetransformationstepsfromglycogentolacticacidinvitro;revealedthatmanyreactionsoflacticacid(muscle)andalcohol(yeast)fermentationswerethesame!DiscoveredthatlacticacidisreconvertedtocarbohydrateinthepresenceofO2(gluconeogenesis);observedthatsomephosphorylatedcompoundsareenergy-rich.GlycolysiswasalsoknownasEmbden-Meyerhofpathway.Thewholepathwayofglycolysis(Glucosetopyruvate)waselucidatedbythe1940s.一、糖酵解途径（Embden-Meyerhofpathway

）葡萄糖经10步酶促反应，分解为2分子丙酮酸，产生2分子ATP和2分子NADH。1.糖酵解的十步反应AnetgainoftwoATP,twoNADH,twopyruvatesareresultedwhenaglucosemoleculeisoxidizedviatheglycolysispathway:

Glucose+2ADP+2Pi+2NAD+

2pyruvate+2ATP+2H2O+2NADH+2H+2.Tenenzymescatalyzethetenreactionsofglycolysis（1）Hexokinase(alsoglucokinaseinliver)催化糖酵解的第一步反应。★Mg2+ATP2-,notATP4-

是该酶的另一个底物;★己糖激酶是一个调节酶，6-磷酸葡萄糖和ATP是该酶的变构抑制剂。★该反应热力学有利，在细胞中为不可逆反应。（2）Phosphohexoseisomerase(alsocalledphosphoglucoseisomerase)catalyzestheisomerizationfromglucose6-Ptofructose6-P,convertinganaldosetoaketose.（3）Phosphofructokinase-1(PFK-1,磷酸果糖激酶-1)催化第二步磷酸化反应。★

PFK-1是一个变构酶，催化效率很低，糖酵解途径进行的速度依赖于该酶的活性水平。★PFK-1是一个调节酶，ATP和H+是该酶的变构抑制剂，AMP可解除ATP对该酶的抑制作用，因此细胞中ATP/AMP值对此酶具有明显的调节作用。★H+

浓度升高抑制该酶的活性。★在动物细胞中，发现三种同工酶：PFKA,B,C。★TheplantPFK-1makesuseofPPi,insteadofATPatthisstep.（4）Aldolase(醛缩酶),namedforthereversereactioncatalyzesthecleavage(“lysis”)offructose1,6-bisphosphatefromthemiddleC-Cbondtoformtwo3-carbonsugars,dihydroxyacetonephosphateandglyceraldehyde3-phosphate;thisisareversalaldolcondensationreaction;thermodynamicallyveryunfavorableunderstandardconditions.（5）Triosephosphateisomerase(anextremelyefficientenzyme)convertsdihydroacetonephosphatetoglyceraldehyde3-phosphate;anintramolecularredoxreaction(ahydrogenatomistransferredfromC-1toC-3).（6）Glyceraldehyde3-phosphatedehydrogenase

催化糖酵解的唯一的氧化还原反应。★NAD+

为该酶的辅酶，从底物接受2e-。★脱氢反应的同时，底物发生磷酸化，无机磷酸直接参与反应，砷酸可取代磷酸参与该反应，但产物为3-磷酸甘油酸。★巯基为该酶的催化基团，形成硫酯中间产物，碘乙酸为是该酶的抑制剂。（7）Thephosphoglyceratekinasecatalyzesthedirecttransferoftheanhydridephosphatein1,3-BPGtoanADPtogenerateanATP;thisiscalledthesubstrate-levelphosphorylation;1,3-BPGisahighenergyintermediatethatleadstoATPformation.（8）Thephosphoglyceratemutasecatalyzestheshiftofphosphorylgroupon3-phosphoglyceratefromC-3toC-2;2,3-bisphosphoglycerateisbothacoenzymeforthemutaseandanintermediateforthereaction.（9）Enolase(烯醇酶)catalyzestheeliminationofaH2Ofrom2-phosphoglyceratetogeneratephosphoenolglycerate(PEP)withthetransferpotentialofthephosphorylgroupdramaticallyincreased(

G0`

changedfrom–17.6to–61.9kJ/mol).（10）Thepyruvatekinase(namedforthereversereaction)催化糖酵解的第二分子ATP的合成。★该酶为变构调节酶，ATP、长链脂肪酸、乙酰辅酶A、和丙氨酸为抑制剂，果糖-1，6-二磷酸为激活剂。★该反应在细胞中为不可逆反应。★该酶至少有三种不同类型的同工酶：L型（肝脏）、M型（肌肉和脑）、A型（其他组织）。二

其他单糖进入糖酵解途径三、糖酵解途径的调节Therateofglycolysisinmammalsismainlycontrolledatthestepactedbyphosphofructokinase-1(PFK-1)(Why?)(1)

PFK-1catalyzesanirreversibleexergonicreaction,whichcommitsglucosetotheglycolysispathway(awayfromthepentosephosphatepathway).(2)PFK-1isacomplextetramericenzymeregulatedbymultipleintracellularsignals(allostericeffectors):ATP,citratebeingnegativeones;AMP,ADPandfructose2,6-bisphosphateaspositiveones.(3)Aregulatedbifunctionalenzyme(PFK-2andFBPase-2)synthesizes(fromFru-6-P)anddegradesfructose2,6-bisphosphate.Afeedforwardstimulation:Fru-6-PstimulatethesynthesisandinhibitsthehydrolysisofFru-2,6-bisphosphate,whichinturnstimulatesPFK-1.2.Hexokinaseandpyruvatekinasealsosetthepaceofglycolysis(1)Thesetwoenzymesalsocatalyzedirreversibleexergonicreactions.(2)MusclehexokinaseisallostericallyinhibitedbyitsreactionproductGlc-6-P,whichaccumulateswhenPFK-1isinhibited.(3)Theliverhexokinase(alsocalledhexokinaseDorglucokinase)hasabout100XlessaffinityforglucosethanthatinmuscleandisnotinhibitedbyGlc-6-P:itsmainroleistoconvertexcessglucosetoGlc-6-Pforglycogensynthesis.(4)PyruvatekinaseisallostericallyinhibitedbyATP,

alanine,acetyl-CoA,andlong-chainfattyacids.(5)Thecatalyticactivityoftheliverpyruvatekinaseisozyme(theLtype)isalsocontrolledbyreversiblephosphorylation.四、无氧条件下，丙酮酸的去路

－－发酵ThisoccurstoregenerateNAD+fortheglycolysispathwaytocontinuewhenO2lacks.乳酸发酵：

PyruvateisreducedtolactatewhenO2lacksinareactioncatalyzedbylactatedehydrogenase(occurringinveryactiveskeletonmuscle,somebacterialikelactobacilli)2.乙醇发酵：pyruvateisfirstdecarboxylatedandthenreducedbyNADH,catalyzedbypyruvatedecarboxylaseandalcoholdehydrogenaserespectively.PresentonlyinthosealcoholfermentativeorganismsPresentinmanyorganismsincludinghumanHexokinaseGlucoseInducedfitIrreversibleincells

Onesubunitofthetetramericphosphofructokinase-1(PFK-1)RegulatoryADPThecommittingstepPhosphorolysis（磷酸解作用）C-1nolongercarriesalargepositivecharge:hydrideionleavesreadilyEnergy-richintermediate(thioester)InactiveenzymeProposedactionmechanismOfglyceraldehyde3-PdehydrogenaseAproposedactionmechanismforphosphoglyceratemutase第二十三章

三羧酸循环TricarboxylicAcidCycle一、葡萄糖的有氧分解代谢在有氧条件下，葡萄糖的分解代谢可分为三个阶段进行讨论：（1）葡萄糖到丙酮酸阶段同糖酵解途径；（2）三羧酸循环；（3）呼吸链电子传递和ATP的合成。1.Pyruvateisoxidizedtoacetyl-CoAbythecatalysisofpyruvatedehydrogenasecomplexStructureofthepyruvatedehydrogenasecomplex

ThreedimensionalimageofPDHcomplex,showingthesubunitstructure:E1,pyruvatedehydrogenase;E2,dihydrolipoyltransacetylase;andE3,dihydrolipoyldehydrogenase.Thecore(green)consists

of60moleculesofE2,arrangedin20trimerstoformapentagonal

dodecahedron.ThelipoyldomainofE2(blue)reachesoutwardto

touchtheactivesitesofE1molecules(yellow)arrangedontheE2core.

AnumberofE3subunits(red)arealsoboundtothecore,wherethe

swingingarmonE2canreachtheiractivesites.Anasteriskmarksthe

sitewherealipoylgroupisattachedtothelipoyldomainofE2.Numberoflipoyl

domainsvariesbyspecies.

2.Thecompleteoxidationofpyruvateinanimaltissueswasproposedtoundergoviaacyclicpathway(1)O2consumptionandpyruvateoxidationinmincedmuscletissueswerefoundtobestimulatedbysomefour-carbondicarboxylicacids(Fumarate,succinate,malateandoxaloacetate,five-carbondicarboxylicacid(a-ketoglutarate),orsix-carbontricarboxylicacids(citrate,isocitrate,cis-aconitate).(2)Asmallamountofanyoftheseorganicacidsstimulatesmanyfoldsofpyruvateoxidation!(3)Malonateinhibitspyruvateoxidationregardlessofwhichactiveorganicacidisadded!(4)HansKrebsproposedthe“citricacidcycle”forthecompleteoxidationofpyruvateinanimaltissuesin1937(hewronglyhypothesizedthatpyruvatecondenseswithoxaloacetateinhisoriginalproposal).(5)Thecitricacidcyclewasconfirmedtobeuniversalincellsbyinvitrostudieswithpurifiedenzymesandinvivostudieswithradioisotopes(“radioisotopetracerexperiments”).(6)KrebswasawardedtheNobelprizeinmedicinein1953forrevealingthecitricacidcycle(thusalsocalledtheKrebscycle).3.Theacetylgroup(carriedbyCoA)iscompletelyoxidizedtoCO2viathecitricacidcycleThe4-carbonoxaloacetate(草酰乙酸)actsasthe“carrier”fortheoxidation.Thetwocarbonsreleasedas2CO2inthefirstcycleofoxidationarenotfromtheacetyl-CoAjustjoined.The8electronsreleasedarecollectedbythreeNAD+andoneFAD.OnemoleculeofATP(orGTP)isproducedpercyclebysubstrate-levelphosphorylation.4.Thecitricacidcycle5.Thecompleteoxidationofoneglucosemayyieldasmanyas32ATPAlltheNADHandFADH2willeventuallypasstheirelectronstoO2afterbeingtransferredthroughaseriesofelectroncarriers.ThecompleteoxidationofeachNADHmoleculeleadstothegenerationofabout2.5ATP,andFADH2ofabout1.5ATP.Overallefficiencyofenergyconservationisabout34%usingthefreeenergychangesunderstandardconditionsandabout65%usingactualfreeenergychangesincells.6.TheregulationofTCAcycle(1)ThepyruvatedehydrogenasecomplexinvertebratesisregulatedallosetericallyandcovalentlyTheformationofacetyl-CoAfrompyruvateisakeyirreversiblestepinanimalsbecausetheyareunabletoconvertacetyl-CoAintoglucose.Thecomplex(inallorganisms)isallostericallyinhibitedbysignalingmoleculesindicatingarichsourceofenergy,e.g.,ATP,acetyl-CoA,NADH,fattyacids;activatedbymoleculesindicatingalack(ordemand)ofenergy,e.g.,AMP,CoA,NAD+,Ca2+.Theactivityofthecomplex(invertebrates,probablyalsoinplants,butnotinE.coli)isalsoregulatedbyreversiblephosphorylationofoneoftheenzymes,E1,inthecomplex:phosphorylationofaspecificSerresidueinhibitsanddephosphorylationactivatesthecomplex.Thekinaseandphosphataseisalsopartoftheenzymecomplex.ThekinaseisactivatedbyahighconcentrationofATP.(2)TherateofthecitricacidcycleiscontrolledatthreeexergonicirreversiblestepsCitratesynthase,isocitratedehydrogenaseanda-ketoglutaratedehydrogenase;Inhibitedbyproductfeedback(citrate,succinyl-CoA)andhighenergycharge(ATP,NADH);Activatedbyalowenergycharge(ADP)orasignalforenergyrequirement(Ca2+).二、三羧酸循环的回补反应Themostcommonanapleroticreactionscoverteitherpyruvateorphosphoenolpyruvatetooxaloacetateormalate.Someanaerobicbacteria,lackingthea-ketoglutaratedehydrogenaseenzyme,makebiosyntheticprecursorsviatheincompletecitricacidcycle;couldbeanearlyevolutionstageofthecitricacidcycle.

三、三羧酸循环的中间产物是重要的生物合成的前体分子Thecitricacidcycleisthehubofintermediarymetabolismservingboththecatabolicandanaboli