生物化学2012-05-04 柠檬酸循环

Questions1、葡萄糖是如何彻底氧化分解提供能量的？2、当纯粹以淀粉为食物时，动物如何获得合成蛋白质和核酸的基本构件分子——氨基酸和核苷酸的？第二讲柠檬酸循环（KrebsCycle）一、柠檬酸循环的准备阶段二、柠檬酸循环的循环阶段三、葡萄糖彻底氧化分解的能量计算四、柠檬酸循环的调控五、柠檬酸循环的意义六、柠檬酸循环的中间物的补充一、柠檬酸循环的准备阶段•丙酮酸形成乙酰-CoACH3CCOO-+NAD+CoA-SH+CH3C-SCoA+CO2pyruvatedehydrogenasecomplexPyruvateCoenzymeAAcetyl-CoAOO+NADH（丙酮酸脱氢酶）（二氢硫辛酰转乙酰基酶）（二氢硫辛酸脱氢酶）丙酮酸脱氢酶复合体（PDH）一、柠檬酸循环的准备阶段PDH特点：–反应发生在线立体（mit）中，mit内膜上有丙酮酸转运酶–反应由丙酮酸脱氢酶复合物（PDH）催化，二氢硫辛酰转乙酰基酶含-SH，受砷化物抑制–∆G°’=-33.4kJ•mol-1，反应为放能反应–反应需要的6种辅助因子：NAD+,FAD,Mg2+,thiaminepyrophosphate（TPP）,coenzymeA（CoA-SH）,andlipoicacid（硫辛酸）–PDH受产物别构调节和共价调节的协同控制，产物调节物有：NADH/NAD+、乙酰-CoA/CoA-SH一、柠檬酸循环的准备阶段PDH的磷酸化共价调节物一、柠檬酸循环的准备阶段Cellsinarestingmetabolicstate–needandusecomparativelylittleenergy–highATP,lowADPimplyhighATP/ADPratio–highNADH,lowNAD+implyhighNAHDH/NAD+ratioCellsinanactivemetabolicstate–needandusemoreenergythanrestingcells–lowATP,highADPimplylowATP/ADPratio–lowNADH,highNAD+implylowNAHDH/NAD+ratio一、柠檬酸循环的准备阶段二、柠檬酸循环的循环阶段•Step1:草酰乙酸（OAA）的缩合;∆G°’=-32.8kJ•mol-1柠檬酸合酶是别构酶，是Krebs循环的限速调节酶，受NADH、ATP、succinyl-CoA调节。CH3C-SCoAAcetyl-CoAC-COO-CH2-COO-Oxaloacetate+C-COO-HOCH2-COO-CH2-COO-+CoA-SHCoenzymeAcitratesynthaseCitrateOOTheCitricAcidCycle•Step2:柠檬酸经脱水、再水化异构化为异柠檬酸–citrateisachiral（非手性的）;ithasnostereocenter–isocitrateischiral;ithas2stereocenters–Aconitase（乌头酸酶）canbeinhibitedbyNOC-COO-HOCH2-COO-CH2-COO-CitrateC-COO-CH2-COO-C-COO-HCH-COO-CH2-COO-AconitateHOIsocitrateCH-COO-TheCitricAcidCycleTheCitricAcidCycle•Step3:异柠檬酸的氧化脱羧–异柠檬酸脱氢酶（IDH）为别构调节酶，被ATP和NADH抑制、ADP和NAD+激活–CO2是来自乙酰-COA?C-COO-HCH-COO-CH2-COO-HOIsocitrateC-COO-HC-COO-CH2-COO-C-HHC-COO-CH2-COO-NADHNAD+-KetoglutarateCO2isocitratedehydrogenaseOOOxalosuccinate异柠檬酸的其他去路？异柠檬酸的其他去路——乙醛酸循环（theglyoxylatecycle）植物和微生物（不是动物细胞）中异柠檬酸可经另一条途径——乙醛酸循环产生四碳酸，并能最终生成葡萄糖。–乙醛酸循环发生在乙醛酸循环体中；–乙醛酸循环与柠檬酸循环和糖异生途径相关；–乙醛酸循环需要2种特殊的酶异柠檬酸裂解酶和苹果酸合酶。GlyoxylateCycleC-COO-CH-COO-CH2-COO-IsocitrateCH2-COO-CH2-COO-SuccinateisocitratelyaseHHO+O=C-COO-GlyoxylateHC-COO-GlyoxylateCH3C-SCoAAcetyl-CoACoA-SHCoenzymeACH-COO-OHCH2-COO-malatesynthetase++MalateOOHGlyoxylateCycle乙醛酸循环体线粒体胞质溶胶TheGlyoxylateCycleHelpsplantsgrowinthedark–seedsarerichinlipids,whichcontainfattyacids–duringgermination,plantsusetheacetyl-CoAproducedinfattyacidoxidationtoproduceoxaloacetateandotherintermediatesforcarbohydratesynthesis–onceplantsbeginphotosynthesisandcanfixCO2,glyoxysomesdisappearTheCitricAcidCycle•Step4:-酮戊二酸氧化脱羧生成琥珀酰-CoA，由-酮戊二酸脱氢酶复合物催化–需要：Mg2+,coenzymeA,thiaminepyrophosphate,lipoicacid,FAD,andNAD+CH2C-COO-CH2-COO--KetoglutarateOCoA-SHNADHNAD+-ketoglutaratedehydrogenasecomplexCH2CCH2-COO-SCoAOSuccinyl-CoA+CO2TheCitricAcidCycle•Step5:琥珀酸生成，由琥珀酰-CoA合成酶催化CH2CCH2-COO-SCoAOSuccinyl-CoAGDP+PiCoA-SHSuccinate+GTP+succinyl-CoAsynthetaseCH2-COO-CH2-COO-+Succinyl-CoA+H2OSuccinate+CoA-SHGDP+PiGTP+H2OG0'(kJ昺ol-1)-33.4+30.1-3.3Succinyl-CoA+GDP+PiSuccinate+CoA-SH+GTP底物水平磷酸化TheCitricAcidCycle•Step6:琥珀酸脱氢生成延胡索酸，由琥珀酸脱氢酶催化FADFADH2CH2-COO-CH2-COO-SuccinatesuccinatedehydrogenaseCCHHCOO--OOCFumarate1、Krebs循环中唯一嵌入线粒体内膜的酶2、是电子传递链（呼吸链）组分II的主要组成3、FAD和酶共价结合4、受丙二酸的强烈抑制TheCitricAcidCycle•Step7:延胡索酸水化为苹果酸，由延胡索酸酶催化CCHHCOO--OOCFumarateH2OCH-COO-HOCH2-COO-L-MalatefumaraseTheCitricAcidCycle•Step8:苹果酸脱氢生成草酰乙酸，由苹果酸脱氢酶催化C-COO-CH2-COO-OxaloacetateNAD+NADHmalatedehydrogenaseCH-COO-HOCH2-COO-L-MalateOFromPyruvatetoCO2CoA-SH+PyruvatedehydrogenasecomplexCitricacidcyclePyruvate+NAD+Acetyl-CoA+NADH+CO2H++Acetyl-CoA+3NAD++FADGDP++Pi2CO2+CoA-SH+3NADH3H+++FADH2+GTPPyruvate4NAD++FADGDP++Pi2H2O+2H2O+3CO2+4NADH+FADH2+GTP4H+++CO2是来自丙酮酸吗？经过几轮柠檬酸循环后，脱下CO2的是来自Acetyl-CoA?PyruvateAcetyl-CoA++CoA-SH+NAD+Acetyl-CoA+NADH+CO2+H+CitrateIsocitrateIsocitrate+OxaloacetateH2OCitrate+CoA-SH+H++NAD+-Ketoglutarate+NADH+CO21.2.G?(kJ昺ol-1)-33.4-32.2+6.3-7.13.4NAD+GTPSuccinateFADFADH24.CoA-SH5.6.7.-Ketoglutarate+NAD++CoA-SHSuccinyl-CoA+NADH+CO2+H+Succinyl-CoA+GDP+Pi++Succinate+Fumarate+Fumarate+H2OMalate8.Malate+Oxaloacetate+NADH+NAD++FAD+GDP+Pi3CO24NADH++FADH2+GTP+4H+Pyruvate-33.4-3.3~0-3.8+29.2-77.7PyruvateAcetyl-CoA++CoA-SH+NAD+Acetyl-CoA+NADH+CO2+H+CitrateIsocitrateIsocitrate+OxaloacetateH2OCitrate+CoA-SH+H++NAD+-Ketoglutarate+NADH+CO21.2.G?(kJ昺ol-1)-33.4-32.2+6.3-7.13.4NAD+GTPSuccinateFADFADH24.CoA-SH5.6.7.-Ketoglutarate+NAD++CoA-SHSuccinyl-CoA+NADH+CO2+H+Succinyl-CoA+GDP+Pi++Succinate+Fumarate+Fumarate+H2OMalate8.Malate+Oxaloacetate+NADH+NAD++FAD+GDP+Pi3CO24NADH++FADH2+GTP+4H+Pyruvate-33.4-3.3~0-3.8+29.2-77.7在有氧代谢中，柠檬酸循环与电子传递链（呼吸链）的电子传递和氧化磷酸化构成氧化分解代谢的完整途径。TheCitricAcidCycle三、葡萄糖彻底氧化分解的能量计算GlucosetoCO2+H2OCytoplasmicReactionsGlucose-Glucose6-PFru-6-P-Fru-1,6-BisP1,3-BisPG-3-PG(x2)PEP-Pyruvate(x2)Oxidofglyceraldehyde-3-P(x2)MitochondrialReactionsPyruvate-acetyl-CoA(x2)NADHFADH2ATP-1(-1)-1(-1)+2(+2)+2(+2)+2+2cont'dCitricAcidCycleSuccinyl-CoA-GTP(x2)Oxidofsuccinate(x2)Oxidationofisocitrate(x2)Oxidof-KG(x2)Oxidofmalate(x2)OxPhos/ElectTransportReoxidofNADH(glycolysis)ReoxidofNADH(pyruvate)ReoxidofFADH2(CAcycle)ReoxidofNADH(CAcycle)+2(+2)+2+2+2+2+3(+5)-2-2-2-6+5(+5)+3(+3)+15(+15)TOTAL00+30(+32)NADHFADH2ATP四、柠檬酸循环的调控•柠檬酸合酶：被ATP、NADH、succinylCoA、citrate抑制•异柠檬酸脱氢酶：被ADP和NAD+激活