121712蛋白質(zhì)降解及氨基酸分解代謝simplified

安徽省岳西縣姚河鄉(xiāng)梯嶺村朱頌成副教授碩士研究生導(dǎo)師,醫(yī)學(xué)樓1205安徽省淮南市淮南八公山豆腐制作中科院上海植物生理研究所(1996-2002)DepartmentofBiochemistry,UniversityofColorado(2002-2007)DerivationofiPScellsfromvariouscells------manyspecies------manytissuesHistoneacetyltransferaseandcancercontrolProteindegradation,AminoAcidMetablismBiosynthesisofAminoAcidsNitogenFixationBiosynthesisandDegradationofNucleotidesMetabolismRegulationLehningerPrinciplesofBiochemistry,4theditionTextbookofbiochemistrywithclinicalcorrelationsThesignificanceofaminoacid1.cells/tissuesgrowing,renewingandmendingProteinDegradation/AminoAcidMetablismThesignificanceofaminoacid2.Takepartinimportantphysiologicalactivities3.Oxidationandsupplyenergy4.NitrogenbalanceBeforewegetonaminoacidmetablism:1.Breakdownofdietaryproteins2.ProteindegradationincellsProteinbreakdownstartsatstomachOptimalpH1.5～2.5，LowspecificityProducesmallpeptideandfreeaminoacids

pepsinogen胃蛋白酶原+SmallPeptideHCl、PepsinPepsinBreakdownofdietaryproteins

Cholecystokinin(縮膽囊素)stimulatessecretionofseveralpancreaticenzymes(胰腺酶）withactivityoptimaatpH7to8.Pancreasexocrinecells:Trypsinogen(胰蛋白酶原）,Chymotrypsinogen（糜蛋白酶原）,Procarboxypeptidases（羧肽酶）A,BThedigestionofproteinsinthesmallintestine（小腸）.trypsinchymotrypsinogenElastase彈性蛋白酶procarboxypeptidasetrypsinogenEnterokinase腸激酶Chymotrypsin糜蛋白酶ProelastaseCarboxypeptidase羧肽酶Enterokinase腸激酶:producedbycellsoftheduodenum

（十二指腸）腸激酶，EK，酶切位點(diǎn)DDDDKSynthesisoftheenzymesasinactiveprecursors:1Protectstheexocrinecellsfromdestructiveproteolyticattack.2Transportation3RegulationElaboratemechanismforgettingactivedigestiveenzymes.ProteinDigestion

ProteolyticenzymesofpancreaticjuiceEndopeptidases內(nèi)肽酶Exopeptidases外切酶Trypsin

:Arg,Lys(C)Chymotrypsin:Tyr,Trp,Phe,Met,Leu(C)Elastase:Ala,Gly,Ser(C)carboxypeptidaseaminopeptidaseCleavagesitesandsubstratesofproteasesCleavagesitesandsubstratesofproteases一Theyhydrolyzethepeptidesveryefficiently.Inhumans,mostglobularproteinsfromanimalsourcesarealmostcompletelyhydrolyzedtoaminoacidsinthegastrointestinaltract,butsomefibrousproteins,suchaskeratin(角蛋白）,areonlypartlydigested.Inaddition,theproteincontentofsomeplantfoodsisprotectedagainstbreakdownbyindigestiblecellulosehusks（外殼）.AbsorptionoffreeaminoacidsTrypsinandProteaseinCellCultureTrypsin和蛋白/肽的質(zhì)譜測(cè)序基于MS/MS的蛋白質(zhì)序列

和修飾鑒定Liquidchromatography-coupledmassspectrometery(LC-MS)TryspindigestionAdiseasecausedbyobstructionofthenormalpathwaybywhichpancreaticsecretionsentertheintestine.Theenzymogensoftheproteolyticenzymesareconvertedtotheircatalyticallyactiveformsprematurely,insidethepancreaticcells,andattackthepancreatictissueitself.Thiscausesexcruciatingpainanddamagetotheorganthatcanprovefatal.Acutepancreatitis(急性胰腺炎）andProtease問(wèn)題：

1蛋白酶水解的是什么化學(xué)鍵？

2人體消化道內(nèi)有哪些蛋白酶?

3酶以酶原的形式存在有什么意義？

思考題：

有哪些方式將蛋白質(zhì)轉(zhuǎn)變?yōu)榘被幔?/p>

TwomajorsystemsfortheturnoverofcellularproteinsLysosomesandUbiquitin-mediatedproteindegradationProteinDegradationinCellLysosomalDegradationPathwayProteinsA.EndocytosedB.CytosolicC.OrganellesbyautophagyLysosomalhydrolasesexhibitacidicpHoptimal溶酶體貯積病UbiquitinpathwayUbiquitinProteasomeE3ligase(Cullin-RINGcomplex)ProteinDegradationSysteminCellWhatisUbiquitin?HumanUb:MQIFKVTLTGKTITLEVEPNDTIENVKAKIQDKEGIPPDQQRLIFAGKQLEDGRTLADYNIQKESTLHLVLRLRGGYeastUb:MQIFKVTLTGKTITLEVESSDTIDNVKSKIQDKEGIPPDQQRLIFAGKQLEDGRTLSDYNIQKESTLHLVLRLRGGUbiquitylation-SubUbiquitinpathwayE1-S-UbE2-S-UbE3substratesUbMono-ubiquitinationChangeproteinconformation,Localization,interactionPolyubiquitinationUbUbUbUbsubstratesProteosomeDegradationE1-Ubactivatingenzyme(Uba)UsesATPAMP+PPitoformthioesterbondbetweenitssulfhydrylgroupandtheterminalcarboxylategroupofUBE2-Ubconjugatingenzyme(Ubc):AcceptsactivatedUBE3-UbLigase:CatalyzestransferofUBfromE2to-aminogroupoflysineontargetproteinforminganisopeptidebond蛋白酶體（Proteasome）19Slid19Slid20ScoreControlofp53byubiquitylationChromatinstructureandtranscriptionalregulation.TurnoverofoncogeneproductsandtumorbiologyCellcyclecontrolRegulationofDNArepairresponseApoptosisEndocytosis一般了解（不考）ControlofCellCycleProgressionbyDistinctE3ComplexesAnaphase-PromotingComplexTwofamiliesofE3ubiquitinligases一般了解（不考）一般了解（不考）TheNobelPrizeinChemistry,2004“forthediscoveryofubiquitin-mediatedproteindegradation”AvramHershkoIrwinRoseAaronCiechanoverQuestions細(xì)胞內(nèi)主要的蛋白質(zhì)降解體系是什么？各有什么特點(diǎn)人體消化道內(nèi)有哪些蛋白酶參與了蛋白質(zhì)的消化？AminoAcidPoolAminoacidmetabolismDeamination（脫氨）Transamination（轉(zhuǎn)氨作用）Ureacycle（尿素循環(huán)）Pyruvategroup

aKetoglutarategroupSuccinyl-CoAgroupOxaloacetategroupFumarate/oxaloacetategroupAlanine/acetoacetategroupAcetyl-CoA/acetoacetategroupDecarboxylation1.DeaminationofAAstransamination

oxidativedeaminationuniondeamination

Importantinthediagnosisofheartandliverdamagecausedbyheartattack,drugtoxicity,orinfection.Afteraheartattack,avarietyofenzymes,includingtheseaminotransferases,leakfromtheinjuredheartcellsintothebloodstream.AssaysforTissueDamagebyAnalysesofcertainenzymeactivitiesinbloodserum轉(zhuǎn)氨酶與器官損傷Transamination(轉(zhuǎn)氨作用）1Anaminogroupistransferredtoanα-ketoacid,withformationofthecorrespondingaminoacidandanotherα-ketoacid.2Usuallyoccursinaminoaciddegradation3Transaminationisreversible,however，transaminationofessentialaminoacidisunidirectional,Ser,LysandProcannotbetransaminated.5KeyrolesofGluandα-ketoglutarateintransaminationTransamination4α-ketoacidproducedintransaminationcanbeintermediatorforTCA6Onlytheα-aminogroupofGluentersUreacycleALT:Alanineaminotransferase(inliver)

丙氨酸氨基轉(zhuǎn)移酶，alsocalledglutamate-pyruvatetransaminase（谷丙轉(zhuǎn)氨酶）,GPTAST:Aspartateaminotransferase(inheart)，alsocalledglutamate-oxaloacetatetransaminase（谷草轉(zhuǎn)氨酶）,GOTTwoimportanttransaminases:嗜酒如命進(jìn)食高蛋白補(bǔ)品服用某些藥物肝病（肝炎、肝損傷…)腎病(谷氨酰轉(zhuǎn)肽酶)

FunctionExpressionActivityCellularLocalizationDomainModificationStructureRegulationCofactorTransamination---needscoenzymepyridoxalphosphate（磷酸吡哆醛，B6）1Covalent2Interaction3Schiff’sbaseVitamineB6亞胺Transamination---needcoenzymepyridoxalphosphate(PLP)ALLAminotransferasesidentifiedsofarusepyrodoxalphosphateascoenzyme.Pyridoxaminephosphate(PMP)(2)OxidativedeaminationGlutamateReleasesItsAminoGroupasAmmoniaintheLiver1Inhepatocyte’smitochondria,2Glutamateundergoesoxidativedeamination3CatalyzedbyL-glutamatedehydrogenase4Producesfreeammonia5ProducesNADH6RequiredforUreaCycle7TheenzymeisstrictlyregulatedHowandWhy？(3)UniondeaminationAlanine+α-ketoglutaratePyruvate+glutamateGlutamate+NAD++H2Oα-ketoglutarate+NADH+NH4+NetReaction:

Alanine+NAD++H2Opyruvate+NADH+NH4+ThecombinedactionofanaminotransferaseandglutamatedehydrogenaseTransaminase,glutamatedehydrogenase.草酰乙酸腺苷酰琥珀酸蘋(píng)果酸延胡索酸腺苷酸次黃苷酸(3)

Purinenucleotidecycle(inmuscle)1.Sources:

①

DeaminationofAAs--mainsource②CatabolismofothernitrogencontainingcompoundsSourceandoutletofammonia（NH3）

2.Outlets:(1)Formationofureainliver(2)FormationofGln(3)SynthesisofAA(4)ExcreteinurineMetabolismofAmmoniamostvertebratesbirds&reptilesfish&otheraquaticvertebratesTransportationofNH31Glucose–Alaninecycle2TransportationofammoniabyGln

UreacycleinlivercellsAmmoniaIsToxictoAnimals1Glucose-AlanineCycleAlanineTransportsAmmoniafromSkeletalMusclestotheLiver2.TransportationofammoniabyGlnExcessammoniaintissuesisaddedtoglutamatetoformglutamine,catalyzedbyglutaminesynthetase(谷氨酰胺合成酶）

Aftertransportinthebloodstream,theglutamineenterstheliverandNH4isliberatedinmitochondriabytheenzymeglutaminase（谷氨氨酰胺酶）bloodcirculationliverotherorgansmusclekidneyureaglucose–alaninecycleGlutaminaseNeutralNon-polarityNon-toxicEasilytransportedthroughmembraneFormationofurea1.Site:liver(mitochondriaandcytosol)2.Process---------ornithinecycleUreaIsProducedfromAmmoniainFiveEnzymaticSteps(氨、瓜、精琥、精、尿）HansCrebPhospho-carbonateHydrogencarbonateCarbamateCarbamoyl-phosphateFormationofCarbamoylphosphatebyCarbamoylphosphateSynthetaseI①Formationofcarbamoylphosphate（氨甲酰磷酸）inmitochondria+2ADP+Pi2ATP+CO2+NH3+H2OH2N-C-PO1The2Nofurea:

NH3andAsp.2Starts/endsupwithornithine3NH3,CO2formcaramoylphosphate,4Citrullineformation,5IncorperationofAsp.6Energeconsuming:Oneurea3ATP(4~P?)8Needs5enzymes：carbamoylphosphatesynthetaseⅠornithinecarbamoyltransferase,OCTArgininosuccinateSynthetaseArgininosuccinaseArginase10Ratelimiting:1ststepUreaformation:RegulationofureasynthesisActivationoftheureacyclebyN-acetyl-glutamateN-acetyl-glutamate

synthetaseIRegulationofureasynthesisEnergyconsuming:oneureawillcosts4~PCarbamoylphasphatesynthetaseISubstrateactivation/inductionResponsivetoconcentrationofammonia/aminoacidinliverProtein-richdietaryStarvation

Aspartate（天冬氨酸）

Fumarate(延胡素酸）GeneticDefectsintheUreaCycle----Life-ThreateningTotallackofanyUreaCycleenzymeislethal.Hyperammonemia-elevatedNH3inblood.ElevatedNH3istoxic,especiallytothebrain.Severementalretardationresulted.Controltheprotein/aminoacidintakeRemovetheexcessiveNH3/aminoacidsAddintermediatesofUreacycle一般了解（不考）Tetrahydrofolicacid(FH4)OnecarbonunitsarecarriedbyFH4.Substanceforsynthesisofnucleicacid.N5,N10ofFH4participateinthetransferofonecarbonunits.葉酸缺乏：new-born，BloodcelldiseaseAnalogasdrug

N5,N10ofFH4participateinthetransferofonecarbonunits.Formationofonecarbonunit(1)Ser→N5,N10-CH2-FH4

(2)Gly→N5,N10-CH2-FH4一般了解（不考）

(3)His→N5-CH=NHFH4一般了解（不考）(4)Trp→N10-CHOFH4一般了解（不考）Tyrosineformationfromphenylalanine(Monooxygenasereaction,單氧化酶)PhenylKetonuria苯丙酮尿癥PhenylalanineHydroxylaseTyrosineAminotransferaseParahydroxyphenylepyruvateDi-oxygenase(ascorbateVc)HomogentisateDioxygenaseMaleylAcetoacetateIsomaeraseFumarylAcetoacetasemetaboliteconcentrationinurine[mmoles/24h]controlpatientphenylalanine0.181.8–6.0phenylpyruvate--1.8–12phenyllactate--1.8–3.3phenylacetate--sign.incr.phenylacetylglutama