氨基酸代謝第一節Thenitrogencycle

Nitrogenexistspredominantlyinanoxidizedstateintheenvironment,occurringprincipallyasN2intheatmosphereorasnitrateion(NO3-)inthesoilsandoceans.Itsacquisitionbybiologicalsystemsisaccompaniedbyitsreductiontoammoniumion(NH4+)andtheincorporationofNH4+intoorganiclinkageasaminogroup.ThereductionofNO3-toNH4+occursingreenplants,variousfungi,andcertainbacteriainatwo-stepmetabolicpathwayknownasnitrateassimilation.第一節Thenitrogencycle

TheformationofNH4+fromN2gasistermednitrogenfixation.N2fixationisanexclusivelyprokaryoticprocess.Noanimalsarecapableofeithernitrogenfixationornitrateassimilation.Animalsreleaseexcessnitrogeninareducedform,eitherasNH4+orasorganicnitrogenouscompoundssuchasurea.ThereleaseofNoccursbothduringlifeandasaconsequenceofmicrobialdecompositionfollowingdeath.第一節Thenitrogencycle

Variousbacteriareturnthereducedformsofnitrogenbacktotheenvironmentbyoxidizingthem.TheoxidationofNH4+toNO3-isperformedbynitrifyingbacteria.NitratenitrogenalsoreturnstotheatmosphereasN2asresultofthemetabolicactivityofdenitrifyingbacteria.第一節ThenitrogencycleDietaryproteinsaredigestedintoaminoacidsinthegastrointestinal(胃腸)tractviatheactionofpepsin,trypsin,chymotrypsin,carboxypeptidasesandaminopeptidases.SourcesofaminoacidsforanimalsProteins(butnotpepsin)unfoldedAbsorbedastri-&dipeptides,andaminoacidsDegradation&absorptionofdietaryproteinsPepsin:thefirstenzymediscovered(18thcentury).proteasesEssentialaminoacidsAminoacidscannotbestoredinanimals:excessbeingcompletelyoxidizedtoreleaseenergyorconvertedtostorablefuels(fattyacidsorcarbohydrates).

Overallfateofexcessaminoacids第二節Aminoaciddegradation

1.氧化脫氨基

氨基酸在酶的作用下脫去氨基生成相應酮酸的過程，叫氧化脫氨基作用。一.氨的去路

Glu+NAD(P)+H2O

a-KG+NH4+

+NADH(P)

+H+

2.脫氫酶作用-GDH一.氨的去路

3.轉氨基作用一.氨的去路

轉氨基作用是α-氨基酸和α-酮酸之間氨基的轉移作用。一種α-氨基酸的α-氨基借助轉氨酶（transaminase）的催化作用轉移到α-酮酸的羰基上，結果生成新的酮酸，而原來的α-酮酸則形成相應的氨基酸。

3.轉氨基作用谷丙轉氨酶催化的轉氨基作用機理

一.氨的去路

4.聯合脫氨作用(轉氨酶-谷氨酸脫氫酶)谷氨酸

α-酮戊二酸

丙氨酸

丙酮酸

轉氨酶

谷氨酸脫氫酶

NAD(P)+H+

NAD(P)+聯合脫氨基作用

PLP

一.氨的去路

在氨基酸脫羧酶催化下進行脫羧作用，生成一個伯胺類化合物和CO2，其反應可以用下式表示二.

脫羧基作用

PLPactsasatemporarycarrierofaminogroupsattheactivesitesofallaminotransferases.PLPfacilitatesseveraldifferenttypesoftransformationaroundthea-carbonofaminoacids.PLPisderivedfromvitaminB6(pyridoxine，吡哆醇)吡哆醛磷酸磷酸吡哆胺SerumaminotransferaseshavebeenusedasclinicalmarkersoftissuedamagesDamagedheartorlivercellsleakaminotransferases.Bloodaspartateaminotransferaseandalanineaminotransferaseareusuallyexaminedforindicationsofillness.三.氨基酸碳架的分解

氨基酸脫羧酶

1.進入TCA循環

Oxidationofthecarbonskeletonsofaminoacidsinmammals2.再合成為氨基酸谷氨酸＋丙酮酸α-酮戊二酸＋丙氨酸谷氨酸＋草酰乙酸α-酮戊二酸＋天冬氨酸三.氨基酸碳架的分解

3.轉變為糖和脂肪

當體內不需要將α-酮酸再合成氨基酸，并且體內的能量供給充足時，α-酮酸可以轉變為糖或脂肪。例如，用氨基酸飼養患人工糖尿病的狗，大多數氨基酸可使尿中的葡萄糖的含量增加，少數幾種可使葡萄糖及酮體的含量同時增加。在體內可以轉變為糖的氨基酸稱為生糖氨基酸，按糖代謝途徑進行代謝；能轉變為酮體的氨基酸稱為生酮氨基酸。

三.氨基酸碳架的分解

硝酸鹽還原分兩步進行：第一步在硝酸還原酶（nitratereductase,NR）催化下，由NAD（P）H提供1對電子，硝酸鹽被還原為亞硝酸鹽，第二步是在亞硝酸還原酶（nitritereductase,NiR）下，由還原型鐵氧還蛋白（Fdred）提供3對電子，使亞硝酸鹽（NO2-）還原成氨。

第三節Nitratereduction硝酸鹽還原分兩步進行：第一步在硝酸還原酶（nitratereductase,NR）催化下，由NAD（P）H提供1對電子，硝酸鹽被還原為亞硝酸鹽，第二步是在亞硝酸還原酶（nitritereductase,NiR）下，由還原型鐵氧還蛋白（Fdred）提供3對電子，使亞硝酸鹽（NO2-）還原成氨。

第三節NitratereductionAmmoniumentersorganiclinkageviathreemajorreactionsthatarefoundinallcells.Theenzymesmediatingthesereactionsare：(1)Cabamoyl-phosphatesynthetaseI(氨甲酰磷酸合成酶)(2)Glutamatedehydrogenase（谷氨酸脫氫酶）,(3)Glutaminesynthetase（谷氨酰氨合成酶）.

第四節AmmoniumassimilationNH4+inhepatocytes(肝細胞)isconvertedintoureaforexcretionviatheureacycleinmostterrestrialvertebratesUreaisformedfromammonia,CO2(asbicarbonate)andAsp.ThepathwaywasalsodiscoveredbyHansKrebsin1932(fiveyearsbeforehediscoveredthecitricacidcycle).FourATPmoleculesareconsumedtoproduceeachurea.

Carbamoyl-phosphatesynthetaseIcatalyzesoneofthestepsintheureacycle.TwoATPareconsumed,oneintheactivationofHCO3-forreactionwithammonium,andtheotherinthephosphorylationofthecarbamateformed:1.Carbamoyl-phosphatesynthetaseI

NH4++HCO3-+2ATPH2N-CO-O-PO3-+2ADP+Pi+2H+

N-acetylglutamateisanessentialallostericactivatorforthisenzyme第四節AmmoniumassimilationThesynthesisofCarbamoyl（氨甲酰）

phosphaterequirestwoactivationsteps,consumingtwoATPmolecules:oneforactivatingHCO3-,theothertophosphorylatecarbamate.

ananhydride1.Carbamoyl-phosphatesynthetaseI

該反應消耗2個ATP分子中的兩個高能磷酸鍵，其中1個是用于活化HCO3-，另1分子ATP則用于磷酸化氨甲酰基。第四節AmmoniumassimilationFumarateisconvertedbacktoAspviaapartialusageofthecitricacidcycle.TherateofureasynthesisiscontrolledattwolevelsAllosteric（別構）regulation:N-acetylglutamate,bybindingtoasitewhichhydrolyzes（水解）Glninanotherisozyme,positivelyregulatescarbamoylphosphatesynthetaseIactivity.Generegulation:synthesesoftheureacycleenzymesareallincreasedduringstarvation(whenenergyhastobeobtainedfrommuscleproteins!)orafterhighproteinuptake.Theratesoftranscriptionofthefivegenesencodingtheenzymesareincreased.GeneticdefectsoftheureacycleenzymesleadtohyperammonemiaandbraindamageHighlevelsofammonialeadtomentaldisorderorevencomaanddeath.Ingeniousstrategiesforcopingwiththedeficiencieshavebeendevisedbasedonathoroughunderstandingoftheunderlyingbiochemistry.StrategyI:dietcontrol,providetheessentialaminoacidsintheira-ketoacidforms.StrategyII:whenargininosuccinatelyaseisdeficient,ingestingasurplusofArgwillhelp(ammoniawillbecarriedoutofthebodyintheformofargininosuccinate,insteadofurea).StrategyIII:whencarbamoylphosphatesynthetaseI,ornithinetranscarbamoylase,orargininosuccinatesythetasearedeficient,theammoniacanbeeliminatedbyingestingcompounds(e.g.,benzoateorphenylacetate),whichwillbeexcretedafteracceptingammonia.

Glutamatedehydrogenasecatalyzesthereductiveaminationofa-ketoglutaratetoyieldglutamate.Reducedpyridinemucleotides(NADHorNADPH)providethereducingpower:2.Glutamatedehydrogenase(GDH)

NH4++a-ketoglutarate+NADPH+H+

glutamate+NADP++H2O第四節AmmoniumassimilationTheglutamatedehydrogenasereaction第四節Ammoniumassimilation3.Glutaminesynthetase(GS)GlutaminesynthetasecatalysestheATP-dependentamindationofthe-carboxylgroupofglutamatetoformglutamine.GSactivitydependsonthepresenceofdivalentcationssuchasMg2+.GlutamineisamajordonorinthebiosynthesisofmanyorganicNcompoundsandGSactivityistightlyregulated.

GDHandGSareresponsibleformostoftheammoniumassimilatedintoorganiccompounds.第四節Ammoniumassimilation谷氨酰胺合成酶谷氨酰胺合成酶第四節AmmoniumassimilationTheGlutaminesynthetaseisaprimaryregulatorypointinnitrogenmetabolism:beingregulatedbyatleasteightallostericeffectorsandreversibleadenylylation.

Thebacterial

glutaminesynthetasehas12subunitsarrangedastworingsofhexamers.Activesites

Tyr397(adenylylationsite)Theglutaminesynthetaseisaccumulativelyinhibitedbyatleast8allostericeffectors,mostlyendproductsofglutaminemetabolism.Glutamatesynthasecatalyesthereductiveaminationofa-ketoglutaratesuingtheamide-NofglutamineastheNdonor:Glutamatesynthase(GOGAT)

Reductant+a-KG+Gln2Glu+oxidizedredctant

第四節AmmoniumassimilationTheglutamatesynthasereaction谷氨酸合酶第四節AmmoniumassimilationOnlycertainbacteriacanfixN2intoammoniaRhizobiaCyanobacteria藍細菌根瘤菌第5節NitrogenfixationThedinitrogenase(固氮酶)complexincertainbacteria(diazotrophs)catalyzestheconversionofN2(azote,“withoutlife”)toNH3,whichistheultimatesourceofnitrogenforallnitrogen-containingbiomolecules.

N2+8H++8e-2NH3+H2

TheHabermethod:N2+3H22NH3

G`o=-33.5kJ/molwithironcatalyst,500oC,300atmospheres.Thenitrogenasecomplexconsistsofdinitrogenaseanddinitrogenaseredutase

bothbeingiron-sulfurproteins.Dinitrogenase(a2b2)orFeMoproteinReductase:adimeroftwoIdenticalsubunitsbridgedbya4Fe-4S.ATPhydrolysisiscoupledtoproteinconformatinalchanges.Dinitrogenasereductase(dimer)orFeproteinADPADP4Fe-4S8Fe-7S(P-cluster)Fe-Mocofactor

e-Fe-Mocofactor8Fe-7S(P-cluster)4Fe-4SADPADPMolybdenum(orvanadium)N2isbelievedtobereducedbytheFe-MocofactorN2FeFeFeFeFeFeFeSSSSSSSSSMo高檸檬酸ElectronsaretransferredthroughaseriesofcarrierstoN2foritsreductiononthenitrogenasecomplex.ElectronsaretransferredtoN2boundintheactivesiteofdinitrogenaseviaferredoxin/flavodoxinanddinitrogenaseReductase.N2+8H++8e-+16ATP+16H2O

2NH3+H2+16ADP+16Pi(orphotophosphorylation)Conformationalchangereducese-affinityTheoxidizeddinitrogenasereductasedissociatesfromthedinitrogenaseReduceddinitrogenasereductaseassociateswiththedinitrogenaseThenitrogenasecomplexisextremelylabiletoO2andvariousprotectivemechanismshaveevolved:livinganaerobically,formingthickwalls,uncouplinge-transportfromATPsynthesis(enteringO2isusedinmediately)orbeingprotectedbyO2-bindingproteins.Genesencodingtheproteincomponentsofthenitrogenasecomplexarebeingtransferredintonon-nitrogen-fixingbacteriaandplants.ReducednitrogenintheformofNH4+isassimilatedintoaminoacidsmainlyviaatwo-enzymepathway:glutaminesynthetaseandglutamatesynthase

(anenzymeonlypresentinbacteriaandplants).GlnsynthetaseGluSynthase(+NADPH+ATP)GlnsynthetaseThepathwaysforammoniatoenterorganiccompounds.GluDehydrogenaseVeryminor)

Asnsynthetase

CarbamoylPhosphateSynthetase

Transamination(orNH4+)