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Sinteza corpilor cetoniciSinteza corpilor cetonici(cetogeneza)(cetogeneza)
Principala cale de metabolizare a acetilPrincipala cale de metabolizare a acetil CoA CoA – includerea în ciclul Krebs (în condiţiile în – includerea în ciclul Krebs (în condiţiile în care scindarea lipidelor şi a glucidelor este care scindarea lipidelor şi a glucidelor este echilibrată)echilibrată) - “lipidele ard în flacăra - “lipidele ard în flacăra glucidelor”glucidelor”
În lipsa glucidelor; inaniţie, diabet - OA se În lipsa glucidelor; inaniţie, diabet - OA se utilizează pentru generarea Gl.utilizează pentru generarea Gl.
În lipsa OA, Acetil Co A recurge la formarea În lipsa OA, Acetil Co A recurge la formarea corpilor cetonici: corpilor cetonici: acetoacetatul, acetoacetatul, ββ--hidrohibutiratul şi acetonahidrohibutiratul şi acetona
Sinteza lor are loc în ficat, dar se utilizează Sinteza lor are loc în ficat, dar se utilizează de ţesuturile perifericede ţesuturile periferice
Au rol energetic (muşchiul cardiac, stratul Au rol energetic (muşchiul cardiac, stratul cortical al rinichilor)cortical al rinichilor)
cetogenezacetogeneza
Utilizarea corpilor Utilizarea corpilor cetonicicetonici
Acetoacetatul – 2 mol de Acetoacetatul – 2 mol de acetil CoA, utilizate ulterior acetil CoA, utilizate ulterior în ciclul Krebs (23 ATP)în ciclul Krebs (23 ATP)
A doua cale de activare a A doua cale de activare a acetoacetatului poate fi:acetoacetatului poate fi:
Acetona: Acetona: 1.1. pînă la propandiol (CH3-CHOH-pînă la propandiol (CH3-CHOH-
CH2OH) , scindat la fragmente CH2OH) , scindat la fragmente acetil şi formilacetil şi formil
2.2. Transformată în piruvat (prin Transformată în piruvat (prin hidroxilare dublă)hidroxilare dublă)
Cetonemie, cetonurieCetonemie, cetonurie Cetonemie- mărirea c% de corpi cetonici Cetonemie- mărirea c% de corpi cetonici
în sîngeîn sînge Cetonurie – apariţia CC în urinăCetonurie – apariţia CC în urină Diete bogate în lipide, sărace în glucide; Diete bogate în lipide, sărace în glucide;
inaniţie, diabet, dereglări gastrointestinale inaniţie, diabet, dereglări gastrointestinale la copii sau gravide; glucozurie renalăla copii sau gravide; glucozurie renală
Eliminarea hidroxibutiratului şi Eliminarea hidroxibutiratului şi acetoacetatului din organism (fiind anioni acetoacetatului din organism (fiind anioni la excreţie) conduce la pierderea de la excreţie) conduce la pierderea de cationi – Na- rezultă cetoacidozacationi – Na- rezultă cetoacidoza
Pierderea H2O – dehidratarea Pierderea H2O – dehidratarea organismuluiorganismului
Biosinteza Biosinteza lipidelorlipidelor
Obiectivele:Obiectivele: Biosintaza acizilor graşi:Biosintaza acizilor graşi:1.1. saturaţi cu număr par de atomi de carbon;saturaţi cu număr par de atomi de carbon;2.2. nesaturaţi cu număr par de atomi de carbon;nesaturaţi cu număr par de atomi de carbon;3.3. saturaţi cu număr impar de atomi de carbon.saturaţi cu număr impar de atomi de carbon. Enzimele, coenzimele, reglarea.Enzimele, coenzimele, reglarea. Biosinteza TAG: substanţele iniţiale, enzimele şi Biosinteza TAG: substanţele iniţiale, enzimele şi
coenzimele, reglarea.coenzimele, reglarea. Biosinteza fosfogliceridelor: substratele, reacţiile Biosinteza fosfogliceridelor: substratele, reacţiile
parţiale ale I şi a II căi; parţiale ale I şi a II căi; Biosinteza sfingolipidelor: precursorii, reacţiile Biosinteza sfingolipidelor: precursorii, reacţiile
principale, enzimele, reglarea.principale, enzimele, reglarea. Metabolismul colesterolului. Biosinteza Metabolismul colesterolului. Biosinteza
colesterolului – substratele, etapele, reacţiile colesterolului – substratele, etapele, reacţiile parţiale ale I etape (până la acidul mevalonic), parţiale ale I etape (până la acidul mevalonic), enzimele, coenzimele, reglarea. Căile de utilizare enzimele, coenzimele, reglarea. Căile de utilizare şi eliminare ale colesterolului.şi eliminare ale colesterolului.
Sinteza AGSinteza AG Sinteza AG şi încorporarea lor în Tg Sinteza AG şi încorporarea lor în Tg
constituie mecanismul principal de stocare constituie mecanismul principal de stocare a excesului de glucide alimentare (Gl nu se a excesului de glucide alimentare (Gl nu se mai transformă în glicogen dar în Tg)mai transformă în glicogen dar în Tg)
EtapeleEtapele:: Sinteza de novo cu formarea acidului Sinteza de novo cu formarea acidului
palmiticpalmitic Elongarea acidului palmiticElongarea acidului palmitic Introducerea de legături duble în AGIntroducerea de legături duble în AG
Particularităţile sintezei Particularităţile sintezei AGAG Are loc în citozolAre loc în citozol
E – acid gras sintetaza – alcătuită din 8 E – acid gras sintetaza – alcătuită din 8 proteine (domenii)- 7 sunt enzime, a 8-a – proteine (domenii)- 7 sunt enzime, a 8-a – proteina proteina ((purtătoarepurtătoare)) transportatoaretransportatoare de de acilacil -ACP -ACP..
ACP cuprinde 2 grupe SHACP cuprinde 2 grupe SH::1.1. – –SH furnizat de un rest de cisteinilSH furnizat de un rest de cisteinil:: SH-Cis SH-Cis2.2. - SH- SH - - fosfopanteteina, ata fosfopanteteina, ataşşatatăă prin prin
leglegăătura fosfat-Sertura fosfat-Ser: SH-Pant: SH-Pant Ca iniţiator este acetil CoA (rezultat din Ca iniţiator este acetil CoA (rezultat din
glicoliză), pe cînd sursa majoră – malonil glicoliză), pe cînd sursa majoră – malonil CoACoA
rolul reducător îi revine NADPH+Hrolul reducător îi revine NADPH+H
Sinteza de novo cu Sinteza de novo cu formarea acidului palmiticformarea acidului palmitic
Etapele:Etapele:1.1. transferul lui Acetil CoA din transferul lui Acetil CoA din
mitocondrii în citozolmitocondrii în citozol2.2. Sinteza de malonil CoASinteza de malonil CoA3.3. Sinteza acidului palmiticSinteza acidului palmitic
TTransferul lui Acetil CoA din ransferul lui Acetil CoA din mitocondrii în citozolmitocondrii în citozol
Sinteza de malonil CoASinteza de malonil CoA acetil-CoA + HCOacetil-CoA + HCO33
+ + ATPATP ADPADP + + PPii + + malonil-CoA malonil-CoA
E- acetil CoAE- acetil CoACarboxilazaCarboxilaza citrat, citrat, InsulinaInsulina palmitoil CoApalmitoil CoAGlucagonulGlucagonul
Sinteza acidului palmiticSinteza acidului palmitic
Sinteza acidului palmiticSinteza acidului palmitic
Sinteza acidului palmiticSinteza acidului palmitic
Ciclu de reacţii este reluat: butiril+ACP Ciclu de reacţii este reluat: butiril+ACP se condensează cu malonil+ACP- formînd se condensează cu malonil+ACP- formînd în final C6-acil ACP.în final C6-acil ACP.
Catena AG creşte pînă la formarea Catena AG creşte pînă la formarea palmitil-S-ACPpalmitil-S-ACP
RReacţia sumară:eacţia sumară: Acetil-ACP+7 malonil-CoA +14 Acetil-ACP+7 malonil-CoA +14 NADPH+HNADPH+H
Palmitat +7CO2+14NADP Palmitat +7CO2+14NADP ++ + 8HSCoA+6H2O + 8HSCoA+6H2O
deoarece malonil CoA se sintetizează deoarece malonil CoA se sintetizează din acetil CoA:din acetil CoA:88 acetacetiil-CoA + 14l-CoA + 14 NADPH +NADPH +H H ++ + 7+ 7
ATPATP palmitate+ 14palmitate+ 14 NADPNADP++ + + 88HSHSCoACoA ++ 77 ADPADP ++ 77 PPii
Elongarea AGElongarea AG Localizată: reticulul endoplasmaticLocalizată: reticulul endoplasmatic AG este activat AG este activat La acidul preexistent (palmitil CoA) se ataşează malonil La acidul preexistent (palmitil CoA) se ataşează malonil
CoACoA
Biosinteza AG nesaturaţiBiosinteza AG nesaturaţi Pot fi sintetizaţi AC mononesaturaţi. Introducerea unei Pot fi sintetizaţi AC mononesaturaţi. Introducerea unei
duble legături are loc prin acţiunea unei monooxigenaze duble legături are loc prin acţiunea unei monooxigenaze (introduce gruparea hidroxil), urmată de deshidratare(introduce gruparea hidroxil), urmată de deshidratare
Acidul linoleic şi linolenic sunt esenţiali (exogen)Acidul linoleic şi linolenic sunt esenţiali (exogen) Acidul linoleic se transformă în acidul arahidonic conform Acidul linoleic se transformă în acidul arahidonic conform
reacţiilorreacţiilor
Sinteza Sinteza TAGTAG 2 căi:2 căi:1.1. calea monoacilglicerolului: are loc în peretele calea monoacilglicerolului: are loc în peretele
intestinal (enterocite)din produşi absorbiţi intestinal (enterocite)din produşi absorbiţi (resinteza lipidelor).(resinteza lipidelor).
2.2. calea glicerolfosfatului: în toate ţesuturile calea glicerolfosfatului: în toate ţesuturile (activă: ţesutul adipos şi ficat)(activă: ţesutul adipos şi ficat)
AG sunt incorporaţi în TAG sub formă activă de AG sunt incorporaţi în TAG sub formă activă de acilCoA:acilCoA:
R-COOH + ATP + HS-CoA R-COOH + ATP + HS-CoA +H2O+H2O R-CO~SCoA R-CO~SCoA + AMP + + AMP + 2 P2 Pii
E- acil Co A sintetazaE- acil Co A sintetaza
1. calea 1. calea monoacilglicerolului monoacilglicerolului
TG împreună cu FL,Col, proteine TG împreună cu FL,Col, proteine sunt incorparate în CM şi secretaţi sunt incorparate în CM şi secretaţi mai departe în vasele limfatice.mai departe în vasele limfatice.
calea glicerolfosfatuluicalea glicerolfosfatului
originea glicerol originea glicerol fosfatului fosfatului
În ficat:În ficat: În ţesut adipos, ficatÎn ţesut adipos, ficat
Sinteza glicerofosfolipidelorSinteza glicerofosfolipidelor 2 căî de sinteză:2 căî de sinteză: Sinteza Sinteza de novo - de novo - utilizează ca utilizează ca
intermediar comun acidul fosfatidicintermediar comun acidul fosfatidic Calea de rezervă – o sinteză din Calea de rezervă – o sinteză din
produse formateproduse formate Particularitatea biosintezei FL este Particularitatea biosintezei FL este
participarea precursorilor în forme participarea precursorilor în forme active de derivaţi ai citidin fosfatului active de derivaţi ai citidin fosfatului (CDP) ca CDP-colina, CDP-(CDP) ca CDP-colina, CDP-etanolamina, CDP-diglicerid.etanolamina, CDP-diglicerid.
Sinteza Sinteza de novode novo
2. sinteza din produse 2. sinteza din produse formateformate
Sinteza sfingolipidelorSinteza sfingolipidelor Se formează din palmitoil CoA şi SerSe formează din palmitoil CoA şi Ser Sfingozina liberă se formează din Sfingozina liberă se formează din
ceramidă ceramidă Sinteza are loc pe suprafaţa Sinteza are loc pe suprafaţa
citozolică a membranelor reticulului citozolică a membranelor reticulului endoplasmaticendoplasmatic
Sinteza sfingolipidelorSinteza sfingolipidelor
Sinteza ColesteroluluiSinteza Colesterolului Se sintetizează din Acetil-CoASe sintetizează din Acetil-CoA Necesită 18 moli de Acetil-CoA şi 18 de ATPNecesită 18 moli de Acetil-CoA şi 18 de ATP Principalul organ de metabolizare este ficatul, Principalul organ de metabolizare este ficatul,
dar are loc şi în intestin, suprarenale, dar are loc şi în intestin, suprarenale, tegumentetegumente
Are loc în 3 etape:Are loc în 3 etape:1.1. Sinteza acidului mevalonicSinteza acidului mevalonic2.2. mevalonatul prin mai multe reacţii - 3∆-mevalonatul prin mai multe reacţii - 3∆-
izopentenil pirofosfat. 6 molecule de 3∆-izopentenil pirofosfat. 6 molecule de 3∆-izopentenil pirofosfat – scualenizopentenil pirofosfat – scualen
3.3. Scualenul se supuine ciclizării – lanosterol -- ColScualenul se supuine ciclizării – lanosterol -- Col
H2CC
CH3HO
CH2C
O O
CH2 OH
H2CC
CH2 CH2 O P O P O
O
O
O
O
CH3
H2CC
CH3HO
CH2C
O O
CH2 O P O P O
O
O
O
O
CO2
ATPADP + Pi
2 ATP2 ADP
mevalonate
5-pyrophosphomevalonate
(2 steps)
isopentenyl pyrophosphate
H2CC
CH2 CH2 O P O P O
O
O
O
O
CH3
H3CC
CH CH2 O P O P O
O
O
O
O
CH3
isopentenyl pyrophosphate
dimethylallyl pyrophosphate
CH CH2CH3C
CH3
CH CH2CCH2
CH3
CH CH2 O P O P O
O
O
O
O
CCH2
CH3
2
O
NADP+
O2 H2O
HO
H+
NADPH
NADP+ + 2 PP i
NADPH
2 farnesyl pyrophosphate
squalene 2,3-oxidosqualene lanosterol
O
NADP+
O2 H2O
HO
H+NADPH
squalene 2,3-oxidosqualene lanosterol
H O H O
lan o ste ro l cho leste ro l
1 9 s tep s
REGLAREA REGLAREA ŞŞI I PATOLOGIAPATOLOGIA
METABOLISMULMETABOLISMULUI LIPIDICUI LIPIDIC
ObiectiveleObiectivele Metabolismul eicosanoizilor. Căile ciclooxigenazică şi lipooxigenazică ale Metabolismul eicosanoizilor. Căile ciclooxigenazică şi lipooxigenazică ale
biosintezei lor. Inactivarea.biosintezei lor. Inactivarea. Metabolismul vitaminelor liposolubile: sursele alimentare, necesităţile Metabolismul vitaminelor liposolubile: sursele alimentare, necesităţile
diurne, transformărilediurne, transformările Reglarea metabolismului lipidelor la nivelul celulei.Reglarea metabolismului lipidelor la nivelul celulei. Reglarea neurohormonală a metabolismului lipidelor. Rolul lipotropinelor, Reglarea neurohormonală a metabolismului lipidelor. Rolul lipotropinelor,
ACTH, hormonilor tiroizi, insulinei, glucagonului, glucocorticoizilor şi ACTH, hormonilor tiroizi, insulinei, glucagonului, glucocorticoizilor şi catecolaminelor.catecolaminelor.
Relaţiile reciproce dintre metabolismul energetic, glucidic şi lipidic.Relaţiile reciproce dintre metabolismul energetic, glucidic şi lipidic. Dereglările digestiei şi absorbţiei lipidelor. Steatoreea pancreatică, hepatică Dereglările digestiei şi absorbţiei lipidelor. Steatoreea pancreatică, hepatică
şi intestinală.şi intestinală. Dislipidemiile:Dislipidemiile: a) hipolipoproteinemiile familiale – afecţiunea Tangier, a) hipolipoproteinemiile familiale – afecţiunea Tangier, - şi - şi --
lipoproteinemia familială;lipoproteinemia familială; b) hiperlipoproteinemiile primare şi familiale;b) hiperlipoproteinemiile primare şi familiale; c) hiperlipoproteinemiile secundare (dobândite) – în diabet zaharat, c) hiperlipoproteinemiile secundare (dobândite) – în diabet zaharat,
alcoolism, afecţiuni ale glandelor endocrine. alcoolism, afecţiuni ale glandelor endocrine. Cauze, mecanismele dereglării metabolismului lipidelor, manifestările Cauze, mecanismele dereglării metabolismului lipidelor, manifestările
biochimice.biochimice. 6. Lipidozele tiszlare:6. Lipidozele tiszlare: a) ereditare – Neimann-Pick, Tay-Sachs, Krabbe, Gaucher, Farber, a) ereditare – Neimann-Pick, Tay-Sachs, Krabbe, Gaucher, Farber,
leucodistrofia metacromatică, gangliozidoza GM1;leucodistrofia metacromatică, gangliozidoza GM1; b) dobândite – obezitate, ateroscleroză, alcoolism. b) dobândite – obezitate, ateroscleroză, alcoolism. Cauze, mecanismele dereglării metabolismului lipidelor, manifestările Cauze, mecanismele dereglării metabolismului lipidelor, manifestările
biochimice.biochimice. 7. A-, hipo- şi hipervitaminozele A, D, E, K – cauze, manifestări metabolice.7. A-, hipo- şi hipervitaminozele A, D, E, K – cauze, manifestări metabolice. 8. Rolul eicosanoizilor în procesele inflamatorii, reacţiile alergice, dereglările 8. Rolul eicosanoizilor în procesele inflamatorii, reacţiile alergice, dereglările
fluidităţii sanguine.fluidităţii sanguine.
Metabolismul Metabolismul eicosanoiziloreicosanoizilor
ATP-dependent carboxylation provides energy ATP-dependent carboxylation provides energy input. input. TheThe COCO22 is lost later during condensation with is lost later during condensation with the growing fatty acid. the growing fatty acid. The spontaneous decarboxylation drives the The spontaneous decarboxylation drives the condensation reaction. condensation reaction.
H 3C C SC o A
O
C H 2 C SC o A
O
O O C
acetyl-C oA
m alonyl-C oA
The input to fatty acid The input to fatty acid synthesis is synthesis is acetyl-acetyl-CoACoA, which is , which is carboxylated to carboxylated to malonyl-CoAmalonyl-CoA. .
HCOHCO33 + + ATPATP + acetyl-CoA+ acetyl-CoA ADPADP + P+ Pii + +
malonyl-CoAmalonyl-CoA
ll
Enzyme-biotin HCO3
- + ATP
ADP + Pi Enzyme-biotin-CO2
-
O CH3-C-SCoA acetyl-CoA O -O2C-CH2-C-SCoA malonyl-CoA
ll
Enzyme-biotin
1
2
Biotin Biotin is linked to the enzyme by an amide bond is linked to the enzyme by an amide bond between the terminal carboxyl of the biotin side between the terminal carboxyl of the biotin side chain and the chain and the -amino group of a -amino group of a lysinelysine residue. residue. The combined biotin and lysine side chains act as a The combined biotin and lysine side chains act as a long flexible armlong flexible arm that allows the biotin ring to that allows the biotin ring to translocate between the 2 active sites. translocate between the 2 active sites.
CHCH
H2CS
CH
NHC
N
O
(CH2)4 C NH (CH2)4 CH
CO
NH
O
CO
O
Carboxybiotin lysine residue
Acetyl-CoA CarboxylaseAcetyl-CoA Carboxylase, which converts acetyl-, which converts acetyl-CoA to malonyl-CoA, is the CoA to malonyl-CoA, is the committed stepcommitted step of the of the fatty acid synthesis pathway. fatty acid synthesis pathway.
The mammalian enzyme is The mammalian enzyme is regulatedregulated, by , by phosphorylationphosphorylation allosteric control by local metabolites.allosteric control by local metabolites.
Conformational changesConformational changes associated with associated with regulation:regulation: In the In the activeactive conformation, Acetyl-CoA conformation, Acetyl-CoA
Carboxylase associates to form multimeric Carboxylase associates to form multimeric filamentousfilamentous complexes. complexes.
Transition to the Transition to the inactiveinactive conformation is conformation is associated with dissociation to yield the associated with dissociation to yield the monomericmonomeric form of the enzyme (protomer). form of the enzyme (protomer).
The The decreaseddecreased production of production of malonyl-CoAmalonyl-CoA prevents prevents energy-utilizing fatty acid synthesis when cellular energy-utilizing fatty acid synthesis when cellular energy stores are depleted. (AMP is abundant only energy stores are depleted. (AMP is abundant only when ATP has been extensively dephosphorylated.)when ATP has been extensively dephosphorylated.)
AMP-Activated Kinase catalyzes phosphorylation of Acetyl-CoA Carboxylase, causing inhibition.
Phosphorylated protomer of Acetyl-CoA Carboxylase (inactive) Dephosphorylated Polymer of Acetyl-CoA Carboxylase (active)
Citrate
Dephosphorylated, e.g., by insulin-
activated Protein Phosphatase
Palmitoyl-CoA
Phosphorylated, e.g., via AMP-activated Kinase when cellular stress or exercise depletes ATP.
Regulation of Acetyl-CoA Carboxylase
When AMP is high (ATP low), malonyl-CoA production is When AMP is high (ATP low), malonyl-CoA production is diminished, releasing fatty acid oxidation from inhibition. diminished, releasing fatty acid oxidation from inhibition. This will lead to This will lead to increased ATP productionincreased ATP production..
AMP-Activated KinaseAMP-Activated Kinase has a significant role has a significant role even in tissues (e.g., even in tissues (e.g., cardiac muscle) that do cardiac muscle) that do not significantly not significantly synthesize fatty acids. synthesize fatty acids. In such tissues In such tissues malonyl-malonyl-CoACoA, produced via one , produced via one isoform of Acetyl-CoA isoform of Acetyl-CoA Carboxylase, functions Carboxylase, functions mainly as an mainly as an inhibitorinhibitor of of fatty acid oxidationfatty acid oxidation. .
H3C C SCoA
O
CH2 C SCoA
O
OOC
acetyl-CoA
malonyl-CoA
ATP + HCO3
ADP + Pi
Acetyl-CoA Carboxylase (inhibited by
AMP-Activated Kinase)
A cAMP cascade, activated by glucagon & A cAMP cascade, activated by glucagon & epinephrine when blood glucose is low, may also epinephrine when blood glucose is low, may also result in phosphorylation of Acetyl-CoA Carboxylase result in phosphorylation of Acetyl-CoA Carboxylase via via cAMP-Dependent Protein KinasecAMP-Dependent Protein Kinase..With Acetyl-CoA Carboxylase inhibited, acetyl-CoA With Acetyl-CoA Carboxylase inhibited, acetyl-CoA remains available for synthesis of ketone bodies, remains available for synthesis of ketone bodies, the alternative metabolic fuel used when blood the alternative metabolic fuel used when blood glucose is low.glucose is low.
H3C C SCoA
O
CH2 C SCoA
O
OOC
acetyl-CoA
malonyl-CoA
The antagonistic effect of The antagonistic effect of insulininsulin, produced , produced when blood glucose is high, is attributed to when blood glucose is high, is attributed to activation of Protein Phosphatase. activation of Protein Phosphatase.
Phosphorylated protomer of Acetyl-CoA Carboxylase (inactive) Dephosphorylated Polymer of Acetyl-CoA Carboxylase (active)
Citrate
Dephosphorylated, e.g., by insulin-
activated Protein Phosphatase
Palmitoyl-CoA
Phosphorylated, e.g., via AMP-activated Kinase when cellular stress or exercise depletes ATP.
Regulation of Acetyl-CoA Carboxylase
Palmitoyl-CoAPalmitoyl-CoA (product of Fatty Acid Synthase) (product of Fatty Acid Synthase) promotes the promotes the inactive inactive conformation, diminishing conformation, diminishing production of malonyl-CoA, the precursor of fatty acid production of malonyl-CoA, the precursor of fatty acid synthesis. synthesis. This is an example of This is an example of feedback inhibitionfeedback inhibition..
Regulation of Acetyl-CoA Carboxylase by local metabolites:
Phosphorylated protomer of Acetyl-CoA Carboxylase (inactive) Dephosphorylated Polymer of Acetyl-CoA Carboxylase (active)
Citrate
Dephosphorylated, e.g., by insulin-
activated Protein Phosphatase
Palmitoyl-CoA
Phosphorylated, e.g., via AMP-activated Kinase when cellular stress or exercise depletes ATP.
Regulation of Acetyl-CoA Carboxylase
[Citrate] is high when there is adequate [Citrate] is high when there is adequate acetyl-CoA entering Krebs Cycle. acetyl-CoA entering Krebs Cycle. Excess acetyl-CoA is then converted via Excess acetyl-CoA is then converted via malonyl-CoA to fatty acids for storage. malonyl-CoA to fatty acids for storage.
Glucose-6-phosphatase glucose-6-P glucose Gluconeogenesis Glycolysis pyruvate fatty acids
acetyl CoA ketone bodies cholesterol oxaloacetate citrate
Krebs Cycle
CitrateCitrate allosterically allosterically activatesactivates Acetyl- Acetyl-CoA Carboxylase. CoA Carboxylase.
Fatty acid synthesisFatty acid synthesis from acetyl-CoA & malonyl- from acetyl-CoA & malonyl-CoA occurs by a series of reactions that are:CoA occurs by a series of reactions that are: in in bacteriabacteria catalyzed by catalyzed by 6 different enzymes 6 different enzymes
plus a separate acyl carrier protein (ACP) plus a separate acyl carrier protein (ACP) in in mammalsmammals catalyzed by individual domains of catalyzed by individual domains of
a very large polypeptide that includes an ACP a very large polypeptide that includes an ACP domain.domain.Evolution of the mammalian Fatty Acid Synthase Evolution of the mammalian Fatty Acid Synthase apparently has involved apparently has involved gene fusiongene fusion. .
NADPHNADPH serves as serves as electron donorelectron donor in the two in the two reactions involving substrate reduction. reactions involving substrate reduction. The NADPH is produced mainly by the Pentose The NADPH is produced mainly by the Pentose Phosphate Pathway. Phosphate Pathway.
Fatty AcidFatty AcidSynthase Synthase prosthetic groups: prosthetic groups: the the thiolthiol of the side- of the side-
chain of a chain of a cysteinecysteine residue of residue of Condensing Enzyme Condensing Enzyme domain.domain.
the the thiolthiol of of phosphopantetheiphosphopantetheinene, equivalent in , equivalent in structure to part of structure to part of coenzyme A. coenzyme A.
N
N N
N
NH2
O
OHO
HH
H
CH2
H
OPOPOH2C
O
O O
O
P
O
O O
C
C
C
NH
CH2
CH2
C
NH
CH3H3C
HHO
O
CH2
CH2
SH
O
-mercaptoethylamine
pantothenate
ADP-3'- phosphate
Coenzyme A
phosphopantetheine
H3N+ C COO
CH2
SH
H
cysteine
PhosphopantetheinPhosphopantetheinee (Pant) is covalently (Pant) is covalently linked via a phosphate linked via a phosphate ester to a serine OH ester to a serine OH of the of the acyl carrier acyl carrier proteinprotein domain of domain of Fatty Acid Synthase. Fatty Acid Synthase.
TheThe long flexible long flexible armarm of of phosphopantetheine phosphopantetheine helps its thiol to move helps its thiol to move from one active site to from one active site to another within the another within the complex. complex.
OPOH2C
O
OC
C
C
NH
CH2
CH2
C
NH
CH3H3C
HHO
O
CH2
CH2
SH
O
CH2 CH
NH
C O
-mercaptoethylamine
pantothenate
serine residue
phosphopantetheine of acyl carrier protein
phosphate
As each of the substrates acetyl-CoA & malonyl-CoA As each of the substrates acetyl-CoA & malonyl-CoA bind to the complex, the initial attacking group is bind to the complex, the initial attacking group is the oxygen of a the oxygen of a serineserine hydroxylhydroxyl group of the group of the Malonyl/acetyl-CoA Transacylase enzyme domain. Malonyl/acetyl-CoA Transacylase enzyme domain. Each acetyl or malonyl moiety is transiently in ester Each acetyl or malonyl moiety is transiently in ester linkage to this serine hydroxyl, before being linkage to this serine hydroxyl, before being transferred into thioester linkage with the transferred into thioester linkage with the phosphopantetheine thiolphosphopantetheine thiol of the acyl carrier of the acyl carrier protein (ACP) domain. protein (ACP) domain. Acetate is subsequently transferred to a Acetate is subsequently transferred to a cysteine cysteine thiolthiol of the Condensing Enzyme domain. of the Condensing Enzyme domain.
Condensing Malonyl/acetyl-CoA Dehydratase Enoyl -Ketoacyl ACP Thioesterase Enzyme (Cys) Transacylase (Ser) Reductase Reductase (Pant) N- -C
Order of domains in primary structure of mammalian Fatty Acid Synthase
The The condensationcondensation reaction (step 3) reaction (step 3) involves decarboxylation of the malonyl involves decarboxylation of the malonyl moiety, followed by attack of the resultant moiety, followed by attack of the resultant carbanion on the carbonyl carbon of the carbanion on the carbonyl carbon of the acetyl (or acyl) moiety. acetyl (or acyl) moiety.
Pant
SH
Cys
SH
Pant
SH
Cys
S
C
CH3
O
Pant
S
Cys
S
C
CH3
OC
CH2
COO
O
Pant
S
Cys
SH
C
CH2
C
O
CH3
O
acetyl-S-CoA HS-CoA malonyl-S-CoA HS-CoA CO2
1 2 3
1 Malonyl/acetyl-CoA-ACP Transacylase 2 Malonyl/acetyl-CoA-ACP Transacylase 3 Condensing Enzyme (-Ketoacyl Synthase)
4.4. The The -ketone is -ketone is reducedreduced to an alcohol by e to an alcohol by e transfer from NADPH. transfer from NADPH.
5.5. DehydrationDehydration yields a trans double bond. yields a trans double bond. 6.6. ReductionReduction by NADPH yields a saturated chain. by NADPH yields a saturated chain.
Pant
S
Cys
SH
C
CH2
C
O
CH3
O
Pant
S
Cys
SH
C
CH2
HC
O
CH3
Pant
S
Cys
SH
Pant
S
Cys
SH
NADPH NADP+NADPH NADP+
C
CH
HC
O
CH3
C
CH2
CH2
O
CH3
OH
H2O
4 5 6
4 -Ketoacyl-ACP Reductase 5 -Hydroxyacyl-ACP Dehydratase 6 Enoyl-ACP Reductase
Following Following transfertransfer of the growing fatty acid of the growing fatty acid from phosphopantetheine to the Condensing from phosphopantetheine to the Condensing Enzyme's cysteine sulfhydryl, the Enzyme's cysteine sulfhydryl, the cycle cycle begins againbegins again, with another malonyl-CoA. , with another malonyl-CoA.
Pant
S
Cys
SH
C
CH2
CH2
O
CH3
Pant
SH
Cys
S
C
CH2
O
CH2
CH3
Pant
S
Cys
S
C
CH2
O
CH2
CH3
C
CH2
COO
O
Malonyl-S-CoA HS-CoA
7 2
7 Condensing Enzyme 2 Malonyl/acetyl-CoA-ACP Transacylase (repeat).
Product release:Product release: When the fatty acid is 16 carbon atoms long, When the fatty acid is 16 carbon atoms long, a a ThioesteraseThioesterase domain catalyzes hydrolysis domain catalyzes hydrolysis of the thioester linking the fatty acid to of the thioester linking the fatty acid to phosphopantetheine. phosphopantetheine. The The 16-C16-C saturated fatty acid saturated fatty acid palmitatepalmitate is is the final product of the Fatty Acid Synthase the final product of the Fatty Acid Synthase complex. complex.
The The primary structureprimary structure of the mammalian Fatty Acid of the mammalian Fatty Acid Synthase protein is summarized above.Synthase protein is summarized above.Fatty Acid SynthaseFatty Acid Synthase in mammals is a in mammals is a homo-dimerhomo-dimer. .
Condensing Malonyl/acetyl-CoA Dehydratase Enoyl -Ketoacyl ACP Thioesterase Enzyme (Cys) Transacylase (Ser) Reductase Reductase (Pant) N- -C
Order of domains in primary structure of mammalian Fatty Acid Synthase
KR KR
DH DH KS KS MAT MAT
ER ER
Arrangement of domains in Fatty Acid Synthase
“arm” “leg”
X-RayX-Ray crystallographic crystallographic analysis at 4.5 Å analysis at 4.5 Å resolution shows the resolution shows the dimeric Fatty Acid dimeric Fatty Acid Synthase to have an Synthase to have an X-shapeX-shape, with , with domains arranged as domains arranged as summarized at right. summarized at right.
The solved structure does The solved structure does not resolvenot resolve the position the position of of ACPACP & & ThioesteraseThioesterase domains, predicted from domains, predicted from primary structure to be near primary structure to be near -Ketoacyl Reductase -Ketoacyl Reductase (KR) domains of lateral "arms" of the complex. (KR) domains of lateral "arms" of the complex.
Condensing Malonyl/acetyl-CoA Dehydratase Enoyl -Ketoacyl ACP Thioesterase Enzyme (Cys) Transacylase (Ser) Reductase Reductase (Pant) N- -C
Order of domains in primary structure of mammalian Fatty Acid Synthase
KR KR
DH DH KS KS MAT MAT
ER ER
Arrangement of domains in Fatty Acid Synthase
“arm” “leg”
These domains may These domains may be too flexible to be be too flexible to be resolved. resolved. KR = KR = -Ketoacyl -Ketoacyl Reductase; ER = Enoyl Reductase; ER = Enoyl Reductase; Reductase; DH = Dehydratase; DH = Dehydratase; KS = KS = -Ketoacyl Synthase -Ketoacyl Synthase (Condensing Enzyme); (Condensing Enzyme); MAT = Malonyl/Acetyl-MAT = Malonyl/Acetyl-CoA Transacylase. CoA Transacylase.
KR KR
DH DH KS KS MAT MAT
ER ER
Arrangement of domains in Fatty Acid Synthase
“arm” “leg”
Fatty Acid Synthase complex is somewhat Fatty Acid Synthase complex is somewhat asymmetricasymmetric..There is evidence for There is evidence for conformational changesconformational changes relating to catalysis. relating to catalysis. Protein flexibilityProtein flexibility may facilitate transfer of ACP- may facilitate transfer of ACP-attached reaction intermediates among the several attached reaction intermediates among the several active sites in each half of the complex. active sites in each half of the complex.
For images see:website (ETH Zurich)
website (Asturias lab, Scripps)
article (Maier, Jenni & Ban; requires subscription to Science).
ExploreExplore with Chime the structure of with Chime the structure of the the E. coli E. coli -Ketoacyl-ACP Synthase -Ketoacyl-ACP Synthase III, equivalent to the domains of the III, equivalent to the domains of the mammalian Fatty Acid Synthase that mammalian Fatty Acid Synthase that catalyze the initial acetylation and catalyze the initial acetylation and condensation reactions. condensation reactions.
-Oxidation & Fatty Acid -Oxidation & Fatty Acid SynthesisSynthesisComparedCompared
Oxidation Pathway Fatty Acid Synthesis
pathway location mitochondrial matrix cytosol
acyl carriers (thiols) Coenzyme-A phosphopantetheine
(ACP) & cysteine
e acceptors/donor FAD & NAD+ NADPH
-OH intermediate L D
2-C product/donor acetyl-CoA malonyl-CoA (& acetyl-CoA)
Fatty Acid Synthase is Fatty Acid Synthase is transcriptionally transcriptionally regulatedregulated. . In liver:In liver: InsulinInsulin, a hormone produced when blood , a hormone produced when blood
glucose is high, glucose is high, stimulatesstimulates Fatty Acid Synthase Fatty Acid Synthase expression.expression.Thus excess glucose is stored as fat.Thus excess glucose is stored as fat.Transcription factors that that mediate the Transcription factors that that mediate the stimulatory effect of insulin include stimulatory effect of insulin include USFsUSFs (upstream stimulatory factors) and (upstream stimulatory factors) and SREBP-1SREBP-1. . SREBPsSREBPs (sterol response element binding (sterol response element binding proteins) were first identified for their proteins) were first identified for their regulation of cholesterol synthesis.regulation of cholesterol synthesis.
Polyunsaturated Polyunsaturated fatty acidsfatty acids diminishdiminish transcription of the Fatty Acid Synthase gene in transcription of the Fatty Acid Synthase gene in liver cells, by suppressing production of SREBPs. liver cells, by suppressing production of SREBPs.
In fat cellsIn fat cells::Expression of SREBP-1 and of Fatty Acid Expression of SREBP-1 and of Fatty Acid Synthase is Synthase is inhibitedinhibited by by leptinleptin, a hormone , a hormone that has a role in regulating food intake and fat that has a role in regulating food intake and fat metabolism. metabolism. LeptinLeptin is produced by fat cells in response to is produced by fat cells in response to excess fat storage. excess fat storage. Leptin regulates body weight by decreasing Leptin regulates body weight by decreasing food intake, increasing energy expenditure, and food intake, increasing energy expenditure, and inhibiting fatty acid synthesis. inhibiting fatty acid synthesis.
ElongationElongation beyond the 16-C length of the palmitate beyond the 16-C length of the palmitate product of Fatty Acid Synthase occurs in mitochondria product of Fatty Acid Synthase occurs in mitochondria and endoplasmic reticulum (ER). and endoplasmic reticulum (ER). Fatty acid elongation within Fatty acid elongation within mitochondriamitochondria involves involves
the the -oxidation pathway-oxidation pathway running in reverse, but running in reverse, but NADPH serves as electron donor for the final NADPH serves as electron donor for the final reduction step. reduction step.
Polyunsaturated fatty acids esterified to CoA are Polyunsaturated fatty acids esterified to CoA are substrates for the substrates for the ER elongation machineryER elongation machinery, , which uses malonyl-CoA as donor of 2-carbon units. which uses malonyl-CoA as donor of 2-carbon units. The reaction sequence is similar to Fatty Acid The reaction sequence is similar to Fatty Acid Synthase but individual steps are catalyzed by Synthase but individual steps are catalyzed by separate proteinsseparate proteins. . A family of enzymes designated A family of enzymes designated Fatty Acid Fatty Acid ElongasesElongases catalyze the initial condensation step for catalyze the initial condensation step for elongation of saturated or polyunsaturated fattyelongation of saturated or polyunsaturated fatty acidsacids..
DesaturasesDesaturases introduce introduce double bondsdouble bonds at at specific positions in a fatty acid chain.specific positions in a fatty acid chain.
Mammalian cells are unable to produce Mammalian cells are unable to produce double bonds at certain locations, e.g., double bonds at certain locations, e.g., 1212. .
Thus some polyunsaturated fatty acids are Thus some polyunsaturated fatty acids are dietary essentialsdietary essentials, e.g., linoleic acid, 18:2 , e.g., linoleic acid, 18:2 cis cis 9,129,12 (18 C atoms long, with cis double (18 C atoms long, with cis double bonds at carbons 9-10 & 12-13). bonds at carbons 9-10 & 12-13).
CO
OH910
oleate 18:1 cis 9
Formation of a double bond in a fatty acid involves Formation of a double bond in a fatty acid involves the following endoplasmic reticulum membrane the following endoplasmic reticulum membrane proteins in mammalian cells:proteins in mammalian cells: NADH-cyt bNADH-cyt b5 5 ReductaseReductase, a flavoprotein with , a flavoprotein with
FADFAD as prosthetic group. as prosthetic group. Cytochrome bCytochrome b55, which may be a separate protein , which may be a separate protein
or a domain at one end of the desaturase. or a domain at one end of the desaturase. DesaturaseDesaturase, with an active site that contains , with an active site that contains two two
iron atomsiron atoms complexed by histidine residues. complexed by histidine residues.
CO
OH910
oleate 18:1 cis 9
The desaturase catalyzes a The desaturase catalyzes a mixed function mixed function oxidationoxidation reaction. reaction. There is a There is a 4-electron4-electron reduction of reduction of OO22 2 H2 H22OO as as a fatty acid is oxidized to form a double bond. a fatty acid is oxidized to form a double bond.
2e2e pass from pass from NADHNADH to the desaturase via the to the desaturase via the FAD-containing reductase & cytochrome b FAD-containing reductase & cytochrome b55, , the order of electron transfer being: the order of electron transfer being: NADHNADH FADFAD cyt bcyt b55 desaturasedesaturase
2e2e are extracted from the fatty acid as the are extracted from the fatty acid as the double bond is formed.double bond is formed.
E.g., the overall reaction for desaturation of E.g., the overall reaction for desaturation of stearate (18:0) to form oleate (18:1 cis stearate (18:0) to form oleate (18:1 cis 99) is:) is:stearate + NADH + Hstearate + NADH + H++ + O + O22 oleate + NADoleate + NAD++ + 2H+ 2H22OO
