Describe the differences between B-Oxidation and Fatty Acid Chain Synthesis in terms of:
1. Location
2. Acyl Carrier
3. Carbon Units
4. Product
5. Substrate
6. Oxidizing Agents
7. Reducing Agent
8. Enyzme Proteins
Beta Ox vs. Synthesis
1. Location: mitochondria vs. cytoplasm
2. Acyl Carrier: CoA vs. Acyl Carrier Prot
3. Carbon Units: 2 vs. 2
4. Product: Acetyl-CoA (only Beta-oX)
5. Substrate: Malonyl-CoA (only Synthesis)
6. Oxidizing Agents: NAD+, FAD (Beta-Ox)
7. Reducing Agent: NADPH (Synthesis)
8. Enzyme Proteins: Separate vs. Single, Multifn'l
Describe the reaction sequence for the formation of Malonyl-CoA from Acetyl-CoA
must be activated, via a carboxylation reaction, to Malonyl CoA
-Acetyl CoA
-Acetyl CoA Carboxylase catalyzes the reaciton using a biotin prosthetic group
Acetyl-CoA + ATP + CO2 → Malonyl-CoA + ADP + Pi
Malonyl-CoA is committed to fatty acid synthesis
Recognize the similarities and differences between the structures of acyl-carrier protein and
coenzyme A
-ACP and CoA both serve as carriers during their respective processes
-Both have Pantetheine (pantothenic acid) group and use a high energy thioester bond (SH)
-ACP is linked to its reactive protein region via a serine
-CoA has no protein region, but has 3`P-ADP
Reaction 1 of fatty acid synthesis
ACETYL TRANSACYLASE (AT)
Begin with: Acetyl-CoA and HS-CE (condensing enzyme)
-cleaves the Acetyl group from the CoA and puts it on the CE
Rxn 2 of Fatty Acid Synthesis
Malonyl Transacylase (MT)
-cleaves the malonyl group from the CoA and puts it on the Acyl Carrier Protein (ACP)
Rxn 3 of Fatty Acid Synthesis
Acyl-Malonyl-ACP Condensing Enzyme
Begin with: Acetyl-CE (rxn 1) and Malonyl-ACP (rxn 2)
-A proton attaches to the S group of Acetyl-CE, breaking off S-CE from Acetyl group and forming HS-CE
-Decarboxylation (-CO2) of the Malonyl-ACP yields Acetyl-ACP and that attaches to the now free Acetyl
group
End with:
Acetoacetyl-ACP (CH3-C(=O)-CH2-C(=O)-S-ACP
+ HS-CE
Rxn 4 of Fatty Acid Synthesis
B-Ketoacyl-ACP Reductase
-NADPH + H+ used to reduce the Acetoacetyl-ACP (an H is added to the double-bonded O on the beta
carbon and to the beta carbon itself, thus beta carbon has only single bonds now)
End-products:
CH3-CH(-OH)-CH2-C(=O)-S-ACP
D-3-hydroxybutyrl-ACP
Rxn 5 of Fatty Acid Synthesis
3-Hydroxyacyl-ACP Dehydratase
-removes the hydroxyl group from the beta carbon (C3) and removes an H from the alpha carbon to
allow a new double bond to form between alpha and beta carbons
end with:
CH3-CH=CH-C(=O)-S-ACP + H2O
Crotonyl-ACP (trans-Δ2-enoyl-ACP)
Rxn 6 of Fatty Acid Synthesis
Enoyl-ACP Reductase
-6th rxn is end of the first cycle
-NADPH +H+ again reduces the substrate, Crotonyl-ACP to get rid of the double bond between alpha and
beta carbon
end with:
CH3-CH2-CH2-C(=O)-S-ACP
butyryl-ACP
so now you've created a fatty acid chain with 2 more C's on it. You can now repeat Rxn 3 with AcylMalonyl-ACP Condensing Enzyme to react a butyryl-CE (C4) with another malonyl-ACP (C3) to end up
with a C6, etc.
Rxn 7 of Fatty Acid Synthesis
for the last cycle, use THIOESTERASE
-Say you start with palmityl-ACP (16 C)
-add H2O to cleave off the -S-ACP group by reducing the thioester bond.
End with:
CH3-(CH2)14-COOH + HS-ACP
Palmitate
Describe the domain structure of fatty acid synthase
-Growing chain on one ACP gets worked on by the reduction unit domain of the opposing strand
-Additional 2-carbon units always come from decarboxylation of Malonyl-ACP
-Growing chain always leaves ACP and goes back to condensing enzyme, making room for Malonyl-CoA
to bind to ACP (via Malonyl-CoA Transacylase)
Describe the stoichiometric summary reactions for the Activation Phase and the Elongation Phase of
fatty acid synthesis
Activation using Acetyl-carboxylation:
7 Acetyl-CoA + 7 (HCO3- + H+) + 7ATP →
7 Malonyl-CoA + 7 ADP + 7 Pi + 7 (H2O + H+)
Elongation using Fatty Acid Synthase:
Acetyl-CoA + 7 Malonyl-CoA + 14 NADPH + 14 H+ →
Palmitate + 7 CO2 + 14 NADP+ + 8 CoA + 6 H2O
Where does the Acetyl-CoA and NADPH used in fatty acid synthesis come from?
formed from pyruvate inside mitochondria
Acetyl-CoA
during fed state (when glucose is available)
List the steps for transport of acetyl CoA from mitochondria to the cytosol via citrate 1. Citrate
Synthase Rxn in mitochondria:
Oxaloacetate + Acetyl CoA → Citrate
2. Citrate transport to cytoplasm:
When [citrate] high from rxn 1, TCA cycle is saturated and Citrate can be transported out of the
mitochondria, into the cytoplasm via transporter protein
3. ATP Citrate Lyase Reaction (in cytoplasm):
Citrate + CoA + ATP → ADP + Pi + Oxaloacetate + Acetyl CoA
Describe the cytoplasmic reactions that generate most of the NADPH required for fatty acid synthesis
Returning oxaloacetate to the mitochondria generates NADPH needed for FA synthesis in cytosol
Oxaloacetate cannot traverse the mitochondrial membrane
1. Malate dehydrogenase catalyzes reduction of Oxaloacetate to form Malate (NADH + H+ → NAD+)
2. Malic enzyme catalyzes oxidation of Malate by NADP+ to form pyruvate (NADP+ → NADPH + H+)
3. Pyruvate can traverse the mitochondrial membrane. Inside Pyruvate carboxylase catalyzes the
addition of CO2 to pyruvate to get oxaloacetate
Metabolic Regulation of Fatty Acid Synthesis
and fatty acids scarce
Synthesis of FA increases when carbs & energy plentiful
Synthesis of FA decreases when energy supplies are low due to starvation, exercise or deficient glucose
utilization
Describe the structure of linoleate and explain why it cannot be made in humans
body cannot synthesize linoleate (18:2 (9,12)) or Linolenate (18:3 (9,12,15)) because we do not have
enzymes to introduce a second double bond after C-9
eg. Palmitate and Stearate can make palmitoleate (16:1(9)) and oleate (18:1(9)) using mammalian
DESATURASE, but beyond C-9 cannot
Explain how linoleate can be converted to arachidonate through desaturation and elongation
reactions
Get Linoleate 18:2(9,12) from diet
1. In ER, use a desaturase to introduce DB at C-6: Octadecatrienoate 18:3(6,9,12)
2. In ER, use malonyl-CoA (2 C addition) in an elongation rxn: Eicosatrienoate 20:3(8,11,14)
3. Use another desaturase to introduce DB at C-5: Arachidonate 20:4(5,8,11,14)
Describe the regulatory effects of citrate, insulin, epinephrine and glucagon, on the activity of AcetylCoA Carboxylase
citrate: upregulates ACC, promoting aggreagation when acetyl-CoA + ATP levels are high (well-fed)
insulin: upregulates ACC, activates Protein Phosphatase 2A which dephosphorylates ACC to activate it
epinephrine: inhibits ACC, activates Protein Kinase A which inhibits protein phosphatase 2A which
dephosphorylates ACC to activate it. Therefore epinephrine inhibits ACC
glucagon: inhibits ACC
Local Regulators of Acetyl-CoA Carboxylase
sequence)
INHIBITOR: Fatty Acyl-CoA (product of reaction
UPREGULATOR: Citrate (promotes aggregation) when acetyl-CoA + ATP levels are high (well-fed)
NOTE: malonyl-CoA also blocks Beta-Oxidation by blocking the CPT-I
Global Regulators of Acetyl-CoA Carboxylase
INHIBITORS:
1. Amp-activated Protein Kinase:
-phosphorylates Acetyl-CoA Carboxylase, inactivating it
-turned off during times of high [ATP], keeping ACC in active, unphosphorylated state (high ATP = FA
Synthesis)
2. Protein Kinase A:
-inhibits protein phosphatase 2A
-active during times of fight/flight, starvation
ACTIVATORS:
1. Protein Phosphatase 2A:
-dephosphorylates (actives) ACC
-active during "fed" state, responds to insulin