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55 terms

Fatty Acid synthesis and Cholesterol biosynthesis

Lectures 45 and 46
Where does fatty acid biosynthesis occur?
brain, kidney, mammary glands, intestine, adipose tissue
Main site of synthesis
Problems associated with neonatal digestion
-Low pancreatic secretion of lipase
-Immature liver unable to produce bile
Overcome problems of neonatal digestion
- very active gastric lipase
- breast milk rich in MCFAs
- Breast milk also contains bile salt-stimulated lipase
Enzymes in De-novo Fatty acid synthesis
- ACCase (acetyl-CoA carboxylase)
- Fatty acid synthase (FAS)
End product of Fatty acid synthesis
Modifications of palmitate
This takes place in the ER

**In mitochondria - Acyl oxidation, acetyl-CoA production, ketogenesis
**Cytosol - NADPH production, FA synthesis, isoprenoid synthesis
Where does fatty acid de novo synthesis take place?
Simplified version of fatty acid synthesis
1. Acetyl CoA» Palmitate (enz = ACCase and FAS)
2. Palmitate » Unsaturated fat (desaturase) »
LCFA, VLCFA (elongase)
Palmitate » LCFA, VLCFA (elongase)
How are acetyl CoA made in the mitochondria transported to the cytosol for FA synthesis?
Malate-citrate shuttle
Malate-citrate shuttle
1. Acetyl-CoA + OAA » Citrate (TCA) (Citrate synthase)
2. Citrate crosses mitochondrial membrane » cytosol
3. Citrate » acetyl CoA + OAA (ATP-Citrate lyase)

Citrate + CoASH + ATP » Acetyl-CoA + OAA + ADP + Pi

(ATP- Citrate lyase) Cytosolic reaction
Why can FAs not be converted to carbohydrate in animals?
*** Acetyl CoA cannot be converted to pyruvate/ OAA
What happens to th OAA released into the cytosol from the citrate lyase reaction?
1. OAA + NADH » Malate +NAD (malate dehydrogenase) --- Malate diffuses through membrane into mitochondria

Also in cytosol
2. Malate + NADP » Pyruvate (Malic enzyme)
CArboxylation of Acetyl-CoA by ACCase in cytosol
1. Acetyl-CoA » Malonyl-CoA ( ACCase)

**ACCase is the only enzyme involved in FA synthesis that is separate from the multifunctional enzyme FAS (Fatty acid synthase)
Components of ACCase
- Biotin carboxylase
- transcarboxylase
-Biotin-carboxyl carrier protein ( BCCP)
Human ACCase
- multifunctional protein
- exists in inactive form
-active when it forms a filamentous polymer

**Rate of FA synthesis dependent on the eqm of those 2 forms:

Protomer (inactive) »« Polymer (active)
ACCase reation
2 phases:

1. Carboxylation of biotin to form carboxybiotin
(Biotin carboxylase)

ATP + HCO₃⁻ + BCCP » CO₂---BCCP + ADP + Pi

2. Transcarboxylation of biotin

CO₂---BCCP + acetyl-CoA » malonyl-CoA + BCCP
Overall ACCase reaction
ACetyl Co-A (2C) + ATP + CO₂ » Malonyl-CoA (3C) + ADP + Pi
FAS (Fatty acid synthase)
** located in cytoplasm
**Several sequential reactions
Simplified FAS reaction
1. ACetyl CoA (2C) »Malonyl CoA (3C) (ACCase)
2. Malonyl CoA (3C) » Palmitic Acid (C16:0) (FAS using ACP intermediates)
3. Palmitic acid » Palmitoyl-CoA (thioester bond formation) *** increases the solubility.
Characteristics of FAS enzyme
- phosphopantetheine binding domain (ACP - acyl carrier protein)
- 2 thiol grps must be loaded onto acyl groups before condensation
- acyl grp from acyl-CoA initially transferred to ACP
- acyl grp then transferred from ACP to ketoacyl-ACP synthase
- malonyl grp from malonyl-CoA transferred to thiol grp of ACP
***ACP arm is flexible - move substrate to active site.
*** FAS has several active sites - 2 FAS work in pairs.
Preliminary reactions of FA synthesis
1. Acetyl-CoA + ACP » Acetyl-ACP + HS-CoA
2. Malonyl-CoA + ACP » Malonyl-ACP + HS-CoA
Condensation reactions of FA synthesis
acetyl synthase (2C) + malonyl-ACP(3C) » acetoacetyl-ACP(4C) + synthase-SH + CO₂(1C)

* enzyme - β-ketoacyl-ACP synthase.
1st Reduction reaction of FA synthesis
acetoacetyl-ACP + NADPH + H⁺ «» βhydroxybutyryl-ACP + NADP⁺

**Enz- βketoacyl-ACP reductase
Dehydration reaction
βhydroxybutyryl-ACP »« crotonyl-ACP + H₂O

enz- βketoacyl-ACP dehydratase
2nd reduction reaction
crotonyl-ACP + NADPH + H⁺ »« butyryl-ACP (4C) + NADP⁺

enz- βenoyl-ACP reductase
Fate of butyryl-ACP
-enters 2nd round of FA synthesis similar to acetyl-ACP

*** Final product is PALMITOYL-ACP.
-palmitoyl-ACP » palmitic acid (thioesterase)

(All C's in palmitic acid are derived from malonyl-CoA except 2C's at methyl end (from original acetyl CoA molecule)
Effect of malonyl Co-A on CPT I
** inhibitory effect on CPT I
- prevents acetyl-CoA from β-oxidation

**Glucagon activates CPT I (liver)
Effect of insulin on FA synthesis
Insulin »» Acetyl CoA » Malonyl CoA » Pamitate

*Malonyl-CoA »»» inhibit CPT I»»»» no acetyl CoA goes into mitochondria»»» stays in cytosol for FA synthesis instead of oxidation
Elongation reactions of FA.
2 pathways:

1. ER (using malonyl-CoA)
2. Mitochondrial (uses acetyl-CoA)
Desaturation of FA reactions
-2 H⁺ removed and H₂O produced
- NADPH used

**Acyl chain must be 16 or 18 C's before desaturation occurs.
Why can mammals not synthesize linoleic and α-linoleic acid?
- mammals cannot introduce Δ¹⁵ and Δ¹² double bonds.
-linoleate (C18:2) and α-linoleic (18:3) have this.

*** mammals can only synthesize up to Δ⁹ double bonds btwn the COOH and Δ⁹ of CH of chain.

**possible to desaturate oleate at Δ⁶ forming -
C18:2Δ⁶,⁹ FA.
Specific process : FA desaturation
Steroyl-CoA (C18:0) »»»oleoyl-CoA (C18:1 Δ⁹) + 2H₂O

*Aerobic desaturation
*microsomal enzymes
*Δ⁹ desaturation
Functions of essential FA's
- membrane structure
- specific enzyme-protein interaction in membranes
- synth eicosanoids
-syn arachidonic acid (C20:4)
-synth docosahexanoic acid (DHA C22:6)
polyunsaturated FA
- not all are converted to eicosanoids bc of limiting activites of some enzymes (elongases and Δ⁵ and Δ⁶ desaturases)
- precursors for resolvins, docosatrienes, neuroprtectins (anti-inflammatory properties)
***** Humans rely on exogenous source of EPA and DHA in diet.
- key PUFA in brain tissue
-imp for brain and Nervous tissue development.
- not regarded as EFAs although they have to be taken in exogenously
What is linoleic acid used to make?
- C18: 2n-6 (linoleic acid) » C18:3n-6 (γ linoleic acid) » C20:3n-6 » C20:3n-6» C20:4n-6 (Arachidonic acid) » C22: 5n-6
What is α-linolenic acid used to make?
-C18:3n-3 (α-linolenic acid) » C18:4n-3 » C20: 4n-3 » C20: 5n-3 (Eicosapentanoic acid, EPA) » C22: 6n-3 (Docosahexanoic acid, DHA)
When there is an EFA deficiency what happens?
Stearic acid is used to make Mead acid, causes dermatosis
Regulation of FA synthesis
-ACCase is key regulatory enzyme.
** strict regulation
1. Short term Rapid Response
2. Long term response
Short term rapid response FA regulation
-allosterically (stimulated) regulated by citrate
-inhibited by palmitoyl-CoA
-regulated by phosphorylation/dephosphorylation
(insulin, epinephrine, norepinephrine, glucagon)
-Activated by insulin in dephosphorylated form,
-Deactivated by glucagon and epinephrine in phosphorylated form.
Long term Response
- Increased expression of ACCase and FAS (at molecular level to maintain high carb diet)
TAG synthesis - Kennedy pathway
- takes place in the liver
- exported to peripheral tissue in blood
- Glycerol-3-Phosphate is immediate precursor for TAG synthesis.
** Glycerol-3-P derived directly from LPLase or indirectly from glycolysis.
- acyl-CoA transferred from cytosolic acyl-CoA to SER
-(TAG molecule is assembled here)
Formation of phosphatidic acid
-2 sequential acylations
1. Glycerol-3-P acyl transferase,
Glycerol-3-P + Acyl-CoA»» lysophoshatidic acid
2. Acylglycerol-3-P acyltransferase
Lysophosphatidic acid+ Acyl-CoA»» Phosphatidic acid

3. Phosphate removed by phosphatase
Phosphatidic acid»» Diacylglycerol + Pi
4. DAG acyl transferase
Diacylglycerol + Acyl-CoA»» Triacylglycerol
Specific to TAG biosynthesis
*** Acylation at the sn-3 position of DAG (DAG acyl transferase)
Important Sites of TAG synthesis
-SI (MAG pathway)
-Liver ( Kennedy pathway)
-Adipose tissue (Kennedy)
-Mammary gland (milk fat lactation) (Kennedy pathway)

** small amount of turnover in myocytes.
-TAG synthesis in enterocytes is primarily controlled by the rate of influx of exogenous dietery lipids.
-TAG synthesis in adipose tissue governed by the supply of glucose-derived glycerol-3-P.
MAG pathway in enterocytes
2 MAG + acyl-CoA »» DAG + Acyl-CoA »» TAG

** Enzyme = acyltransferase
Important points to note about FA synthesis
1. De novo FA synthesis from glucose (ACCase and FAS) --- not very active in adipocytes due to Western diet
2. TAG synthesis utilizing glycerol-3-phosphate
3. TAG mobilization by HSL
4. FA uptake from TAG-rich blood lipoproteins following LPLase activity.
Cholesterol facts
- most common steroid in animal cells
- absent from plant tissue and vegetable oils
- OH makes it amphipatic
- cholesteryl esters are hydrophobic
- vital in cell membranes
- precursor to steroid hormones
-cannot be degraded to CO₂
- liver converts it to bile acids
Plant sterols
- plants synthesize sterols: stigmasterol, campesterol, β-sistosterol
- not readily absorbed by human intestines
- inhibitory effect on cholesterol absorption
- use in cholesterol lowering drugs
Main site of cholesterol synthesis
Other sources of cholesterol
- dietary cholesterol
- Import of cholesterol from blood
Difference btwn testosterone and estrogen
- Testosterone has an =O grp while estrogen has an OH grp.
Cholesterol synthesis simplified
*** in cytosol
-------acetyl-CoA + acetoacetl-CoA » HMG-CoA

1. HMGCoA » Mevalonate (HMG-CoA reductase)
2. Mevalonate » C5 Isopentyl PPi
3. C5 Isopentyl PPi » C10 Geranyl PPi
4.C10 Geranyl PPi » C15 Farnesyl PPi
5. C15 Farnesyl PPi » Squalene
6Squalene » Lanosterol » Cholesterol

** Farnesyl is precursor for
- CoQ (Ubiquinone)
- Dolichol (glycoprotein synthesis)