Biosynthesis of fatty acids

/De-novo synthesis of fatty

acids/
Introduction to De-novo synthesis of Fatty
acids
 Diet supplies a large proportion of fatty acids used by the
body.

 Excess amounts of carbohydrate and proteins obtained

from diet in a well- fed state can be converted to fatty acids
which are stored as triacylglycerols (de- novo synthesis of
fatty acids from acetyl CoA obtained from the metabolism
of these fuels). In humans fatty acid synthesis occurs
primarily in the liver and lactating mammary gland and to
a lesser extent in the adipose tissue and kidney.
 The process incorporates carbons from acetyl CoA into a
growing fatty acid chain, utilizing ATP and NADPH.

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Fatty acid synthesis is not the exact reversal of β-oxidation.

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Sources of NADPH in the body
 NADPH is involved as donor of reducing equivalents

 The oxidative reactions of the pentose phosphate pathway are the chief
source of the hydrogen (NADPH) required for the reductive synthesis of
fatty acids.
 Tissues specializing in active lipogenesis i.e, liver, adipose tissue, and the
lactating mammary gland—possess an active pentose phosphate pathway.
 Other sources of NADPH include the reaction that converts malate to
pyruvate catalyzed by the "Malic enzyme" (NADP malate
dehydrogenase) and the extra mitochondrial isocitrate dehydrogenase
reaction (probably not a substantial source, except in ruminants).
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The PPP/HMP Pathway/ - Source of NADPH

In hepatocytes, adipose tissue and the lactating mammary glands,

the NADPH is supplied primarily by the pentose phosphate
pathway.

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The Malic enzyme- Source of NADPH

Reversible reaction, pyruvate produced in the reaction

reenters the mitochondrion for further utilization.
De - novo synthesis of fatty acids occurs in the cytosol of the cell.
 Fatty acids can be synthesized from acetyl-CoA. This is the
major way of utilizing excess dietary carbohydrates.
Production of cytosolic acetyl CoA:
1. The source of mitochondrial acetyl CoA is the oxidation of
pyruvate, degradation of fatty acids, ketone bodies or amino
acids. Acetyl-coA for fatty acids synthesis is never derived from
fatty acid oxidation rather it is from glucose (primarily) or
ketogenic amino-acids.
2. The Coenzyme A portion of acetyl CoA cannot cross the
mitochondrial membrane. Why? Only the acetyl portion is
transported to the cytosol in the form of citrate.

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3. A high mitochondrial concentration of citrate and ATP
enhances the de- novo synthesis of fatty acids.
Mitochondrial citrate concentration increases when
isocitrate dehydrogenase is inhibited by the presence
of large amounts of ATP, causing citrate and isocitrate
to accumulate. Therefore, cytosolic citrate may be
viewed as a high- energy signal. Also a large amount
of ATP is needed for fatty acid synthesis

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Transportation of Acetyl co A as citrate

Intra-mitochondrial acetyl-CoA first reacts with

oxaloacetate to form citrate, in the TCA cycle catalyzed by
citrate synthase.
 Citrate is transported from mitochondria by a
tricarboxylic acid transporter.
 In the cytoplasm, citrate is cleaved to oxaloacetate and
acetyl CoA. The enzyme is ATP citrate lyase.
The oxaloacetate can return to the mitochondria as malate
or pyruvate
Transportation of Acetyl co A as citrate…
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Enzymes and cofactors involved in the process of Fatty acid
synthesis

Two main enzymes-

Acetyl co A carboxylase
 Fatty acid Synthase

Both the enzymes are multienzyme complexes

Coenzymes and cofactors are-
 Biotin
 NADPH, Mn++ Mg++
Details of enzymes
1. Acetyl co A carboxylase
 Is catalyze the initial & controlling Step in Fatty Acid Synthesis
 Multienzyme complex containing:-
 Biotin
 Biotin Carboxylase
 Biotin carboxyl carrier protein
 Transcarboxylase
 A regulatory allosteric site
2. Fatty acid Synthase complex

 The Fatty Acid Synthase Complex is a polypeptide containing seven

enzyme activities and a terminal acyl carrier protein (ACP).
 In mammals, the fatty acid synthase complex is a dimer comprising
two identical monomers, each containing all seven enzyme activities of
fatty acid synthase on one polypeptide chain
Advantages of Multi-enzyme Complex
a. Intermediates of the reaction can easily interact with the active
sites of the enzymes.
b. One gene codes all the enzymes; so all the enzymes are in
equimolecular concentrations.
c. So the efficiency of the process is enhanced.

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Fatty acid synthase …
Fatty Acid Synthase prosthetic groups are:

a. The thiol (-SH)of the side-chain of a cysteine residue of keto

acyl synthase enzyme(also called condensing enzyme)

a. The thiol (-SH)of phosphopantetheine in the ACP, equivalent

in structure to part of coenzyme A.
The acyl carrier protein is a polypeptide chain having a
phospho-pantotheine group, to which the acyl groups are attached
in thioester linkage. So ACP acts like the CoA carrying fatty acyl
groups.
Steps of fatty acid synthesis
Formation of Malonyl co A

 The input to fatty acid synthesis is acetyl-

CoA, which is carboxylated to malonyl-
CoA.
 The reaction is catalyzed by Acetyl co A
carboxylase and committed step in the
pathway.
The acetyl CoA carboxylase is not a part of
the multienzyme complex. But it is the rate-
limiting enzyme.
Formation of Malonyl co A…
 ATP-dependent carboxylation provides energy input.

As for other carboxylation reactions, the enzyme prosthetic group is biotin. 

 The reaction takes place in two steps: carboxylation of biotin (involving

ATP) and transfer of the carboxyl to acetyl-CoA to form malonyl-CoA.

The CO2 is lost later during condensation with the growing fatty acid. The
spontaneous decarboxylation drives the condensation reaction.

The elongation of the fatty acid occurs by addition of 2 carbon atoms at a

time. But the 2-carbon units are added as 3-carbon, malonyl units. The whole
reaction sequence occurs while the intermediates are bound to ACP (acyl
carrier protein).
Formation of Malonyl co A…
 Remember Biotin is
linked to the enzyme by an
amide bond between the
terminal carboxyl of the
biotin side chain and the e-
amino group of a lysine
residue.
 Fatty acid synthase reactions
All the remaining steps are catalyzed by Fatty acid synthase
complex.
[1] A molecule of acetate is transferred from acetyl CoA to the –SH
group of the ACP. Domain: Acetyl CoA-ACP acetyltransacylase.
[2] Next, this two-carbon fragment is transferred to a temporary
holding site, the thiol group of a cysteine residue on the enzyme.
[3] The now-vacant ACP accepts a three-carbon malonate unit from
malonyl CoA. Domain: Malonyl CoA-ACP transacylase.
[4] The acetyl group on the cysteine residue condenses with the
malonyl group on ACP as the CO2 originally added by acetyl
CoA carboxylase is released. The result is a four-carbon unit
attached to the ACP domain. The loss of free energy from the
decarboxylation drives the reaction. Domain: 3-Ketoacyl-ACP
synthase.
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 Fatty acid synthase reactions…
 The next three reactions convert the 3-ketoacyl group to the
corresponding saturated acyl group by a pair of reductions
requiring NADPH and a dehydration step.
[5] The keto group is reduced to an alcohol. Domain: 3-Ketoacyl-
ACP reductase.
[6] A molecule of water is removed to introduce a double bond
between carbons 2 and 3 (the α- and β-carbons). Domain: 3-
Hydroxyacyl-ACP dehydratase.
[7] The double bond is reduced. Domain: Enoyl-ACP reductase.

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Fatty acid synthase reactions…
 The result of these seven steps is production of a four-carbon
compound (butyryl) whose three terminal carbons are fully
saturated, and which remains attached to the ACP.
 These seven steps are repeated, beginning with the transfer of the
butyryl chain from the ACP to the Cys residue [2*], the attachment
of a molecule of malonate to the ACP [3*], and the condensation of
the two molecules liberating CO2 [4*]. The carbonyl group at the β-
carbon (carbon 3—the third carbon from the sulfur) is then reduced
[5*], dehydrated [6*], and reduced [7*], generating hexanoyl-ACP.
This cycle of reactions is repeated five more times, each time
incorporating a two-carbon unit (derived from malonyl CoA) into
the growing fatty acid chain at the carboxyl end.
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Fatty acid synthase reactions…
 When the fatty acid reaches a length of 16 carbons, the
synthetic process is terminated with palmitoyl- S-ACP.
Note: Shorter-length fatty acids are important end
products in the lactating mammary gland.
 Palmitoyl thioesterase /de-acylase activity/ cleaves the
thioester bond, releasing a fully saturated molecule of
palmitate (16:0).
 Note: All the carbons in palmitic acid have passed
through malonyl CoA except the two donated by the
original acetyl CoA, which are found at the methyl-
group (ω) end of the fatty acid.
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Domains of fatty acid synthase complex

1.Acetyl CoA – ACP transacylase.

2. Malonyl CoA- ACP transacylase.
3. β- Ketoacyl – ACP synthase.
4. β- Ketoacyl- ACP reductase.
5. β –Hydroxyacyl – ACP dehydratase.
6. Enoyl – ACP reductase.
7. Palmitoyl thioesterase.
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 The overall reaction of palmitate synthesis

 8 acetyl CoA + 14 NADPH + 14 H + + 7 ATP

Palmitic acid + 8 CoA + 14 NADP + + 7 ADP + 7 Pi + 6 H 2O

 The cycle of reactions are repeated 7 times, each time

incorporating a two- carbon unit into a growing fatty acid chain.

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Fates of palmitate
1. Essterification:- Palmitate esterified with glycerol to
form acylglycerols or with cholesterol to cholesteryl
ester.
2. Chain elongation: Palmitate may be elongated to form
longer fatty acids
3. Desaturation i.e. synthesis of unsaturated fatty acids:
palmitate may undergo desaturation at C9-C10 to forrm
palmitoleic acid.
4. For synthesis of spingosine

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Regulation of fatty acid synthesis

When a cell has more energy, Glycerol-P

Glucose
the excess is generally converted Triacylglycerol
to Fatty Acids and stored as
Fatty acyl CoA
lipids such as triacylglycerol.

Pyruvate Malonyl CoA

Acetyl CoA

TCA cycle
Regulation of Acetyl-coA carboxylase
A. Short term regulation of acetyl CoA carboxylase
1. Allosteric regulation of acetyl CoA carboxylase:
 Citrate causes polymerization of the enzyme leading to its
activation , while malonyl CoA and long chain Fatty acyl CoA
such as palmitoyl CoA (the end product of the pathway)
depolymerize the enzyme leading to its inactivation.
2 .Reversible phosphorylation:
 In the presence of epinephrine, the enzyme is phosphorylated and
thereby inactivated and in the presence of insulin, the enzyme is
de-phosphorylated and thereby, activated.

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Allosteric & covalent (reversible phosphrylation) of ACC

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Regulation of fatty acids synthesis and long chain
fatty acids oxidation in liver

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B. Long term regulation of acetyl CoA carboxylase

In the long term by changes in gene expression governing rates

of synthesis of enzymes.

Prolonged consumption of high- carbohydrate or fat -free diet

↓
Increases acetyl CoA carboxylase synthesis and increases fatty
acid synthesis.
High- fat diet or fasting

↓
Decreases acetyl CoA carboxylase synthesis
Note: Nutritional state regulates lipogenesis

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Fatty acid elongation/ Micrsomal path way for fatty acid
synthesis
 Palmitate in animal cells is the precursor of other long-chained FAs.
 By further additions of acetyl groups, fatty acid chain length is elongated
through the action of FA elongation systems present in the smooth
endoplasmic reticulum.
This system is becomes active during myelination of neurons in order to
provide C22 and C24 fatty acids present in spingolipids.
Desaturation of fatty acids/Synthesis of unsaturated fatty
acids

 Animals can readily introduce one double bond to palmitate and stearate.
 Humans have carbon 9, 6, 5 and 4 desaturases, but lack the ability to
introduce double bonds from carbon 10 to the ω end of the chain.
 This is the basis for the nutritional essentiality of the polyunsaturated
linoleic and linolenic acids.
The desaturation of Fatty Acids…

 Palmitate and stearate serve as precursors of the two

most common monounsaturated fatty acids of animal
cells: palmitoleate (16:1Δ9), and Oleate (18:1Δ9).
 The double bond is introduced by fatty acyl-CoA
desaturase in the smooth endoplasmic reticulum.
Lipogenesis (synthesis of
triacylglycerols)
Lipogenesis (synthesis of triacylglycerols)

Fatty acids are stored in the adipose tissue in the form of neutral
TAGs which serve as the body’s major fuel storage reserve.
 TAGs provide concentrated stores of metabolic energy
because they are highly reduced and largely anhydrous.
 The yield from complete oxidation of fatty acids to CO2 and
H2O is 9 Kcal / g of fat, as compared to 4 Kcal / g of protein or
carbohydrate.

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A triacylglycerol with an unsaturated fatty acid on carbon 2.
Steps of synthesis of triacylglycerols(TAGs)
a. Synthesis of glycerol – 3- phosphate

b. Conversion of a free- fatty acid to its activated form which is

attached to coenzyme A ( fatty acyl CoA) , before it participates in
TAG synthesis, catalyzed by Fatty acyl CoA synthetase
(thiokinase).
c. Synthesis of a molecule of TAG from glycerol - 3- phosphate and
fatty acyl- CoA , by the sequential addition of two fatty acyl- CoA
molecules at carbons 1 and 2 catalyzed by acyl-transferase.

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Steps of synthesis of triacylglycerols(TAGs)…

 The subsequent removal of

the phosphate group at
carbon number 3 catalyzed
by phosphatase, followed by
the addition of the third fatty
acid catalyzed by acyl-
transferase.

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Pathways for production of glycerol phosphate in liver and adipose
tissue. Remember lack of glycerol kinase in adipose tissue.
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 Adipocytes can take up glucose only in the presence of the
hormone insulin (remember GLUT-4). Thus , when plasma glucose
is low and therefore, plasma insulin level is low , adipocytes have
only a limited ability to synthesize glycerol–3–phosphate and
cannot produce TAGs.

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 Different fates of TAG in the liver and adipose tissue
 In white adipose tissue, TAG is stored in a nearly
anhydrous form as fat droplets in the cytosol of the cells.
It serves as “depot fat,” ready for mobilization when the
body requires it for fuel.
 Little TAG is stored in the liver. Instead, most is
exported, packaged with other lipids and apoproteins to
form lipoprotein particles called very-lowdensity lipo
proteins (VLDL). Nascent VLDL are secreted directly
into the blood where they mature and function to deliver
the endogenously derived lipids to the peripheral tissues.
 Note: Recall that chylomicrons deliver primarily dietary
(exogenously derived) lipids.
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Metabolism of Adipose tissue

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 Lipolysis (degradation of TAG)

Mobilization of stored fats from the adipose tissue is

initiated by the hydrolytic release of fatty acids and
glycerol from their TAG form by hormone – senstive
lipase, that removes a fatty acid from either carbon 1 or
carbon 3 of the TAG.
Additional lipases specific for mono-acylglycerol or di-
acylglycerol remove the remaining fatty acids.
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Regulation of lipolysis
 Hormone-sensitive lipase is activated when phosphorylated by a
cAMP- dependent protein kinase.
 cAMP is produced in the adipocyte when one of several hormones
(primarily epinephrine) binds to receptors on the cell membrane.
 In the presence of high plasma levels of insulin and glucose,
hormone- sensitive lipase is de-phosphorylated and becomes inactive.
 When lipolysis is turned on , fatty acid synthesis is turned off because
acetyl CoA carboxylase the enzyme catalyzing the first reaction in the
de- novo synthesis of fatty acids is inhibited by hormone – directed
-phosphorylation.
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Regulation of lipolysis…
 Thus , if the cAMP-mediated cascade is activated for example
by epinephrine as in starvation or diabetes mellitus , fatty acid
synthesis is turned off, whereas, TAG degradation is turned on.
 In the presence of high plasma levels of insulin and glucose in
a well fed state, hormone- sensitive lipase is de-phosphorylated
and becomes inactive, so lipolysis is turned off and fatty acid
synthesis and lipogenesis is turned on.

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Control of
lipolysis

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 Fatty liver
Imbalance in the Rate of Tri- acylglycerol(TAG) formation & export
causes Fatty Liver. The balance between the factors causing fat deposition
in liver versus factors causing removal of fat from liver determines the
outcome.
 For a variety of reasons, lipid mainly as tri- acylglycerol (TAG) can
accumulate in the liver.
 Extensive accumulation is regarded as a pathologic condition.

 When accumulation of lipid in the liver becomes chronic, fibrotic

changes occur in the cells that progress to cirrhosis and impaired liver
function.

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 Fatty livers fall into two main categories
1. The first type is associated with raised levels of plasma free fatty
acids resulting from mobilization of fat from adipose tissue or from
the hydrolysis of lipoprotein triacylglycerol (TAG) by lipoprotein
lipase in extrahepatic tissues.
 The production of VLDL does not keep pace with the increasing
influx and esterification of free fatty acids, allowing TAG to
accumulate, causing a fatty liver.
 This occurs during starvation, Diabetes Mellitus and the feeding of
high-fat diets.

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 The ability to secrete VLDL may also be impaired ( eg, in starvation).
 In uncontrolled diabetes mellitus and ketosis, fatty infiltration is
sufficiently severe to cause visible pallor (fatty appearance) and
enlargement of the liver with possible liver dysfunction and ultimately
cirrhosis.
 High carbohydrate diet:- liver is first saturated with glycogen, then any
further amount of carbohydrate will be converted to triacyglycerols.
2. The second type of fatty liver is usually due to a metabolic
block in the production of plasma lipoproteins (VLDL),thus
allowing triacylglycerol to accumulate in the liver cells.

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Theoretically, the blocks may be due to
(1) Decrease in lipotropic factors
(2)a block in apolipoprotein synthesis,
(3) a block in the synthesis of the lipoprotein from lipid
and apolipoprotein (conjugation),
(4) A failure in provision of phospholipids that are
found in lipoproteins, or
(5) a failure in the secretory mechanism itself.

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Lipotropic factors
They are required for the normal mobilization of fat from liver.
Therefore, deficiency of these factors may result in fatty liver.
They can afford protection against the development of fatty liver.
1. Choline. Feeding of choline has been able to reverse fatty
changes in animals.
2. Lecithin and methionine. They help in synthesis of apoprotein
and choline formation. The deficiency of methyl groups for
carnitine synthesis may also hinder fatty acid oxidation.
3. Vitamin E and selenium give protection due to their anti-
oxidant effect.
4. Omega 3 fatty acids present in marine oils have a protective
effect against fatty liver.
5. Proteins of high biological values.

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Summary on causes of Fatty Liver
A .Causes of fat deposition in liver
1. Mobilization of NEFA from adipose tissue.
2. Excess synthesis of fatty acid from glucose.
B. Reduced removal of fat from liver
3. Toxic injury to liver. Secretion of VLDL needs synthesis
of apo B-100 and apo C.
4. Decreased oxidation of fat by hepatic cells.
An increase in factors (1) and (2) or a decrease in factors (3)
and (4) will cause excessive accumulation, leading to fatty
liver.

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Causes of fatty liver

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Ethanol also causes Fatty Liver
 Alcoholism leads to fat accumulation in the liver,
hyperlipidemia, and ultimately cirrhosis.
 The exact mechanism of action of ethanol in the long term is
still uncertain.
 Ethanol consumption over a long period leads to the
accumulation of fatty acids in the liver that are derived from
endogenous synthesis rather than from increased
mobilization from adipose tissue.
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 There is no impairment of hepatic synthesis of protein after ethanol
ingestion.
 Oxidation of ethanol by alcohol dehydrogenase leads to the
production of acetaldehyde and excess NADH. Acetaldehyde is
oxidized by aldehyde dehydrogenase, producing acetate.
 The NADH generated competes with reducing equivalents from other
substrates, including fatty acids, for the respiratory chain, inhibiting
their oxidation, and decreasing activity of the citric acid cycle.
 The net effect of inhibiting fatty acid oxidation is to cause increased
esterification of fatty acids in triacylglycerol, resulting in the fatty liver.

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 Other effects of ethanol may include increased lipogenesis and
cholesterol synthesis from acetyl-CoA, and lipid peroxidation.
 The increased [NADH]/[NAD+] ratio also causes increased
[lactate]/[pyruvate], resulting in hyperlacticacidemia, which
decreases excretion of uric acid, aggravating gout.
 Some metabolism of ethanol to acetaldehyde takes place via
a cytochrome P450-dependent microsomal ethanol oxidizing
system (MEOS) involving NADPH and O2.

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 This system increases in activity in chronic alcoholism and may
account for the increased metabolic clearance in this condition.

 Ethanol will also inhibit the metabolism of some drugs, eg,

barbiturates, by competing for cytochrome P450-dependent
enzymes.

 In some Asian populations and Native Americans, alcohol

consumption results in increased adverse reactions to acetaldehyde
owing to a genetic defect of mitochondrial aldehyde
dehydrogenase.

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