BCH 2212

METABOLISM OF CARBOHYDRATES & LIPIDS

(FOR ANATOMY/PHYSIOLOGY STUDENTS)
METABOLISM OF LIPIDS
(Lipolysis and the Oxidation of Fatty Acids)
Lipid Digestion
• Utilization of dietary lipids requires that they first be absorbed
through the intestine.
• As these molecules are oils they would be essentially insoluble in the
aqueous intestinal environment.
• Solubilization (emulsification) of dietary lipid is accomplished initially
via the agitation action as food passes through the stomach
• emulsification process continues within the intestine via bile salts that
are synthesized in the liver and secreted from the gallbladder.
Lipid Digestion
• Dietary lipids, in the form of triglycerides (triacylglycerides),
phospholipids, and cholesterol, are digested by various lipases.
• The lipases found in the gastrointestinal tract include:
• lingual lipase (secreted by acinar cells of von Ebner glands of the
tongue),
• gastric lipase (secreted by Chief cells of the stomach),
• pancreatic lipase (PNLIP gene), and
• pancreatic lipase-related protein 2 (PNLIPRP2 gene).
• These enzymes generate free fatty acids and a mixture of mono- and
diglycerides from dietary triglycerides.
Lipid Digestion
• The acid lipases are distinct from pancreatic lipases in that they do not require a
lipid-bile acid interface for activity nor do they require the presence of the protein
colipase.
• Pancreatic lipases, on the other hand, only function in the neutral pH environment
generated in the small intestine by the secretion of pancreatic bicarbonate
(HCO3–).
• Also, pancreatic lipases require the presence of colipase and a lipid-bile acid
interface for their activity.
• The role of colipase in pancreatic lipase function is to anchor the lipase to the
surface of an emulsified lipid droplet and to prevent it from being removed by bile
salts.
• Pancreatic lipase degrades triglycerides at the sn-1 and sn-3 positions sequentially
to generate 1,2-diacylglycerides (DAG) and 2-monoacylglycerides (MAG).
• Phospholipids are degraded at the sn-2 position by pancreatic phospholipase
A2 releasing a free fatty acid and the lysophospholipid.
Lipid Digestion
• The triglycerides are then solubilized in lipoprotein
complexes (complexes of lipid and protein) called chylomicrons.
• Triglycerides synthesized in the liver are packaged into VLDLs and
released into the blood directly.
• The triglyceride components of VLDLs and chylomicrons are
hydrolyzed to free fatty acids and glycerol in the capillaries of tissues
such as adipose tissue, heart and skeletal muscle by the actions of
lipoprotein lipase (LPL) and in the liver through the actions of hepatic
triglyceride lipase [HTGL) gene].
• The free fatty acids are then absorbed by the cells and the glycerol is
returned via the blood to the liver (principal site) and kidneys.
• The glycerol can then be converted to the glycolytic intermediate
dihydroxyacetone phosphate DHAP or phosphorylated by glycerol
kinase to glycerol-3-phosphate for reuse in triglyceride synthesis.
Mobilization of Fat Stores
• The primary sources of fatty acids for oxidation are dietary and
mobilization from cellular stores.
• Fatty acids from the diet are absorbed from the gut, packaged
into lipoprotein particles called chylomicrons within intestinal enterocytes
and then delivered to cells of the body via transport in the blood.
• Fatty acids are stored in the form of triglycerides (triacylglycerides: TAGs
or TGs) within all cell but predominantly within adipose tissue.
• In response to energy demands, the fatty acids of stored triglycerides can
be mobilized for use by peripheral tissues. The release of metabolic
energy, in the form of fatty acids, is controlled by a complex series of
interrelated cascades that result in the activation of triglyceride hydrolysis.
• The primary intracellular lipases are adipose triglyceride lipase (ATGL, also
called desnutrin), hormone-sensitive lipase (HSL), and lipase A (LIPA; also
called lysosomal acid lipase, LAL).
Monoglyceride lipase: MGL
• MGL is considered to be the rate-limiting enzyme for the breakdown
of monoglycerides that are the result of both extracellular and
intracellular lipolysis pathways.
• The extracellular generation of monoglycerides is the result of the
action of endothelial cell lipoprotein lipase (LPL) on lipoprotein
particle-associated triglycerides.
• Intracellular hydrolysis of triglycerides by ATGL and HSL, as well as
intracellular phospholipid hydrolysis by phospholipase C (PLC) and
membrane-associated diglyceride lipase α and β results in the
generation of MGL substrates.
Cellular Uptake of Fatty Acids
• When fatty acids are released from adipose tissue stores they enter
the circulation as free fatty acids (FFAs) and are bound to albumin for
transport to peripheral tissues.
• When the fatty acid–albumin complexes interact with cell surfaces the
dissociation of the fatty acid from albumin represents the first step of
the cellular uptake process.
• Uptake of fatty acids by cells involves membrane proteins with high
affinity for fatty acids. There are several members of the fatty acid
receptor family including fatty acid translocase (FAT/CD36), plasma
membrane-associated fatty acid-binding protein (FABPpm), and fatty
acid transport proteins (FATPs).
• The FATPs facilitate the uptake of very long-chain (VLCFA) and long-
chain fatty acids (LCFA).
Mitochondrial (beta) β-Oxidation Reactions - Fatty Acid
Activation
• Oxidation of fatty acids occurs in the mitochondria and the
peroxisomes.
• Fatty acids of between 4–8 and between 6–16 carbon atoms in length,
referred to as short- and medium-chain fatty acids (SCFAs and MCFAs,
respectively), are oxidized exclusively in the mitochondria.
• Long-chain fatty acids (LCFAs: 10–18 carbons long) are oxidized in both
the mitochondria and the peroxisomes with the peroxisomes
exhibiting preference for 14-carbon and longer LCFAs.
• Very-long-chain fatty acids (VLCFAs: C18–C26; often designated as any
fatty acid of 22 carbons and longer) are preferentially oxidized in
the peroxisomes. However, given that peroxisomes are unable to
completely oxidize fatty acids, the chain shortened free fatty acids or
the carnitine esters are released and taken up by the mitochondria for
complete oxidation to CO2 and H2O.
Mitochondrial (beta) β-Oxidation Reactions - Fatty Acid
Activation
• Very long-chain and long-chain fatty acids must be activated in the
cytoplasm before being oxidized in the mitochondria or the peroxisomes.
• Medium-chain and short-chain fatty acids are activated within the matrix of
the mitochondria.
• Fatty acid activation is catalyzed by fatty acyl-CoA synthetases (also called
acyl-CoA ligases or thiokinases).
• The net result of this activation process is the consumption of two molar
equivalents of ATP.
Fatty Acid Transport into Mitochondria
• Short-chain and medium-chain fatty acids require no specific transport
mechanism to enter the mitochondria for oxidation and, as indicated
earlier, are activated by CoA attachment within the mitochondrial matrix.
• Also, because dietary short-chain and medium-chain fatty acids directly
enter the portal circulation (they are not packaged into chylomicrons)
they are rapidly oxidized within the liver.
• Long-chain fatty acids, present in the triglycerides of chylomicrons, or
VLDL, or as free fatty acids released from adipose tissue require a
specific mitochondrial transport mechanism to be oxidized.
• The transport of very long-chain and long-chain fatty acyl-CoA into the
mitochondria is accomplished via an acyl-carnitine intermediate, which
itself is generated by the action of carnitine palmitoyltransferase 1 (CPT-
1 or CPT-I) an enzyme that resides in the outer mitochondrial
membrane.
 Fatty Acid Transport into Mitochondria
• Once a fatty acyl-carnitine is generated at the outer mitochondrial
membrane it is transported into the mitochondria through the action of
carnitine acyl-carnitine translocase, CACT.
• The carnitine acyl-carnitine translocase is located in the inner
mitochondrial membrane where it facilitates acyl-carnitine transport
across the outer and inner mitochondrial membranes in exchange for
free carnitine.
• Following CACT-mediated transfer of the CPT-1-generated fatty acyl-
carnitines across the inner mitochondrial membrane, the fatty acyl-
carnitine molecules are acted on by the inner mitochondrial membrane-
associated carnitine palmitoyltransferase 2 (CPT-2 or CPT-II) regenerating
the fatty acyl-CoA molecules.
Transport of fatty acids from the cytoplasm to the inner mitochondrial space for oxidation. Following
activation of a long-chain fatty acid to a fatty-CoA, the CoA is exchanged for carnitine by CPT-1. The fatty-
carnitine is then transported to the inside of the mitochondrion through the action of CACT where a reversal
exchange takes place through the action of CPT-2. Once inside the mitochondrion the fatty-CoA is a substrate
for the β-oxidation machinery.
Processes of Mitochondrial Fatty Acid β-Oxidation
• The process of mitochondrial fatty acid oxidation is termed β-
oxidation since it occurs through the sequential removal of 2-carbon
units by oxidation at the β-carbon position of the fatty acyl-CoA
molecule.
• The oxidation of fatty acids and lipids in the peroxisomes also occurs
via a process of β-oxidation but the enzymes are distinct from those
used within the mitochondria.
• Each round of mitochondrial fatty acid β-oxidation involves four steps
that, in order, are oxidation, hydration, oxidation, and cleavage.
• The first oxidation step in mitochondrial β-oxidation involves a family
of FAD-dependent acyl-CoA dehydrogenases that act on saturated
fatty acids. Each of these dehydrogenases has a range of substrate
specificities determined by the length of the fatty acid. Short-chain
acyl-CoA dehydrogenase (SCAD, also called butyryl-CoA
dehydrogenase) prefers fats of 4–6 carbons in length.
Processes of Mitochondrial Fatty Acid β-Oxidation
• Medium-chain acyl-CoA dehydrogenase (MCAD; also known as
ACAD1) prefers fats of 4–16 carbons in length with maximal activity
for C10 acyl-CoAs.
• Long-chain acyl-CoA dehydrogenase (LCAD; also known as ACAD4)
prefers fats of 10-18 carbons in length with maximal activity for C12
acyl-CoAs.
• Very long-chain acyl-CoA dehydrogenase (VLCAD; also known as
ACAD6) prefers fats of 16-24 carbons and is inactive on any fatty acid
less than 12 carbons.
• Unlike the localization of SCAD, MCAD, and LCAD to the mitochondrial
matrix, VLCAD is an inner mitochodrial membrane-localized enzyme.
Processes of Mitochondrial Fatty Acid β-Oxidation
• Following the FAD-dependent acyl-CoA dehydrogenase step, the next
three steps in mitochondrial β-oxidation involve a hydration step,
another oxidation step, and finally a hydrolytic reaction that requires
CoA and releases acetyl-CoA and an acyl-CoA two carbon atoms
shorter than the initial substrate.
• In the oxidation of these substrates, until they become medium-chain
length, the water addition is catalyzed by an enoyl-CoA hydratase
activity, the second oxidation step is catalyzed by an NAD-dependent
long-chain hydroxyacyl-CoA dehydrogenase activity (3-hydroxyacyl-
CoA dehydrogenase activity), and finally the cleavage into an acyl-CoA
and an acetyl-CoA is catalyzed by a thiolase activity.
• For the oxidation of very long-chain and long-chain saturated fatty
acids these three activities are encoded in a multifunctional enzyme
called the mitochondrial trifunctional protein, MTP.
Processes of Mitochondrial Fatty Acid β-Oxidation
• Medium-chain and short-chain saturatred fatty acids are oxidized by
soluble mitochondrial matrix enzymes. Following the action of MCAD
and SCAD the resulting enoyl-CoAs are substrates for the enoyl-CoA
hydratase.
• The next oxidation step is catalyzed by hydroxyacyl-CoA
dehydrogenase. This dehydrogenase exhibits highest specificity for
medium-chain saturated fatty acids.
• Mutations in the HADH gene are associated with familial
hyperinsulinemic hypoglycemia.
• The thiolase reaction is catalyzed by the ACAA2 encoded enzyme
which is also referred to as medium-chain 3-ketoacyl-CoA thiolase,
MKCAT.
Processes of Mitochondrial Fatty Acid β-Oxidation
• Each round of β-oxidation produces one mole of FADH2, one mole of NADH,
and one mole of acetyl-CoA.
• The acetyl-CoA, the end product of each round of β-oxidation, then enters
the TCA cycle, where it is further oxidized to CO2 with the concomitant
generation of three moles of NADH, one mole of FADH2 and one mole of ATP.
• The NADH and FADH2 generated during the fat oxidation and acetyl-CoA
oxidation in the TCA cycle then can enter the respiratory pathway for the
production of ATP via oxidative phosphorylation.
• The oxidation of fatty acids yields significantly more energy per carbon atom
than does the oxidation of carbohydrates.
• The net result of the oxidation of one mole of oleic acid (an 18-carbon fatty
acid) will be 146 moles of ATP (2 mole equivalents are used during the
activation of the fatty acid), as compared with 114 moles from an equivalent
number of glucose carbon atoms.
Pathway of mitochondrial β-oxidation of very long-chain and long-chain fatty acids: Mitochondrial oxidation of long-chain fatty acids occurs
in a series of four steps once a fatty acid is activated by one of several fatty acyl-CoA synthetases. The first reaction of mitochondrial fatty acid β-
oxidation is catalyzed by one of several FAD-dependent acyl-CoA dehydrogenases. The remaining three reactions are carried out by an inner
mitochondrial membrane associated multifunctional enzyme complex termed the mitochondrial trifunctional protein, MTP.
Peroxisomal (beta) β-Oxidation Reactions
• In addition to mitochondrial oxidation of fatty acids, the peroxisomes also play an important
role in overall fatty acid metabolism.
• Very-long-chain fatty acids (VLCFAs: C18–C26) are preferentially oxidized in the peroxisomes
with cerotic acid (a 26:0 fatty acid) being solely oxidized in this organelle.
• Peroxisomal fatty acid oxidation does not proceed to CO2 and H2O as in the mitochondria.
The chain-shortened free fatty acids, or carnitine esters, are released from the peroxisomes
and taken up by the mitochondria where their complete oxidation takes place.
• The peroxisomes also metabolize di– and trihydroxycholestanoic acids (bile
acid intermediates); long-chain dicarboxylic acids that are produced by ω-oxidation of long-
chain monocarboxylic acids; pristanic acid via the α-oxidation pathway; certain
polyunsaturated fatty acids (PUFAs) such as tetracosahexaenoic acid (24:6), which by β-
oxidation yields the important PUFA docosahexaenoic acid (DHA); and
certain prostaglandins and leukotrienes.
• The clinical significance of the activity of the acyl-CoA oxidases of peroxisomal β-oxidation is
related to tissue specific oxidation processes. In the pancreatic β-cell there is little, if any,
catalase expressed so that peroxisomal oxidation of VLCFAs results in an increased release of
ROS that can damage the β-cell contributing to the progressive insulin deficiency seen in
obesity.
Microsomal (omega) ω-Oxidation Reactions
• The microsomal (endoplasmic reticulum, ER) pathway of fatty acid ω-
oxidation represents a minor pathway of overall fatty acid oxidation.
However, in certain pathophysiological states, such as diabetes,
chronic alcohol consumption, and starvation, the ω-oxidation
pathway may provide an effective means for the elimination of toxic
levels of free fatty acids.
• The pathway refers to the fact that fatty acids first undergo a
hydroxylation step at the terminal (omega, ω) carbon.
 Phytanic Acid α-Oxidation Reactions
• Phytanic acid (3,7,11,15-tetramethylhexadecanoic acid) is a fatty acid present in the tissues of
ruminants, certain fishes, and in dairy products and is, therefore, an important dietary component
of fatty acid intake.
• Phytanic acid is a 3-methyl branched fatty acid, it cannot act as a substrate for any of the first
enzymes of the mitochondrial β-oxidation pathway (the acyl-CoA dehydrogenases).
• Because the first step in phytanic acid oxidation involves an α-oxidation step, the process is
termed α-oxidation.
• Phytanic acid is first converted to its CoA-ester and then phytanoyl-CoA serves as a substrate in an
α-oxidation process. The α-oxidation reaction (as well as the remainder of the reactions of
phytanic acid oxidation) occurs within the peroxisomes and requires a specific α-hydroxylase
(specifically phytanoyl-CoA hydroxylase, PhyH), which adds a hydroxyl group to the α-carbon of
phytanic acid. PhyH is also called phytanoyl-CoA 2-hydroxylase or phytanoyl-CoA dioxygenase.
• Following two more reactions, the resulting product of phytanic acid metabolism is the 19-carbon
fatty acid, pristanic acid (2,6,10,14-tetramethylpentadecanoic acid).
• Pristanic acid then serves as a substrate for the peroxisomal process of β-oxidation.
• After three steps of peroxisomal β-oxidation, the medium-chain fatty acid product (4,8-
dimethylnonanoic acid) is esterified to carnitine and transferred to the mitochondria for further
oxidation in the mitochondrial β-oxidation pathway.
Regulation of Fatty Acid Metabolism (Oxidation)
• Fat metabolism can also be regulated by malonyl-CoA-mediated
inhibition of CPT-1.
• Such regulation serves to prevent de novo synthesized fatty acids from
entering the mitochondria and being oxidized.
• The predominant source of the malonyl-CoA that inhibits CPT-1 is
from the ACC2 enzyme.
Clinical Significance of Fatty Acids
• The majority of clinical problems related to fatty acid metabolism are associated with
processes of oxidation. These disorders fall into four main groups:
1. Deficiencies in Carnitine: Deficiencies in carnitine lead to an inability to transport fatty
acids into the mitochondria for oxidation. This can occur in newborns and particularly in
pre-term infants.
• Symptoms can range from mild occasional muscle cramping to severe weakness or even
death.
• Treatment is by oral carnitine administration.
2. Carnitine palmitoyltransferase deficiencies: Deficiencies in CPT-1 are relatively rare and
affect primarily the liver and lead to reduced fatty acid oxidation and ketogenesis.
• The most common symptom associated with CPT-1 deficiency is hypoketotic
hypoglycemia. There is also an elevation in blood levels of carnitine.
• Symptoms of this form appear within hours to 4 days after birth and include respiratory
failure, hepatomegaly, seizures, hypoglycemia, and cardiomegaly. The cardiomegaly will
lead to fatal arrhythmias.
• Carnitine acyltransferases may also be inhibited by sulfonylurea drugs such as
tolbutamide and glyburide.
Clinical Significance of Fatty Acids
• 3. Deficiencies in Acyl-CoA Dehydrogenases: A group of inherited diseases
that impair β-oxidation result from deficiencies in acyl-CoA dehydrogenases.
The enzymes affected may belong to one of three categories:
• very long-chain acyl-CoA dehydrogenase (VLCAD)
• medium-chain acyl-CoA dehydrogenase (MCAD)
• short-chain acyl-CoA dehydrogenase (SCAD)
• MCAD deficiency is the most common form of acyl-CoA dehydrogenase
deficiency.
• Symptoms include vomiting, lethargy and frequently coma. Excessive urinary
excretion of medium-chain dicarboxylic acids as well as their glycine and
carnitine esters is diagnostic of this condition.
• In the case of this enzyme deficiency taking care to avoid prolonged fasting is
sufficient to prevent clinical problems.
Clinical Significance of Fatty Acids
4. Refsum Disease: Refsum disease is a rare inherited disorder in which patients harbor
a defect in the peroxisomal α-oxidizing enzyme, phytanoyl-CoA hydroxylase (PhyH).
Although mutations in PhyH are the primary cause of Refsum disease, the syndrome
can also result from defects in the peroxisomal protein (PEX7) responsible for the
import of PhyH into the peroxisome.
• Patients accumulate large quantities of phytanic acid in their tissues and serum.
• This leads to severe symptoms, including cerebellar ataxia, retinitis pigmentosa, nerve
deafness and peripheral neuropathy.
• As expected, the restriction of dairy products and ruminant meat from the diet can
ameliorate the symptoms of this disease.
• Phytanic acid accumulation is also seen when there are inherited defects in
peroxisome function leading to Zellweger syndrome, neonatal
adrenoleukodystrophy and infantile Refsum disease. In addition, rhizomelic
chondrodysplasia punctata, type 1 (RCDP1) results in phytanic acid accumulation.
• Refsum disease due to deficiency in PhyH is properly referred to as classical Refsum
disease to distinguish it from infantile Refsum due to peroxisome dysfunction.
METABOLISM OF LIPIDS

(FATTY ACID BIOSYNTHESIS)

28
 INTRODUCTION
• Fatty acid synthesis occurs mainly in the liver in humans,
although the process also occurs in adipose tissue.
• When an excess of dietary carbohydrate is consumed,
glucose is converted to acetyl CoA, which provides the
2-carbon units that condense in a series of reactions on
the fatty acid synthase complex, producing palmitate.
• Palmitate is then converted to other fatty acids.

29
Reactions of Fatty Acid Synthesis
• The synthesis of malonyl-CoA is the first committed
step of fatty acid synthesis and the enzyme that
catalyzes this reaction, acetyl-CoA carboxylase (ACC),
is the major site of regulation of fatty acid synthesis.
Like other enzymes that transfer CO2 to substrates,
ACC requires a biotin as a co-factor. Acetyl-CoA
carboxylase is called an ABC enzyme due to the
requirements for ATP, Biotin, and CO2 for the
reaction.
• Acetyl-CoA is generated in the mitochondria primarily
from two sources, the pyruvate dehydrogenase (PDH)
reaction and fatty acid oxidation.

30
Conversion of Glucose to Cytosolic Acetyl
CoA
The fatty acid synthase complex is
located in the cytosol, and,
therefore, it uses cytosolic acetyl
CoA.

Fig. 33.6. Conversion of glucose to cytosolic acetyl CoA.

OAA = oxaloacetate. 31
 Sources of NADPH for fatty acid synthesis

Fig. 33.9. Sources of NADPH for fatty acid synthesis. NADPH is produced by the pentose
phosphate pathway and by malic enzyme (decarboxylating or NADP-dependent malate
dehydrogenase) reaction. 32
Acetyl-CoA carboxylase (ACC) Reaction
33
• The synthesis of fatty acids from acetyl-CoA and malonyl-CoA is
carried out by fatty acid synthase, FAS.
• All of the reactions of fatty acid synthesis are carried out by the
multiple enzymatic activities of FAS.
• Fat synthesis involves four primary enzymatic activities. These are
(in order of reaction):
• β-ketoacyl-ACP synthase,
• β-ketoacyl-ACP reductase,
• 3-hydroxyacyl-ACP dehydratase and,
• enoyl-CoA reductase.
• The two reduction reactions require NADPH oxidation to
NADP+.
34
1 and 2 = malonyl/acetyl-CoA ACP transacetylase (also called malonyl/acetyltransferase,
MAT. 3 = Acyl carrier protein portion of FAS (ACP-SH) (also catalyzed by malonyl/acetyl-CoA
ACP transacetylase). Cysteine sulfhydryl (Cys-SH). 4 = β-ketoacyl-ACP synthase. 5 = β-
ketoacyl-ACP reductase. 6 = 3-hydroxyacyl-ACP dehydratase . 7 = enoyl-CoA reductase. 8 =
CYS-SH.
35
• FAS is initially activated by the incorporation of the acetyl group from acetyl-
CoA. The acetyl group is initially attached to the sulfhydryl of the 4'-
phosphopantothenate of the acyl carrier protein portion of FAS (ACP-SH).
• This is catalyzed by malonyl/acetyl-CoA ACP transacetylase (1 and 2; also
called malonyl/acetyltransferase, MAT).
• This activating acetyl group represents the omega (ω) end of the newly
synthesized fatty acid.
• Following transfer of the activating acetyl group to a cysteine sulfhydryl in
the β-ketoacyl-ACP synthase portion of FAS, the three carbons from a
malonyl-CoA are attached to ACP-SH (3) also catalyzed by malonyl/acetyl-
CoA ACP transacetylase.
• The acetyl group attacks the methylene group of the malonyl attached to
ACP-SH catalyzed β-ketoacyl-ACP synthase (4) which also liberates the
CO2 that was added to acetyl-CoA by ACC.
36
• The resulting 3-ketoacyl group then undergoes a series of three reactions
catalyzed by the β-ketoacyl-ACP reductase (5), 3-hydroxyacyl-ACP
dehydratase (6), and enoyl-CoA reductase (7) activities of FAS that reduce,
dehydrate, and reduce the substrate.
• This results in a saturated four carbon (butyryl) group attached to the ACP-
SH.
• This butyryl group is then transferred to the CYS-SH (8) as for the case of the
activating acetyl group. At this point another malonyl group is attached to
the ACP-SH (3b) and the process begins again.
• Reactions 4 through 8 are repeated another six times, each beginning with a
new malonyl group being added.
• At the completion of synthesis the saturated 16 carbon fatty acid, palmitic
acid, is released via the action of the thioesterase activity of FAS (palmitoyl
ACP thioesterase) located in the C-terminal end of the enzyme.
37
 Elongation of Fatty Acids

• After synthesis on the fatty acid synthase complex, palmitate is activated,

forming palmityl CoA. Palmityl CoA and other activated long-chain fatty acids
can be elongated, two carbons at a time, by a series of reactions that occur
in the endoplasmic reticulum (Fig. 33.17). Malonyl CoA serves as the donor
of the 2-carbon units, and NADPH provides the reducing equivalents.
• The series of elongation reactions resemble those of fatty acid synthesis
except that the fatty acyl chain is attached to coenzyme A rather than to the
phosphopantetheinyl residue of an ACP.
• The major elongation reaction that occurs in the body involves the
conversion of palmityl CoA (C16) to stearyl CoA (C18).
• During fatty acid synthesis, malonyl CoA levels, being high, inhibits
carnitine:palmitoyltransferase (CPTI) (also called carnitine:acyltransferase I),
which is involved in the transport of long-chain fatty acids into mitochondria
for β-oxidation. This mechanism prevents newly synthesized fatty acids from
undergoing immediate oxidation. 38
Fig. 33.17. Elongation of long-chain fatty acids in the endoplasmic reticulum.
39
 Desaturation of Fatty Acids

• Desaturation of fatty acids involves a process that requires

molecular oxygen (O2), NADH, and cytochrome b5. The
reaction, which occurs in the endoplasmic reticulum, results
in the oxidation of both the fatty acid and NADH (Fig.
33.18).
• The most common desaturation reactions involve the
placement of a double bond between carbons 9 and 10 in
the conversion of palmitic acid to palmitoleic acid (16:1,∆ 9)
and the conversion of stearic acid to oleic acid (18:1,∆9).
• Other positions that can be desaturated in humans include
carbons 4, 5, and 6. 40
Fig. 33.18. Desaturation of fatty acids. The process occurs in the endoplasmic reticulum and uses molecular oxygen. Both the fatty acid and
NADH are oxidized. Human desaturases cannot introduce double bonds between carbon 9 and the methyl end. Therefore, m is equal to or
less than 7.
Plants are able to introduce double bonds into fatty acids in the region between C10 and the ω-end and therefore can synthesize 3 and 6
polyunsaturated fatty acids.
Fish oils also contain 3 and 6 fatty acids, particularly eicosapentaenoic acid (EPA; ω3, 20:5, ∆ 5, 8, 11, 14, 17) and docosahexaenoic acid
(DHA; ω3, 22:6, ∆ 4,7,10,13,16,19). The fish obtain these fatty acids by eating phytoplankton (plants that float in water). 41
• Arachidonic acid is listed in some textbooks as an essential fatty acid. Although it
is an ω6 fatty acid, it is not essential in the diet if linoleic acid is present because
arachidonic acid can be synthesized from dietary linoleic acid (see Fig. 33.19).
• The essential fatty acid linoleic acid is required in the diet for at least three
reasons:
• (a) It serves as a precursor of arachidonic acid from which eicosanoids are
produced.
• (b) It covalently binds another fatty acid attached to cerebrosides in the skin,
forming an unusual lipid (acylglucosylceramide) that helps to make the skin
impermeable to water. This function of linoleic acid may help to explain the red,
scaly dermatitis and other skin problems associated with a dietary deficiency of
essential fatty acids.
• (c) It is the precursor of C22:6ω3, an important neuronal fatty acid.
• The other essential fatty acid, α-linolenic acid (18:3, ∆9, 12, 15), also forms
eicosanoids.
42
Fig. 33.19. Conversion of linoleic acid to arachidonic acid. Dietary linoleic acid (as linoleoyl
CoA) is desaturated at carbon 6, elongated by 2 carbons, and then desaturated at carbon 5
to produce arachidonyl CoA. 43
SYNTHESIS OF TRIACYLGLYCEROLS AND VLDL PARTICLES

Fig. 33.20. Synthesis of triacylglycerol in liver and adipose tissue. Glycerol 3-phosphate is produced from
glucose in both tissues. It is also produced from glycerol in liver, but not in adipose tissue, which lacks
glycerol kinase. The steps from glycerol 3-phosphate are the same in the two tissues.
44
FA = fatty acyl group.
Regulation of Fatty Acid Metabolism
• In order to understand how the synthesis and degradation of fats needs
to be exquisitely regulated, one must consider the energy requirements
of the organism as a whole.
• The blood is the carrier of triglycerides in the form of VLDLs and
chylomicrons, fatty acids bound to albumin, amino acids, lactate, ketone
bodies and glucose.
• The pancreas is the primary organ involved in sensing the organism's
dietary and energetic states by monitoring glucose concentrations in the
blood.
• Low blood glucose stimulates the secretion of glucagon, whereas,
elevated blood glucose calls for the secretion of insulin.
Regulation of Fatty Acid Metabolism
• The metabolism of fat is regulated by two distinct mechanisms.
• One is short-term regulation, which can come about through events such as
substrate availability, allosteric effectors and/or enzyme modification.
• The other mechanism, long-term regulation, is achieved by alteration of the
rate of enzyme synthesis and turn-over.
• ACC is the rate-limiting (committed) step in fatty acid synthesis. There are
two major isoforms of ACC in mammalian tissues. These are identified as
ACC1 and ACC2.
• Both isoforms of ACC are allosterically activated by citrate and inhibited by
palmitoyl-CoA and other short- and long-chain fatty acyl-CoAs. Citrate
triggers the polymerization of ACC1 (and ACC2) which leads to significant
increases in its activity.
• Glutamate and other dicarboxylic acids can also allosterically activate both
ACC isoforms.
• ACC activity can also be affected by phosphorylation.
Regulation of Fatty Acid Metabolism
• Phosphorylation of ACC1 by AMPK leads to inhibition of the enzyme. AMPK-
mediated phosphorylation appears to be the most significant for the
regulation of ACC1 activity.
• Glucagon-stimulated increases in cAMP, and subsequently to increased PKA
activity, also lead to phosphorylation of ACC.
• ACC2 is a better substrate for PKA than is ACC1.
• The activating effects of insulin on ACC is known to dephosphorylate the ACC1
that are AMPK targets in the heart enzyme. This insulin-mediated effect has
not been observed in hepatocytes or adipose tissues cells. At least a portion
of the activating effects of insulin are related to changes in cAMP levels.
• Activation of α1-adrenergic receptors in liver and skeletal muscle cells inhibits
ACC activity as a result of phosphorylation by a kinase.
• Insulin, a product of the well-fed state, stimulates ACC1 and FAS synthesis,
whereas starvation leads to a decrease in the synthesis of these enzymes.
Regulation of Fatty Acid Metabolism
• Adipose tissue contains hormone-sensitive lipase (HSL), which is activated by
PKA-dependent phosphorylation; this activation increases the release of fatty
acids into the blood. This in turn leads to the increased oxidation of fatty acids
in other tissues such as muscle and liver. In the liver, the net result (due to
increased acetyl-CoA levels) is the production of ketone bodies.
• The activity of HSL is also affected via phosphorylation by AMPK. In this case
the phosphorylation inhibits the enzyme.
• It has been proposed that inhibition of HSL by AMPK mediated-
phosphorylation is a mechanism to ensure that the rate of fatty acid release
does not exceed the rate at which they are utilized either by export or
oxidation.
• Insulin has the opposite effect to glucagon and epinephrine: it increases the
synthesis of triglycerides (and glycogen). One of the many effects of insulin is
to lower cAMP levels, which leads to increased dephosphorylation through
the enhanced activity of protein phosphatases such as PP-1.
 Regulation of Fatty Acid Metabolism

Acetyl CoA carboxylase is regulated allosterically, both positively and negatively, by phosphorylation (circled P) and
dephosphorylation, and by diet-induced induction (circled ↑). It is active in the dephosphorylated state when citrate
causes it to polymerize. Dephosphorylation is catalyzed by an insulin-stimulated phosphatase. Low energy levels, via
activation of an AMP-dependent protein kinase, cause the enzyme to be phosphorylated and inactivated. The ultimate
product of fatty acid synthesis, palmitate, is converted to its CoA derivative palmityl CoA, which inhibits the enzyme. A
high-calorie diet increases the rate of transcription of the gene for acetyl CoA carboxylase, whereas a low-calorie diet
reduces transcription of this gene. 49
CHOLESTEROL SYNTHESIS
• Cholesterol is an alicyclic compound whose basic structure includes
the perhydrocyclopentanophenanthrene nucleus containing four
fused rings (Figure 34.1).
• In its “free” form, the cholesterol molecule contains 27 carbon atoms,
a simple hydroxyl group at C3, a double bond between C5 and C6, an
eight-membered hydrocarbon chain attached to carbon 17 in the D
ring, a methyl group (carbon 19) attached to carbon 10, and a second
methyl group (carbon 18) attached to carbon 13 (Figure 34.2).

50
51
• Approximately one third of plasma cholesterol exists in the free (or
unesterified) form. The remaining two thirds exists as cholesterol esters in
which a long-chain fatty acid (usually linoleic acid) is attached by ester
linkage to the hydroxyl group at C-3 of the A ring.
• All of the 27 carbons are derived from one precursor, acetyl CoA. Acetyl
CoA can be obtained from several sources, including the beta oxidation of
fatty acids, the oxidation of ketogenic amino acids, such as leucine and
lysine, and the pyruvate dehydrogenase reaction. Carbons 1, 2, 5, 7, 9, 13,
15, 18, 19, 20, 22, 24, 26, and 27 of cholesterol are derived from the
methyl group of acetyl CoA and the remaining 12 carbons of cholesterol
from the carboxylate atom of acetyl CoA.
• The synthesis of cholesterol requires significant reducing power, which is
supplied in the form of NADPH. The latter is provided by glucose-6-
phosphate dehydrogenase and 6-phosphogluconate dehydrogenase of the
hexose monophosphate shunt pathway.
• Cholesterol synthesis occurs in the cytosol, requiring hydrolysis of high-
energy thioester bonds of acetyl CoA and phosphoanhydride bonds of ATP.
• Its synthesis occurs in four stages.
52
A. Stage 1: Synthesis of Mevalonate from Acetyl CoA
• The first stage of cholesterol
synthesis leads to the production of
the intermediate mevalonate (Fig.
34.3). The synthesis of mevalonate
is the committed, rate-limiting step
in cholesterol formation.
• The HMG-CoA synthase in this
reaction is present in the cytosol
and is distinct from the
mitochondrial HMG-CoA synthase
that catalyses HMG-CoA synthesis
involved in ketone body production.
• The committed step and major
point of regulation of cholesterol
synthesis in stage 1 involves
reduction of HMG-CoA to
mevalonate, a reaction catalyzed by
HMG-CoA reductase, an enzyme
embedded in the membrane of the
endoplasmic reticulum.
NB: β-hydroxy- β -methylglutaryl-CoA
(HMG-CoA)
53
B. Stage 2: Conversion of Mevalonate to Two Activated Isoprenes

• In the second stage of

cholesterol synthesis,
three phosphate
groups are transferred
from three molecules
of ATP to mevalonate
(Fig. 34.5).
• The purpose of these
phosphate transfers is
to activate both carbon
5 and the hydroxyl
group on carbon 3 for
further reactions in
which these groups will
leave the molecule.
54
C. Stage 3: Condensation of Six Activated 5-Carbon Isoprenes to Form the 30-Carbon
Squalene • The next stage in the biosynthesis of
cholesterol involves the head-to-tail
condensation of
isopentenylpyrophosphate and
dimethylallyl pyrophosphate. In this
reaction, one pyrophosphate group is
displaced, and a 10-carbon chain,
known as geranyl pyrophosphate, is
generated (Fig. 34.6). (The “head”
refers to the end to which
pyrophosphate is joined.)
• Geranyl pyrophosphate then
undergoes another head-to-tail
condensation with isopentenyl
pyrophosphate, resulting in the
formation of the 15-carbon
intermediate, farnesyl pyrophosphate.
• After this, two molecules of farnesyl
pyrophosphate undergo a head-to-
head fusion, and both pyrophosphate
groups are removed to form squalene
(30 carbons). 55
D. Stage 4: Conversion of Squalene to the Four-Ring Steroid Nucleus
• The enzyme squalene
monooxygenase adds a single oxygen
atom from O2 to the end of the
squalene molecule, forming an
epoxide.
• NADPH then reduces the other
oxygen atom of O2 to H2O.
• The unsaturated carbons of the
squalene 2, 3- epoxide are aligned in
a way that allows conversion of the
linear squalene epoxide into a cyclic
structure.
• The cyclization leads to the formation
of lanosterol, a sterol with the four-
ring structure characteristic of the
steroid nucleus.
• A series of complex reactions,
containing many steps, leads to the
formation of cholesterol (Fig. 34.7).
56
ANATOMIC AND BIOCHEMICAL ASPECTS OF ATHEROSCLEROSIS
• The initial step in the development of an
atherosclerotic lesion within the wall of
an artery is the formation of a fatty
streak (accumulation of lipid-ladened
macrophages or foam cells in the
subintimal space).
• These fatty streaks are visible as a yellow-
white linear streak that bulges slightly
into the lumen of the vessel.
• These streaks are initiated when one or
more known “vascular risk factors for
atherosclerosis,” all of which have the
potential to injure the vascular
endothelial cells, reach a critical
threshold at the site of future lesions.
• Examples of such risk factors include
elevated intra-arterial pressure (arterial
hypertension), elevated circulating levels
of various lipids such as LDL, chylomicron
remnants, and VLDL remnants, or low
levels of circulating HDL, cigarette
smoking, chronic elevations in blood
glucose levels, high circulating levels of
the vasoconstricting octapeptide
angiotensin II, and others. 57
• The resulting insult to endothelial cells may trigger these cells to secrete adhesion
molecules that bind to circulating monocytes and markedly slow their rate of movement
past the endothelium. When sufficiently slowed, these monocytic cells accumulate and
have access to the physical spaces that exist between endothelial cells. This
accumulation of monocytic cells resembles the classical inflammatory response to injury.
These changes have led to the suggestion that atherosclerosis is, in fact, an inflammatory
disorder and, therefore, is one that might be prevented or attenuated through the use of
anti-inflammatory agents such as acetylsalicylic acid (e.g., aspirin) and HMG CoA
reductase inhibitors (statins), which have been shown to suppress the inflammatory
cascade as well as to inhibit the action of HMG CoA reductase.
• The monocytic cells are transformed into macrophages that migrate through the spaces
between endothelial cells. They enter the subintimal space under the influence of
chemoattractant cytokines (e.g., chemokine macrophage chemoattractant protein I)
secreted by vascular cells in response to exposure to oxidatively modified fatty acids
within the lipoproteins. The macrophages can replicate and exhibit augmented
expression of receptors that recognize oxidatively modified lipoproteins. Unlike the
classic LDL receptors on liver and many nonhepatic cells, these macrophage-bound
receptors are highcapacity, low-specificity receptors (scavenger receptors). They bind to
and internalize oxidatively modified fatty acids within LDLs to become subintimal foam
cells.
• As these foam cells accumulate, they deform the overlying endothelium, causing
microscopic separations between endothelial cells, exposing these foam cells and
underlying extracellular matrix to the blood. These exposed areas serve as sites for
58
platelet adhesion and aggregation.
• Activated platelets secrete cytokines that perpetuate this process and increase the
potential for thrombus (clot) formation locally. As the evolving plaque matures, a fibrous
cap forms over its expanding “roof,” which now bulges into the vascular lumen, thereby
partially occluding it. Vascular smooth muscle cells now migrate from the tunica media to
the subintimal space and secrete additional plaque matrix material. The smooth muscle
cells also secrete metalloproteinases that thin the fibrous cap near its “elbow” at the
periphery of the plaque. This thinning progresses until the fibrous cap ruptures, allowing
the plaque contents to physically contact the procoagulant elements present within the
circulation.
• This leads to acute thrombus formation. If this thrombus completely occludes the
remaining lumen of the vessel, an infarction of tissues distal to the occlusion (i.e., an
acute myocardial infarction) may occur (Fig. 34.22). Most plaques that rupture also
contain focal areas of calcification, which appears to result from the induction of the
same cluster of genes as those that promote the formation of bone. The inducers for this
process include oxidized sterols as well as transforming growth factor beta (TGF-) derived
from certain vascular cells.
• Finally, high intraluminal shear forces develop in these thinning or eroded areas of the
plaque’s fibrous cap, inducing macrophages to secrete additional metalloproteinases that
further degrade the arterial-fibrous cap matrix. This contributes further to plaque rupture
and thrombus formation (see Fig. 34.22). The consequence is a macrovascular ischemic
event such as an acute myocardial infarction (AMI) or an acute cerebrovascular accident
(CVA).
59
60
READING ASSIGNMENT -Synthesis of Omega-3 and -6 Fatty
Acids
• Most of the omega-6 PUFA consumed in the diet is from vegetable
oils such as soybean oil, corn oil, safflower oil, and borage oil, and
consists of the 18-carbon (18:2) PUFA linoleic acid. Linoleic acid,
which is an essential fatty acid, is converted to arachidonic acid.
• The synthesis of omega-3 fatty acids, EPA and DHA, utilizes the other
essential fatty acid, linolenic acid, which is also an 18-carbon (18:3)
PUFA. The overall pathway for EPA and DHA synthesis involves
three endoplasmic reticulum (ER) fatty acid elongation enzyme
systems, three desaturation reactions involving two different
desaturases, and a peroxisomal β-oxidation step as the final step in
DHA synthesis as outlined in the Figure below.
Pathway for ALA conversion to EPA
and DHA: Alpha(α)-linolenic acid
(ALA) is converted to the omega-3
polyunsaturated fatty acids (PUFA),
eicosapentaenoic acid (EPA),
docosapentaenoic acid (DPA), and
docosahexaenoic acid (DHA), via the
actions of a series of microsomal
(endoplasmic reticulum, ER) enzymes.
The final step in DHA synthesis
involves peroxisomal β-oxidation to
remove the 2-carbon acetyl-CoA from
the 24:6 fatty acid called
tetracosahexaenoic acid (also called
nisinic acid).
 Clinical Significance of Omega-3, and -6 PUFAs
• The most important omega-6 PUFA is arachidonic acid. When cells are stimulated by a variety of external stimuli,
arachidonic acid is released from cell membranes through the action of phospholipase A2 (PLA2). The released
arachidonate then serves as the precursor for the synthesis of the biologically active eicosanoids, the prostaglandins
(PGs), thromboxanes (TXs), and leukotrienes. (LTs) The arachidonate-derived eicosanoids function in diverse biological
phenomena such as platelet and leukocyte activation, signaling of pain, induction of bronchoconstriction, and regulation of
gastric secretions. These activities are targets of numerous pharmacological agents such as the non-steroidal anti-
inflammatory drugs (NSAIDs), COX-2 inhibitors, and leukotriene antagonists.
• Dietary omega-3 PUFAs compete with the inflammatory, pyretic (fever), and pain promoting properties imparted by omega-
6 PUFAs because they displace arachidonic acid from phospholipids present in cell membranes. In addition, omega-3
PUFAs compete with the enzymes that convert arachidonic acid into the biological eicosanoids (PGs, TXs, and LTs). The
net effect of increasing dietary consumption of omega-3 PUFAs, relative to omega-6 PUFAs, is to decrease the potential
for monocytes, neutrophils, and eosinophils (i.e. leukocytes) to synthesize potent mediators of inflammation and to reduce
the ability of platelets to release TXA2, a potent stimulator of the coagulation process.
• The omega-3 fatty acids DHA and EPA have also been shown to be important for normal brain development and function.
Several studies have demonstrated that DHA is essential for proper development of the prenatal and postnatal central
nervous system. The benefits of EPA appear to be in its effects on behavior and mood. In clinical studies with DHA and
EPA there has been good data demonstrating benefit in treating attention deficit hyperactivity disorder (ADHD), autism,
dyspraxia (motor skills disorder), dyslexia, and aggression. In patients with affective disorders consumption of DHA and
EPA has confirmed benefits in major depressive disorder and bipolar disorder. In addition, some studies have
demonstrated promising results in treatment of schizophrenia with some minor benefits in patients with borderline
personality disorder. Although originally purported to have positive effects in children with autism or autism spectrum
disorder, recent clinical studies have shown that omega-3 PUFA supplementation does not have any positive effect on
these disorders. Of significance to these effects of EPA and DHA on cognition, mood and behavior is the fact that
administration of omega-3 fatty acid containing phospholipids (such as those present in Krill oils) are significantly better
than omega-3 containing triacylglycerides such as those that predominate in fish oils.