CHOLESTEROL SYNTHESIS-

STEPS AND REGULATION

Biochemistry for medics

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CHOLESTEROL INTRODUCTION

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Cholesterol is the major sterol in the animal tissues.
Cholesterol is present in tissues and in plasma either
as free cholesterol or as a storage form, combined

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with a long-chain fatty acid as cholesteryl ester. In
plasma, both forms are transported in lipoproteins
Plasma low-density lipoprotein (LDL) is the vehicle of
uptake of cholesterol and cholesteryl ester into many
tissues.
Free cholesterol is removed from tissues by plasma
high-density lipoprotein (HDL) and transported to
the liver, where it is eliminated from the body either
unchanged or after conversion to bile acids in the
process known as reverse cholesterol transport . 2
STRUCTURE OF CHOLESTEROL

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The structure of cholesterol consists of four fused
rings (The rings in steroids are denoted by the letters

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A, B, C, and D.), with the carbons numbered in the
sequence, and an eight numbered, and branched
hydrocarbon chain attached to the D ring.
Cholesterol contains two angular methyl groups: the
C-19 methyl group is attached to C-10, and the C-18
methyl group is attached to C-13.
The C-18 and C-19 methyl groups of cholesterol lie
above the plane containing the four rings.

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STRUCTURE OF CHOLESTEROL (CONTD.)

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Steroids with 8 to 10 carbon atoms in the side chain
and an alcohol hydroxyl group at C-3 are classified as
sterols. Much of the plasma cholesterol is in the
esterified form (with a fatty acid attached at carbon 3), 4
which makes the structure even more hydrophobic .
FUNCTIONS OF CHOLESTEROL

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Cholesterol is the most abundant sterol in humans and
performs a number of essential functions. For example-

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It is a major constituent of the plasma membrane and of
plasma lipoproteins.
It is a precursor of bile salts,
It is a precursor of steroid hormones that include
adrenocortical hormones, sex hormones, placental
hormones etc
Also a precursor of vitamin D, cardiac glycosides,
Sitosterol of the plant kingdom, and some alkaloids.
It is required for the nerve transmission. Cholesterol is
widely distributed in all cells of the body but particularly 5
abundant in nervous tissue.
SOURCES OF CHOLESTEROL

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Cholesterol is derived from
diet

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de novo synthesis and
from the hydrolysis of cholesteryl esters.
A little more than half the cholesterol of the body
arises by synthesis (about 700 mg/d), and the
remainder is provided by the average diet.
The liver and intestine account for approximately
10% each of total synthesis in humans.
Virtually all tissues containing nucleated cells are
capable of cholesterol synthesis, which occurs in the
endoplasmic reticulum and the cytosol. 6
STEPS OF SYNTHESIS OF CHOLESTEROL

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Acetyl co A acts as a precursor of cholesterol. All
27 carbon atoms of cholesterol are derived from

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acetyl CoA in a three-stage synthetic process Stage
one is the synthesis of Isopentenyl
pyrophosphate, an activated isoprene unit that is
the key building block of cholesterol.
Stage two is the condensation of six molecules of
Isopentenyl pyrophosphate to form Squalene. In
stage three, Squalene cyclizes in an astounding
reaction and the tetracyclic product is
subsequently converted into cholesterol. 7
STAGE 1 OF CHOLESTEROL SYNTHESIS

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The first stage in the synthesis of cholesterol is the
formation of Isopentenyl pyrophosphate from acetyl

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CoA.
This set of reactions, which takes place in the cytosol,
starts with the formation of 3-hydroxy-3-
methylglutaryl CoA (HMG CoA) from acetyl CoA.
Initially, two molecules of acetyl-CoA condense to
form Acetoacetyl-CoA catalyzed by cytosolic thiolase.
Acetoacetyl-CoA condenses with a further molecule
of acetyl-CoA catalyzed by HMG-CoA synthase to form
HMG-CoA, that is reduced to mevalonate by NADPH
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catalyzed by HMG-CoA reductase.
The synthesis of
mevalonate is the
committed step in
cholesterol formation.

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The enzyme catalyzing
this irreversible step,

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3-hydroxy-3-
methylglutaryl CoA
reductase (HMG-CoA
reductase), is an
important control site in
cholesterol biosynthesis,
and is the site of action of
the most effective class of
cholesterol-lowering
drugs, the HMG-CoA
reductase inhibitors
(statins). 9
STAGE 1 OF CHOLESTEROL SYNTHESIS
(CONTD.)

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Mevalonate is converted into 3-isopentenyl
pyrophosphate in three consecutive reactions requiring
ATP.

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Decarboxylation yields Isopentenyl pyrophosphate,
an activated isoprene unit that is a key building block
for many important biomolecules.

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STAGE -2 OF CHOLESTEROL SYNTHESIS

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Synthesis of Squalene
Squalene (C30) is synthesized from six molecules of

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Isopentenyl Pyrophosphate (C5) and the reaction
sequence is-
C5 C10 C15 C30
This stage in the synthesis of cholesterol starts with
the isomerization of isopentenyl pyrophosphate to
dimethylallyl pyrophosphate.
Isopentenyl diphosphate is isomerized by a shift of
the double bond to form dimethylallyl diphosphate,
then condensed with another molecule of isopentenyl
diphosphate to form the ten-carbon intermediate
geranyl diphosphate. 11
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STAGE -2 OF CHOLESTEROL SYNTHESIS
(CONTD.)

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A further condensation with isopentenyl
diphosphate forms farnesyl diphosphate. Two

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molecules of farnesyl diphosphate condense at the
diphosphate end to form squalene. Initially,
inorganic pyrophosphate is eliminated, forming
presqualene diphosphate, which is then reduced
by NADPH with elimination of a further inorganic
pyrophosphate molecule.

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STAGE-3- FORMATION OF CHOLESTEROL
FROM SQUALENE

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Squalene can fold into a
structure that closely
resembles the steroid

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nucleus .
Before ring closure occurs,
squalene is converted to
squalene 2,3-epoxide by a
mixed-function oxidase in the
endoplasmic reticulum,
squalene epoxidase.
The methyl group on C14 is
transferred to C13 and that on
C8 to C14 as cyclization occurs,
catalyzed by oxidosqualene:
lanosterol cyclase.
The newly formed cyclized
structure is Lanosterol

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STAGE-3- FORMATION OF CHOLESTEROL
FROM SQUALENE (CONTD.)
The formation of cholesterol from

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lanosterol takes place in the
membranes of the endoplasmic

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reticulum and involves changes in the
steroid nucleus and side chain .
The methyl groups on C14 and C4 are
removed to form 14-desmethyl
lanosterol and then zymosterol. The
double bond at C8–C9 is subsequently
moved to C5–C6 in two steps, forming
desmosterol.
Finally, the double bond of the side
15
chain is reduced, producing cholesterol.
REGULATION OF CHOLESTEROL BIOSYNTHESIS

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Regulation of cholesterol synthesis is exerted near the
beginning of the pathway, at the HMG-CoA reductase

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step.
Following mechanisms are involved at the regulatory
step-
Competitive inhibition
Feed back inhibition
Covalent modification(Role of hormones)
Sterol mediated regulation of transcription

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REGULATION OF CHOLESTEROL BIOSYNTHESIS

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Competitive inhibition
Statins (Lovastatin,

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Mevastatin, Atorva Statin
etc.) are the reversible
competitive inhibitors of
HMG Co A reductase. They
are used to decrease
plasma cholesterol levels in
patients of
hypercholesterolemia.

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REGULATION OF CHOLESTEROL BIOSYNTHESIS

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Feed back inhibition
HMG Co A reductase is

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inhibited by Mevalonate and
Cholesterol.
Mevalonate is the immediate
product of HMG Co A reductase
catalyzed reaction whereas
Cholesterol is the ultimate
product of the reaction
pathway.

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REGULATION OF CHOLESTEROL BIOSYNTHESIS

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Covalent modification (Role of hormones)
Phosphorylation decreases the activity of the

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reductase.
Glucagon favors formation of the inactive
(phosphorylated form) form, hence decreases the
rate of cholesterol synthesis
In contrast , insulin favors formation of the
active(dephosphorylated )form of HMG Co A
reductase and results in an increase in the rate of
cholesterol synthesis
Cholesterol synthesis ceases when the ATP level is 19
low
REGULATION OF CHOLESTEROL BIOSYNTHESIS

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Sterol mediated regulation of transcription
The synthesis of cholesterol is also regulated by the

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amount of cholesterol taken up by the cells during
lipoprotein metabolism.
Chylomicron remnants internalized by liver cells, and
low density lipoproteins internalized by liver cells
and peripheral tissues provide cholesterol which
causes a decrease in the transcription of HMG CoA
reductase gene, leading to a decrease in cholesterol
synthesis.
The rate of synthesis of reductase mRNA is controlled
20
by the sterol regulatory element binding protein
(SREBP).
REGULATION OF CHOLESTEROL BIOSYNTHESIS

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Sterol mediated regulation of transcription
This transcription factor binds to a short DNA sequence

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called the sterol regulatory element (SRE) on the 5 side of
the reductase gene.
In its inactive state, the SREBP is anchored to the
endoplasmic reticulum or nuclear membrane.
When cholesterol levels fall, the protein is released
The released protein migrates to the nucleus and binds
the SRE of the HMG-CoA reductase gene, to enhance
transcription.
When cholesterol levels rise, the proteolytic release of 21
the SREBP is blocked, and the SREBP in the nucleus is
rapidly degraded
TRANSPORT OF CHOLESTEROL

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Cholesterol is transported in plasma in lipoproteins, and
in humans the highest proportion is found in LDL.

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Cholesteryl ester in the diet is hydrolyzed to cholesterol,
which is then absorbed by the intestine together with
dietary unesterified cholesterol and other lipids.
With cholesterol synthesized in the intestines, it is then
incorporated into chylomicrons.
Ninety-five percent of the chylomicron cholesterol is
delivered to the liver in chylomicron remnants,
Most of the cholesterol secreted by the liver in VLDL is
retained during the formation of IDL and ultimately LDL,
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which is taken up by the LDL receptor in liver and extra
hepatic tissues.
UPTAKE OF LDL CHOLESTEROL

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The LDLs (containing cholesteryl esters) are taken up
by cells by a process known as receptor-mediated

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endocytosis.
The LDL receptor mediates this endocytosis and is
important to cholesterol metabolism.
After LDL binding to the LDL receptor, the ligand-
receptor complexes cluster on the plasma membrane
in coated pits, which then invaginate forming coated
vesicles.
These coated vesicles are internalized and clathrin,
the protein composing the lattice in membrane
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coated pits, is removed.
UPTAKE OF LDL CHOLESTEROL

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These vesicles are now called endosomes and these
endosomes fuse with the lysosome.

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The LDL receptor–containing membrane buds off and
is recycled to the plasma membrane.
Fusion of the lysosome and endosome releases
lysosomal proteases that degrade the apoproteins
into amino acids.
Lysosomal enzymes also hydrolyze the cholesteryl
esters to free cholesterol and fatty acids.
The free cholesterol is released into the cell’s
cytoplasm, and this free cholesterol is then available 24
to be used by the cell.
UPTAKE OF LDL CHOLESTEROL

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Excess cholesterol is reesterified by acyl-CoA:
cholesterol acyltransferase (ACAT), which uses fatty

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acyl-CoA as the source of activated fatty acid. Free
cholesterol affects cholesterol metabolism by
inhibiting cholesterol biosynthesis. Cholesterol
inhibits the enzyme hydroxy- methylglutaryl-CoA
reductase (HMG-CoA reductase), which catalyzes an
early rate-limiting step in cholesterol biosynthesis.

In addition, free cholesterol inhibits the synthesis of

the LDL receptor, thus limiting the amount of LDLs
25
that are taken up by the cell.
UPTAKE OF LDL CHOLESTEROL

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Receptor mediated endocytosis of LDL 26
VARIATION OF SERUM CHOLESTEROL
LEVELS

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The normal serum cholesterol concentration ranges
between 150- 220 mg/dl

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High cholesterol concentration is found in-
Diabetes mellitus
Nephrotic syndrome
Obstructive jaundice
Familial hypercholesterolemia
Biliary cirrhosis
Hypothyroidism

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VARIATION OF SERUM CHOLESTEROL
LEVELS

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Hypocholesterolemia- Low serum cholesterol
concentration is observed in-

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Hyperthyroidism
Malnutrition
Malabsorption
Anemia
Physiologically lower levels are found in children
Persons on cholesterol lowering drugs

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HYPERCHOLESTEROLEMIA AND THE
CONSEQUENCES

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Atherosclerosis is characterized
by the deposition of cholesterol

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and cholesteryl ester from the
plasma lipoproteins into the
artery wall.
Diseases in which prolonged
elevated levels of VLDL, IDL,
chylomicron remnants, or LDL
occur in the blood are often
accompanied by premature or
more severe atherosclerosis.
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HYPOLIPIDEMIC DRUGS

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Statins - The statins act as competitive inhibitors of the
enzyme HMG-CoA reductase.

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Fibrates such as Clofibrate and gemfibrozil act mainly
to lower plasma triacylglycerols by decreasing the
secretion of triacylglycerol and cholesterol-containing
VLDL by the liver.
Ezetimibe- ezetimibe, reduces blood cholesterol levels
by inhibiting the absorption of cholesterol by the
intestine
Bile Acid Sequestrants (Resins)-Bile acid sequestrants
bind bile acids in the intestine and promote their
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excretion in the stool. To maintain the bile acid pool size,
the liver diverts cholesterol to bile acid synthesis
HYPOLIPIDEMIC DRUGS

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Bile acid sequestrants (contd.)-The decreased hepatic
intracellular cholesterol content results in up regulation

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of the LDL receptor and enhanced LDL clearance from the
plasma. Bile acid sequestrants, including
Cholestyramine, Colestipol, and colesevelam. Omega
3 Fatty Acids (Fish Oils)-The most widely used n-3
PUFAs for the treatment of hyperlipidemia are the two
active molecules in fish oil: Eicosapentaenoic acid (EPA)
and Docosahexaenoic acid (DHA).
Niacin inhibits the release of free fatty acids from adipose
tissue which leads to a decrease of free fatty acids
entering the liver and decreased VLDL synthesis in the 31
liver.
ROLE OF DIET IN REGULATING CHOLESTEROL
LEVELS

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Polyunsaturated fatty acids have a cholesterol
lowering effect

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There is the up-regulation of LDL receptors by poly-
and monounsaturated as compared with saturated
fatty acids, causing an increase in the catabolic rate of
LDL, the main atherogenic lipoprotein.
In addition, saturated fatty acids cause the formation
of smaller VLDL particles that contain relatively more
cholesterol, and they are utilized by extra hepatic
tissues at a slower rate than are larger particles and
thus may be regarded as atherogenic.
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LIFESTYLE AND THE SERUM CHOLESTEROL
LEVELS

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Additional factors considered to play a part in coronary
heart disease include high blood pressure, smoking,

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male gender, obesity (particularly abdominal obesity)
and lack of exercise
Premenopausal women appear to be protected against
many of these deleterious factors, and this is thought to
be related to the beneficial effects of estrogen.
There is an association between moderate alcohol
consumption and a lower incidence of coronary heart
disease. This may be due to elevation of HDL
concentrations resulting from increased synthesis of
apo A-I 33

Regular exercise lowers plasma LDL but raises HDL.