Download Biochem 34 [5-29

Cholesterol absorption, synthesis, metabolism, and fate
Biochem Ch. 34
I.
II.
III.
Intestinal absorption of Cholesterol

ABC protein family is ATP-binding cassete that uses ATP hydrolysis to transport unwanted/excessive cholesterol and plant
sterols from enterocyte back into gut lumen (ABCG5 and ABCG8)

ABCA1 is required for reverse cholesterol transport and biogenesis of HDL

cholesterol is eliminated from the body in feces as unreabsorbed sterols and bile acids

increase in ABC protein expression = increase in elimination of sterols through feces

phytosterolemia is a disease with defect in function of ABCG5 or ABCG8 in enterocytes

accumulates cholesterol in cells then leads to elevated cholesterol and phytosterol in blood

ezetimibe is compound that lowers serum cholesterol levels

targets NPC1L1 which is believed to transport cholesterol into cells
Cholesterol synthesis

Facts:

27 carbon atoms in its free form

1/3 of plasma cholesterol is in its free unesterified form; other 2/3 are in cholesterol esters with a long FA chain
attached

structure of cholesterol suggest its synthesis involves significant reducing power

all 27 carbons derived from acetyl-CoA

reducing power of cholesterol synthesis is NADPH

Stage 1: synthesis of mevalonate from acetyl-CoA

rate-limiting step in cholesterol formation

2 acetyl-CoA condense  acetoacetyl-CoA

this condenses with another acetyl-CoA HMG-CoA

through HMG-CoA synthase

HMG-CoA is reduced by HMG-CoA reductase to make mevalonate

HMG-CoA reductase is embedded in the ER and uses 2 NADPH molecules; its regulation is controlled
through:
1. Transcriptional control

SREBPs are integral proteins of the ER and bound to SCAP when cholesterol
levels are high

when cholesterol levels drop, the sterols cleave off of SCAP and SREBP:SCAP
complex are transported to Golgi which will then release the DNA binding domain
of SREBP

this domain will travel to nucleus and bind to the sterol-regulatory element (SRE)
upstream of the gene and increase txn
2. Proteolytic degradation of HMG-CoA reductase

increased levels of cholesterol and bile salts cause reductase to be more
susceptible to proteolysis
3. Regulation by covalent modification (phosphorylation)

elevated glucagon levels, increased intracellular sterols, and increased thyroid
hormones increase phosphorylation (inactivates HMG-CoA)

elevated insulin and increased glucocorticoids cause dephysphorylation
(activates)

phosphorylation by AMP-activated protein kinase; cholesterol synthesis
increases when ATP levels are high

Stage 2: conversion of mevalonate to 2 activated isoprenes

3 phosphate groups are transferred from 3 ATP molecules to mevalonate (prepping for further rxns)

phosphate group and carboxyl group are removed to form 2 isoprenes (isopentenyl pyrophosphate &
dimethylallyl pyrophosphate)

Stage 3: condensation of 6 activated 5-carbon isoprenes to form the 30-carbon squalene

isopentyl pyrophosphate + dimethylallyl pyrophosphate  Geranyl pyrophosphate

Geranyl pyrophosphate + isopentyl pyrophosphate  farnesyl pyrophosphate

farnesyl pyrophosphate + farnesyl pyrophosphate  squalene

Stage 4: conversion of squalene to the 4-ring steroid nucleus

enzyme squalene monoxygenase adds oxygen to end of squalene making an epoxide

unsaturated carbons allow cyclization and formation of lanosterol and then complex rxns lead to final formation
of cholesterol
Several fates of cholesterol

most cholesterol is formed in the liver and will be released from hepatocytes as either cholesterol esters, biliary cholesterol, or
bile acids

cholesterol ester production in liver is catalyzed by acyl-CoA-cholesterol acyl transferase (ACAT)

cholesterol esters are more hydrophobic than free cholesterol; liver packages esterified cholesterol into hollow core of
liporproteins (VLDL)

VLDL is secreted from hepatocyte into blood and transports cholesterol esters to tissues that need cholesterol (used for
membrane, steroid hormone, vit. D synth.)
IV.
V.
VI.

extra cholesterol esters stored in liver for later use

hepatic cholesterol pool serves as source for synthesis of bile acids and salts
Synthesis of Bile Salts

Conversion of cholesterol to cholic acid & chenocholic acid

7-a-hydroxylase adds an a-hydroxyl group to carbon 7 on cholesterol

activity of the hydroxylase is decreased by increase in bile salt conc.

double bond in B ring is reduced and additional hydroxylation occurs

2 sets of compounds produced:

cholic acid series: a-hydroxyl groups at positions 3, 7, and 12

chenodeoxychoilic acid series: a-hydroxyl groups 3 and 7

pKa of bile acids is ~6 (same as intestinal lumen); 50% are protonated and other half form bile salts

Conjugation of bile salts

carboxyl group from bile salt activated by rxn that requires ATP and CoA

it will then react with gycine or taurine to form conjugated bile salts

glycine bile salts: glycocholic and glycochenodeoxycholic acid (pKa~4)

taurine bile salts: taurocholic and taurochenodeoxycholic acid (pKa~2)
Fate of bile salts

bile salts are produced in the liver and secreted into the bile; stored in the gallbladder and released into the intestine during a
meal where they will aid digestion of lipids

intestinal bacteria deconjugate and dehydroxylate bile salts (remove glycine and taurine residues and hydroxyl group at
position 7

bile salts lacking hydroxyl group at position 7 = secondary bile salts

deconjugated and dehydroxylated bile salts are less soluble (less readily absorbed) from lumen

>95% of bile salts are resorbed in ileum and return to liver where they are recycled and secreted as bile

steroid nucleus can’t be degraded in body, so excretion as bile salts is way of removal of steroid nucleus &
cholesterol
Transport of cholesterol by the blood lipoproteins

Blood lipoproteins

cholesterol must be transported through these b/c they are hydrophobic and insoluble in the blood

the core is made up of cholesterol esters and triacylglycerols and surrounded by a shell of phospholipids
embedded with a variety of apoproteins and free cholesterol

through these carriers, lipids leave their tissue of origin, enter bloodstream, and are transported to tissues where
their components are used

apoproteins add hydrophilicity & structural stability and also

activate enzymes required for lipoprotein metabolism

are ligands on surface of lipoprotein that target certain tissues

Chylomicrons

largest of lipoproteins and least dense b/c rich triacylglycerol content

synthesized from dietary lipids (exogenous lipoprotein pathway)

major apolipoproteins: apoB48, apoCII, and apoE

apoCII activates LPL

LPL, when activated goes into capillaries of adipose tissue, cardiac muscle, skeletal muscle, and acinar cells of
mammary tissues to hydrolyze chylomicrons and release FA into target cells

muscle cells oxidize FA as fuel

adipocytes and mammary cells store as triacylglycerols (fat)

mammary tissue may use for milk production

chylomicrons now depleted of their core triacylglycerols and have lost their apoCII are picked up by the liver
(through apoE still embedded on chylomicron remnants) through receptor-med. endocytosis

VLDL

excess intake of carbohydrates leads to formation of triacylglycerols which will from nascent VLDL (w/ free and
esterified cholesterol, phospholipids, and apoB100

nascent VLDL are then secreted from the liver (endogenous pathway of lipoprotein metabolism) into
bloodstream where they’ll accept apoCII and apoE from circulating HDL particles (now, it is a mature VLDL)

mature LDL are transported to tissues; apoCII activates LDL to hydrolyze triacylglycerol and release FA &
glycerol; used as fuel in muscle cells, resynthesize triacyclycerol in fat cells, and milk production in lactating
breast

50% of VLDL remnants are taken up from blood by liver through binding of VLDL apoE

IDL & LDL

leftover 50% of VLDL not taken up by liver will have their additional triacylglycerols removed to from IDL

hepatic triglyceride lipase will remove more triacylglycerol to from LDL (rich in cholesterol and cholesterol
esters)

~60% of LDL is transported to liver and endocytosed through binding of apo 100

other 40% go to extrahepatic tissues where they are made into steroid hormones for membrane synthesis and
vitamin D

excess LDL can oversaturate receptors on both hepatic and non-hepatic tissues

excess LDL is more readily uptaken by macrophages near endothelial cells of arteries  inflammatory response
 can initiate cascade of artherclerosis

HDL

Synthesis (3 possible mechanisms)

synthesis of nascent HDL by liver with a very hollow core
budding of apoproteins from chylomimcrons and VLDDL particles as they are being ingested by LDL
free apoAI shed from other circulating lipoproteins acquires cholesterol and phospholipids from others
and cell membranes forming a nascent HDL particle

Maturation of nascent HDL

accumulates phospholipids and cholesterol from cells lining blood vessels

central hollow core fills taking on a globular shape to form the mature HDL (filling with lipids doesn't
require enzyme activity)

Reverse cholesterol transport

removal of cholesterol from cholesterol-laden cells and return to liver

in vascular cells, reduces cholesterol levels and reduces chance of foam cells (macrophages that engulf
oxidized LDL-cholesterol and represent early stage of development of artherosclerotic plaque)

ABCA1 + ATP hydrolysis moves cholesterol from inner to outer leaflet of cell membrane, where HDL can
accept it

to trap inside HDL, HDL acquires LCAT (lecithin-cholesterol acylransferase) by transferring a FA from
lecithin of phospholipid shell to the cholesterol, forming a cholesterol ester, which then migrates into the
core of HDL and can no longer leave

elevated levels of lipoprotein-associated cholesterol in blood (particularly LDL but also triacylglycerolrich lipoproteins) are associated with formation of atheromatous plaques in vessel wall leading to diffuse
atherosclerotic vascular disease

high levels of HDL in the blood are therefore vaculoprotective b/c the increase reverse cholesterol
transport

Fate of HDL

mature HDL is cleared from blood primarily through uptake by scavenger receptor SR-B1

SR-B1 receptor is on many cells, and when HDL is bound, its cholesterol and cholesterol esters enter into
the cells; emptied HDL will dissociate from SR-B1 and reenter circulation

SR-B1 receptors are upregulated when cells need cholesterol for biosynthetic purposes but not
downregulated when cholesterol lvls high

HDL interaction with other particles

HDL also exchanges apoproteins and lipids with other lipoproteins in the blood (HDL transfers apoE and
apoCII to chylomicrons and to VLDL; apoCII activates LPL to degredate triacylglycerol)

when HDL obtains free cholesterol, the cholesterol is esterified via LCAT rxn

HDL transports the free cholesterol or cholesterol esters to the liver directly or by CETP (cholesterol
ester transfer protein) to circulating triacylglycerol-rich lipoproteins such as VLDL and VLDL remnants
(in exchange, triacylglycerol will be transferred to HDL); the greater the concentration of triacylglycerolrich lipoproteins in blood, the greater will be the rate of these exchanges

Mature HDL are HDL3; undergoes reverse cholesterol transport and becomes artherogenic protective
form HDL2; CETP rxn will then lead to loss of cholesterol and gain of triacylglycerol and regenerate HDL3;
hepatic lipase then removes triacylglycerol to regenerate HDL2
Lipoprotein enter cells by receptor-mediated endocytosis

interaction of ligand and receptor (apoprotein and plasma membrane receptor on target tissues) starts endocytosis

receptors for LDL are found in clathrin coated pits on plasma membrane of target cell

plasma membrane will invaginate and fuse into an endocytic vesicle which will fuse with lysosomes

the cholesterol esters are hydrolyzed and then quickly reesterfied by ACAT into monounsaturated FA (as opposed to previous
polyunsaturated FA)

LDL receptor is subject to feedback inhibition by increasing lvls of cholesterol within cell (also uses SREBP mechanism)

when cholesterol lvls decrease both cholesterol synthesis from acetyl-CoA and synthesis of LDL receptors are stimulated
Lipoprotein receptors

LDL receptor

recognizes apoB100 and apoE (binds to VLDL, IDL, chylomicrons, and LDL)

has six different regions

LDL binding region (cysteine-rich, when bound to LDL, bound to calcium ion)

2nd region is like a propeller

3rd region has N-linked oligosaccharides

4th region rich in serine & threonine with O-linked sugars (may extend receptor to allow LDL better
access to molecule)

5th region is transmembrane domain

6th region extends in cytosol where it regulates interaction between C-terminal of the receptor and the
clathrin coated pit

number of LDL receptors, binding, and post-receptor binding process can be diminished leading to LDL
cholesterol in blood  artherosclerosis

abnormalities can result from mutations for the LDL receptor (familial hypercholesterolemia) there are 4 classes
of mutations:

“null” alleles make no protein at all

receptor proteins made but they cannot be transported to cell surface

receptor proteins make it to surface but don’t bind LDL normally

reach surface and bind LDL but can’t cluster and internalize LDL particles

LDL receptor-related protein (LRP)

related to LDL receptor but recognizes broader spectrum of ligands

recognizes apoE or lipoproteins and binds remnants produced by digestion of triacylglycerols of chylomicrons
and VLDL by LPL (one of its functions is to clear this from blood)

LRP receptor abundants in cell membranes of liver, brain, placenta


VII.
VIII.

IX.
X.
synthesis is not affected by increase in intracellular cholesterol; insulin causes number of receptors to increase
(chylomicrons need to be removed after meal)

macrophage scavenger receptor

nonspecific receptors on macrophages that bind various types of molecules including oxidatively modified LDL
particles

SR-B1 is used for HDL; SR-A1 and SR-A2 are expressed on macrophages

LDL often modified and undergoes oxidative damage due to superoxide radicals, nitric oxide, hydrogen peroxide,
etc.

scavenger receptors are not subject to downregulation, so they can continue to take up oxidatively modified LDL
(even after elevated cholesterol lvls)

macrophages engorged with lipids are called foam cells and accumulation of them in subendothelial space of BVs
form early developing atherosclerotic plaque (fatty streak)

antioxidants (vitamin E, absorbic acid, carotenoids, may help protect LDL from oxidation)
Anatomic and biochemical aspects of atherosclerosis

initial step of atherosclerotic lesion in wall of artery is formation of a fatty streak

fatty streak is an accumulation of foam cells in the subintimal space that is visible as a yellow-white streak that bulges slightly
into lumen of vessel

risks involve arterial hypertension, elevated levels of LDL/chylomicron remnants/VLDL remnants, low HDL lvls,
smoking, chronic elevation of blood glucose, high lvls of angiotensin II (vasoconstrictor)

insult to endothelial cells trigger cells to secrete adhesion molecules that bind to monocytes which leads to a slow rate of
movement past endothelium and accumulation of monocytes; they will then gain entry into spaces between endothelial cells
(this is a classical inflammatory response therefore can be prevented through use of anti-inflammatory such as aspirin and
statins)

monocytes then become macrophages that migrate thru spaces between endothelial cells and enter subintimal space
(influenced by chemoattractant cytokines)

macrophages replicate and start internalizing FA leading to foam cell formation; foam cells accumulate in the subintimal space
and deform overlying endothelium; foam cells eventually become exposed through spaces between endothelial cells and also
expose underlying ECM to the blood

exposed areas will adhere to platelets  aggregation of platelets

activated platelets secrete cytokines  thrombus formation

plaque matures; fibrous cap forms over expanding “roof” which starts to occlude vessel

cap ruptures and plaque contents contact procoagulant elements in blood leading to acute thrombus formation which can
eventually lead to an infarction to surrounding tissues
Steroid hormones

steroid hormones must be complexed with a serum protein because of their hydrophobicity; albumin is a nonspecific carrier
but specific carriers exist

glucocorticoids (cortisol) synthesis and secretion are stimulated by ACTH from anterior pituitary gland

mineralocorticoids (aldosterone) secreted in response to angiotensin II or III, rising potassium lvls in blood, or hyponatremia
(low sodium lvl in blood)

androgens (testosterone) synthesized in leydig cells and secreted in response to LH; estrogens are synthesized in ovarian
follicle and corpus luteum stimulated by FSH

progestogens (progesterone) synthesized in corpus luteum and stimulated by LH

cholesterol is converted to progesterone in first 2 steps of steroid hormone synthesis

cytochrome P450SCC side-chain cleavage enzyme in mitochondrial inner membrane removes 6 carbons from side
chain of cholesterol  pregnenolone (21C)

pregnenolone to progesterone is catalyzed by 3-β-hydroxysteroid dehydrogenase

certain enzymes are used in more than one pathway of steroid synthesis so a defect in these enzymes will lead to multiple
abnormalities in steroid synthesis

synthesis of cortisol

free cholesterol goes to inner mitochondria to become pregnenolone

pregnenolone returns to cytosol to become progesterone

C17 of progesterone is hydroxylated in ER membranes (catalyzed by P450 C17) and becomes 17-αhydroxyprogesterone (this may be cleaved to form androgen & estrogen precursors)

non-cleaved 17-α-hydroxyprogesterone are hydroxylated again by P450C21 to form 11-deoxycortisol

11-deoxycortisol is transpoted into mitochondria inner membrane and P450C11 β-hydroxylates substrate at C21
(using oxygen and NADPH) to form cortisol

rate of formation depends on stimulation of adrenal cortical cells by ACTH

synthesis of aldosterone

cholesterol is formed into progesterone in the same way as above

progesterone is hydroxylated at C21 by P450C21 to form DOC

DOC is catalyzed by P450C11 into corticosterone

corticosterone is catalyzed by P450 aldosterons system into 18-hydroxycorticosterone

18-hydroxycorticosterone is oxidized into aldosterone

primary stimulus is octapeptide angiotensin II (also hyperkalemia aka high blood potassium or hyponatremia aka
low blood sodium)

synthesis of adrenal androgens

proceeds from cleavage of 2C sidechain of 17-hydroxypregnenolone at C17 to form 19C adrenal androgen
dehydroepiandrosterone (DHEA) and its sulfate derivative (DHEAS) in zona reticulosum of adrenal cortex

androstenedione is produced by oxidation of β-hydroxy group to a carbonyl group by 3-β-hydroxysteroid
dehydrogenase; this androgen is converted to testosterone

synthesis of testosterone

stimulated by LH from anterior pituitary

most usual pathway is one described above

rate-limiting step is conversion of cholesterol to pregnenolone

synthesis of estrogens and progesterone

ovarian estrogens are C18 steroids with phenolic hydroxyl group at C3 and either a hydroxyl group (estradiol) or
ketone group (estrone) at C17

ovarian granulosa cell, in response to stimulation by FSH from anterior pituitary gland and through catalytic
activity of P450 aromatase converts testosterone to estradiol
XI.
Vitamin D synthesis

vitamin D can be obtained from diet or be synthesized from a cholesterol precursor

calciferols are a family of steroids that affect calcium homeostasis

cholecalciferol requires UV light for its production from 7-dehydrocholesterol

irradiation cleaves C-C bond at C9-C10 opening the B-ring to form cholecalciferol (in inactive precursor of 1,25(OH)2-cholecalciferol (calcitriol); calcitriol is the most potent biologically active form of vitamin D

formation of calcitriol from cholecalciferol begins in the liver and ends in the kidney; carbon 25 of vitamin D 2 or D3 is
hydroxylated in liver to form calcidiol (25-hydroxycholecalciferol); calcidiol circulates to the kidney bound to vitamin Dbinding globulin (transcalciferin); in the proxima convolued tubule, a mixed0function oxidase that requires molecular O 2 and
NADPH, hydroxlyates carbon 1 on A-ring to form calcitriol

this last step is the rate-limiting step and is activated by PTH from parathyroid gland

calcitriol is 100x more potent than 25-(OH)D3 yet this is present in much greater concentration (suggests it plays a role in
calcium and phosphorus homeostasis)

Vitamin D like other sterol hormones diffuse passively thru plasma membrane

in intestinal cells, D3 and receptor complex activate genes that make proteins that transport Ca 2+ from gut lumen
into circulation
CLINICAL COMMENTS

When cholesterol levels are extremely high with normal triacylglycerol levels in a patient and similar lipid abnormalities are present in
other family members and no secondary cause are present, then familial hypercholesterolemia (FH), type IIA should be suspected

FH is a genetic disorder caused by abnormality in 1 or more alleles responsible for formation of LDL receptors

chronic hypercholesterolemia may cause deposition of lipid in vascular tissues leading to atherosclerosis
o
also may deposit in the skin and eye (medial aspect of upper and lower eyelids is called xanthelasma and iris of the eye is
called xanthomas)

ezetimibe is a drug that blocks cholesterol absorption in intestine (decreases chylomicron-based cholesterol)

HMG-CoA reductase inhibitor (atorvastatin) stimulate the synthesis of additional LDL receptors by inhibiting HMG-CoA reductase (ratelimiting enzyme for cholesterol synthesis); this leads to decreased intracellular free cholesterol and stimulates the synthesis of additional
LDL receptors

adrenal cortex makes DHEA and ovary makes very little so testing for this can see where the source of male hormone is coming from

when a patient is deficient in the P450C11 enzyme, it can’t convert 11-deoxycortisol to cortisol so this drop in blood cortisol level will
produce ACTH; however, ACTH will also signal for increased production of adrenal androgens (DHEA) causing virilization
o
prednisone is a synthetic glucocorticoid that can prevent ACTH-induced overproduction of adrenal androgens
BIOCHEMICAL COMMENTS
Metformin
o
treatment for type 2 diabetes
o
reduces blood glucose lvls by inhibiting hepatic gluconeogenesis and lipogenesis

AMPK phosphorylates a coactivatory of CREB txn factor (TORC2)

when TORC2 is phosphorylated, CREB inhibited from upregulating PGC1α (activator for gluconeogenic enzymes)

AMPK decreases txn of key lipogenic enzymes (FA synthase and acetyl-CoA carboxylase)

AMPK inhibits txn of SREBP-1 which regulates txn of HMG-CoA reductase as well as other lipogenic
enzymes
o
activates AMP-activated protein kinase (AMPK) by activating an upstream protein kinase, LKB1
o
AMPK when active phosphorylates and reduces activity of acetyl-CoA carboxylase (req. for FA synthesis) and HMG-CoA
reductase (reducing biosynthesis of cholest.); also activates glucose uptake by muscle (reducing circulating blood glucose)
Fibrates
o
used to decrease lipid levels (triglycerides)
o
major target is peroxisome proliferator-activated receptor-α (PPARα); when fibrate binds, it activates, which then leads to txn
of many genes that degrade lipids

PPARα activation enhances LPL expression, represses apoCIII expression (apoCIII inhibits apoCII activation of LPL),
stimulates apoAI and apoAII synthesis (major proteins of HDL)
Thiaxolidinediones (TZDs)
o
treatment of insulin resistance and type 2 diabetes mellitus
o
activates PPARγ class of txn factors which is expressed in adipose tissue
o
this txn factor is responsible for activating the txn of adiponectin, leading to increased circulating adiponectin levels which
reduces fat content of liver and enhances insulin sensitivity via AMPK-dependent pathway
o
also leads to reduction in plasma free FA levels, leading to enhanced insulin sensitivity
Disease or Disorder
Hypercholesterolemia
Environmental/Genetic
Both
Familial hypercholesterolemia, type II
Genetic
Virilization
Congenital adrenal hyperplasia (CAH)
Both
Genetic
Rickets
Environmental
Comments
Defined by elevated levels of cholesterol in the blood, often leading
to coronary artery disease
Defect in LDL receptor, leading to elevated cholesterol levels, and
premature death caused by coronary artery disease
Excessive release of androgenic steroid due to a variety of causes
CAH is a constellation of disorders caused by mutations in enzymes
required for cortisol synthesis. One potential consequence is
excessive androgen synthesis, which may lead to prenatal
masculinization of females. The different symptoms observed
between patients are caused by different enzyme deficiencies in the
patients.
Due to lack of vitamin D, calcium metabolism is altered, leading to
skeletal deformities