Transcript Fatty Acid Synthesis

Chapter 24
Lipid Biosynthesis
Biochemistry
by
Reginald Garrett and Charles Grisham
Outline
1. How Are Fatty Acids Synthesized?
2. How Are Complex Lipids Synthesized?
3. How Are Eicosanoids Synthesized, and What
Are Their Functions?
4. How Is Cholesterol Synthesized?
5. How Are Lipids Transported Throughout the
Body?
6. How Are Bile Acids Biosynthesized?
7. How Are Steroid Hormones Synthesized and
Utilized?
24.1 – How Are Fatty Acids
Synthesized?
The Biosynthesis and Degradation
Pathways are Different
• As in cases of glycolysis/gluconeogenesis and
glycogen synthesis/breakdown, fatty acid
synthesis and degradation go by different
routes
• There are five major differences between fatty
acid breakdown and biosynthesis
The Differences Between fatty acid
biosynthesis and breakdown
1. Intermediates in synthesis are linked to
-SH groups of acyl carrier proteins (as compared to
-SH groups of CoA)
2. Synthesis in cytosol; breakdown in mitochondria
3. Enzymes of synthesis are one polypeptide (fatty
acid synthase) in animals (bacteria and plants
employ separate enzymes)
4. Biosynthesis uses NADPH/NADP+; breakdown
uses NADH/NAD+ & FAD
5. Stereochemistry (D-b-hydroxyacyl)
Activated form: Malonyl-CoA
Acetate Units are Activated for Transfer in
Fatty Acid Synthesis by Malonyl-CoA
1. Fatty acids are built from 2-C units -- acetylCoA
2. Acetyl-CoA are activated by formation of
malonyl-CoA
3. Decarboxylation of malonyl-CoA and reducing
power of NADPH drive chain growth
4. Chain grows to 16-carbons (palmitate)
5. Other enzymes add double bonds and more Cs
The sources of Acetyl-CoA
1. Amino acid degradation produces cytosolic
acetyl-CoA
2. Fatty acid oxidation produces mitochondrial
acetyl-CoA
3. Glycolysis yields cytosolic pyruvate which
is converted to acetyl-CoA in mitochondria
• Citrate-malate-pyruvate shuttle provides
cytosolic acetate units and reducing
equivalents for fatty acid synthesis
Citrate-malate-pyruvate shuttle
The reducing power: NAPDH
1. Produced in pentose phosphate pathway
(chapter 22)
2. Produced by malic enzyme
Malate + NADP+ → pyruvate + CO2 + NADPH + H+
The "ACC enzyme" commits acetate
to fatty acid synthesis
Acetyl-CoA Carboxylase (ACC)
• Carboxylation of acetyl-CoA to form malonylCoA is the irreversible and committed step in
fatty acid biosynthesis
• ACC uses bicarbonate and ATP (& biotin)
– E.coli enzyme has three subunits
– Animal enzyme is one polypeptide with all three
functions - biotin carboxyl carrier protein, biotin
carboxylase and transcarboxylase
Acetyl-CoA + ATP + HCO3- → malonyl-CoA + ADP + Pi + H+
Malonyl-CoA
Figure 24.2 (a) The acetyl-CoA carboxylase
reaction produces malonyl-CoA for fatty acid
synthesis. (b) A mechanism for the acetylCoA carboxylase reaction.
Figure 24.3 In the acetyl-CoA carboxylase reaction, the biotin ring, on its flexible tether,
acquires carboxyl groups from carbonylphosphate on the carboxylase subunit and transfers
them to acyl-CoA molecules on the transcarboxylase subunits.
Acetyl-CoA Carboxylase
ACC forms long, active filamentous polymers
from inactive protomers
• As a committed step, ACC is carefully regulated
– Palmitoyl-CoA (product) favors monomers
– Citrate favors the active polymeric form
• Phosphorylation modulates citrate activation and
palmitoyl-CoA inhibition
Citrate
Inactive protomers
active polymer
Acyl-CoA
Figure 24.4 Schematic of the acetyl-CoA carboxylase,
with domains and phosphorylation sites indicated, along
with the protein kinases responsible.
Phosphorylation of ACC modulates
activation by citrate and inhibition by
palmitoyl-CoA
• The animal enzyme is phosphorylated at 8 to 10 sites
on each enzyme subunit (Figure 24.4)
• Unphosphorylated E
– Has high affinity for citrate and is active at low [citrate]
– Has low affinity for palmitoyl-CoA and needs high
[palmitoyl-CoA] to inhibit
• Phosphorylated E
– Has low affinity for citrate and needs high [citrate] to activate
– Has high affinity for palmitoyl-CoA and is inhibited at low
[palmitoyl-CoA]
Figure 24.5 The activity of acetyl-CoA carboxylase is modulated by phosphorylation and
dephosphorylation.
The dephospho form of the enzyme is activated by low [citrate] and inhibited only by high
levels of fatty acyl-CoA. In contrast, the phosphorylated form of the enzyme is activated
only by high levels of citrate, but is very sensitive to inhibition by fatty acyl-CoA.
The Acyl Carrier Protein (ACP)
• The acyl carrier protein carry the intermediates
in fatty acid synthesis
• ACP is a 77 residue protein in E. coli - with a
phosphopantetheine
• The same functional group of CoA
Fatty Acid Synthase (FAS)
• Fatty acid synthesis in mammals occurs on
homodimeric fatty acyl synthase I (FAS I)
– consists of 270 kD polypeptides which contain
all reaction centers required to produce a fatty
acid
• In yeast and fungi (lower eukaryotes), the
activities of FAS are distributed on two
multifunctional peptide chains (a6b6)
• In plants and bacteria, the enzymes of FAS
are separated and independent, and this
collection of enzymes is referred to as fatty
acid synthase II (FAS II)
Structure of the Fatty Acid Synthase
Fatty Acid Synthesis
• The individual steps of fatty acid synthesis are
similar across all organisms
• The mammalian pathway (Figure 24.7) is a cycle
of elongation that involves six enzyme activities
• Elongation is initiated by transfer of the acyl
moiety of acetyl-CoA to the acyl carrier protein
by the malonyl-CoA-acetyl-CoA-ACP
transacylase (MAT)
• This enzyme also transfers the malonyl group
of malonyl-CoA to ACP
Fatty Acid Synthase
1. Malonyl-CoA-acetyl-CoA-ACP transacylases (MAT)
2.
b-ketoacyl-ACP synthase (KS)
3.
b-ketoacyl-ACP reductase (KR)
4.
b-hydroxyacyl-ACP dehydratase (DH)
5.
Enoyl-ACP reductase (ER)
6.
Thioesterase (TE) in animals
The pathway of palmitate
synthesis from acetyl-CoA
and malonyl-CoA.
Acetyl and malonyl building
blocks are introduced as acyl
carrier protein conjugates.
Decarboxylation drives the bketoacyl-ACP synthase and
results in the addition of twocarbon units to the growing
chain.
The first turn of the cycle
begins at “1” and goes to
butyryl-ACP; subsequent
turns of the cycle are
indicated as “2” through “6”.
Figure 24.7
Acetyl-ACP
Malonyl -ACP
Acetoacetyl -ACP
Butyryl -ACP
Figure 24.8 An acetyl group is transferred from CoA to MAT, then to the acyl
carrier protein, and then to the ketoacyl synthase. Next a malonyl group is
tranferred to MAT and then to the acyl carrier protein.
Fatty Acid Synthesis
Acetyl-CoA + 7 malonyl-CoA + 14 NAPDH + 14 H+
Palmitoyl-CoA + 7 HCO3- + 14 NAPD+ + 7 CoA-SH
The formation of malonyl-CoA:
7 Acetyl-CoA + 7 HCO3- + 7 ATP47 malonyl-CoA + 7 ADP3- + 7Pi2- +7H+
The overall reaction:
8 Acetyl-CoA + 7 ATP4- + 14 NAPDH + 14 H+
Palmitoyl-CoA + 14 NAPD+ + 7 CoA-SH + 7 ADP3- + 7Pi2-
Further Processing of C16 FAs
Additional elongation & Unsaturation
• Additional elongation occurs in
mitochondria and the surface of ER
• In ER
– Similar to fatty acid synthase
– Involving malonyl-CoA
• In mitochondrion (Figure 24.12)
– A reversal of fatty acid oxidation
– The reducing coenzyme for the second step is
NADH, whereas the reductant for the fourth
step is NADPH.
( in mitochondria)
Figure 24.12 Elongation of fatty
acids in mitochondria is initiated
by the thiolase reaction.
Introduction of cis double bonds
Introduction of cis double bonds:
1. Prokaryotes use an O2-independent process
2. Eukaryotes use an O2-dependent process
•
E.coli add double bonds during the fatty
acid synthesis
–
–
After four normal cycles, b-hydroxydecanoyl
thioester dehydrase forms a double bond b, g to
the thioester
O2-independent
Figure 24.13 Double bonds
are introduced into the
growing fatty acid chain in
E. coli by specific
dehydrases.
Eukaryotes add double bond until the fatty acyl
chain has reached its full length (usually 16
to 18 carbons)
– Stearoyl-CoA desaturase
– Cytochrome b5 reductase & Cytochrome b5
– All three proteins are associated with the ER
membrane
– NADH & O2 are required; O2-dependent
Mammals cannot synthesize most
polyunsaturated fatty acids
– Plants Can manufacture double bonds between the D9
and the methyl end of the chain, but mammals cannot
– Mammals can introduce double bonds between the
double bond at the 8- or 9- position and the carboxyl
group
– Form a double bond at 5-position (6) if one already
exists at the 8-position (9)
Essential fatty acids: mammals require
polyunsaturated fatty acids (PUFAs) but must
acquire them in their diet.
Arachidonic acid is
systhesized from
linoleic acid
• Linoleic acid is
acquired from diet
• Mammals can add
double bonds to
unsaturated fatty acids
• Is the precursor for
prostaglandins and
other biologically
active derivatives
Figure 24.15 Arachidonic acid is synthesized
from linoleic acid in eukaryotes.
ω3 and ω6 – Essential Fatty Acids with Many Functions
Regulation of Fatty Acid Synthesis
1.
•
•
•
Allosteric regulation
Citrate activates acetyl-CoA carboxylase
Fatty acyl-CoAs inhibit acetyl-CoA carboxylase
Malonyl-CoA blocks the carnitine acyltransferase
and thus inhibits b-oxidation
Regulation of FA Synthesis
2. Hormone signals regulate ACC and fatty
acid biosynthesis
–
–
–
–
Glucagon activates lipases/inhibits ACC
Insulin inhibits lipases/activates ACC
Phosphorylation causes inhibition of fatty acid
biosynthesis
Inactivated ACC can be reactivated by a
specific phosphatase induced by insulin
Figure 24.17
Hormonal signals regulate fatty acid synthesis, primarily through
actions on acetyl-CoA carboxylase. Availability of fatty acids also
depends upon hormonal activation of triacylglycerol lipase.
24.2 – How Are Complex Lipids
Synthesized?
Complexed lipids:
1. Glycerolipid
–
–
Triacylglycerols
Glycerophospholipids
2. Sphingolipids
Phospholipids (membrane components)
1. Glycerophospholipids
2. Sphingolipids
Different organisms posses greatly different
complements of lipids
Synthetic pathways depend on different
organism
• Sphingolipids and triacylglycerols only
made in eukaryotes
• Phosphatidylethanolamine (PE) accounts
for 75% of phospholipids in E.coli
– With phosphatidylglycerol (PG) and cardiolipin
– No PC, PI, sphingolipids, cholesterol in E.coli
– But some bacteria do produce PC
Glycerolipid Biosynthesis
Glycerolipids are synthesized by
phosphorylation & acylation of glycerol
• Phosphatidic acid (PA) is the precursor for all
other glycerolipids in eukaryotes
• Eukaryotic systems can also utilize DHAP as
a starting point
(PA)
Figure 24.19
Satuated
Fatty Acid
Unsatuated
Fatty Acid
Figure 24.18 Synthesis of glycerolipids in eukaryotes
begins with the formation of phosphatidic acid, which
may be formed from dihydroxyacetone phosphate or
glycerol as shown.
Glycerolipid Biosynthesis
Phosphatidic acid (PA) is converted either
to DAG or CDP-DAG
• DAG is a precursor for synthesis of TAG,
phosphatidylethanolamine (PE) and
phosphatidylcholine (PC)
– TAG is synthesized mainly in adipose tissue, liver,
and intestines
• CDP-DAG is a precursor for synthesis of
phosphotidylserine (PS), phosphatidylglycerol
(PG), cardiolipin and phosphatidylinositol (PI)
Figure 24.19
Diacylglycerol and CDPdiacylglycerol are the
principal precursors of
glycerolipids in eukaryotes.
Phosphatidylethanolamine
and phosphatidylcholine
are formed by reaction of
diacylglycerol with CDPethanolamine or CDPcholine, respectively.
Glycerolipid Biosynthesis
•
PE synthesis
1. begins with phosphorylation of ethanolamine to
form phosphoethanolamine
2. Transfer of a cytidylyl group from CTP to from
CDP-ethanolamine
3. Phosphoethanolamine transferase link
phosphoethanolamine to the DAG
•
•
Synthesis of PC is entirely analogous
PC can also be converted from PE by
methylation reactions
• Exchange of ethanolamine for serine converts
PE to PS (phosphatidylserine)
Figure 24.21 The interconversion
of phosphatidylethanolamine and
phosphatidylserine in mammals.
Other PLs from CDP-DAG
CDP-diacylglycerol is used in eukaryotes to
produce:
• Phosphatidylinositol (PI) in one step
– 2-8% in animal membrane
– Breakdown to form inositol-1,4,5-triphosphate
& DAG (second messengers)
• Phosphatidylglycerol (PG) in two steps
• Cardiolipin in three steps
Figure 24.22 CDP-diacylglycerol is
a precursor of phosphatidylinositol,
phosphatidylglycerol, and
cardiolipin in eukaryotes.
Plasmalogen Biosynthesis
Dihydroxyacetone phosphate (DHAP) is the
precursor to the plasmalogens
1. Acylation of DHAP
2. Exchange reaction produces the ether linkage
by long-chain alcohol (acyl-CoA reductase)
3. Ketone reduction
4. Acylation again
5. CDP-ethanolamine delivers the head group
6. A desaturase produces the double bond in the
alkyl chain
Figure 24.23 Biosynthesis of
plasmalogens in animals.
DHAP
1-Acyl-DHAP
1-Alkyl-DHAP
1-Alkyl-Glycero-3-P
1-Alkyl-2-acyl-Glycero-3-P
1-Alkyl-2-acyl-Glycero-3phosphoethanolamine
Plasmalogen
Acyl-CoA
Acyl-CoA reductase
Figure 24.24 Platelet-activating
factor, formed from 1-alkyl-2lysophosphatidylcholine by
acetylation at C-2, is degraded by
the action of acetylhydrolase.
Dilate blood vessels
Reduce blood pressure
Aggregate platelet
Sphingolipid Biosynthesis
High levels made in neural tissue
1. Initial reaction is a condensation of serine
and palmitoyl-CoA (by 3-ketosphinganine
synthase)
–
2.
3.
4.
•
3-ketosphinganine synthase is PLP-dependent
Ketone is reduced with help of NADPH
Acylation to form N-acyl-sphinganine
Desaturated to form ceramide
Ceramide is precursor for other sphingolipids
Sphingolipid Biosynthesis
Ceramide is precursor for other sphingolipids
• Sphingomyelin
–
–
–
•
Cerebrosides
–
–
•
Rich in myelin sheath
Insulates nerve axons
By transfer of phosphocholine from PC
Glycosylation of ceramide
Galactosylceramide makes up ~15% of the lipid of
myelin sheath
Gangliosides
–
Cerebrosides contain one or more sialic acid
Figure 24.26 Glycosylceramides
(such as galactosylceramide),
gangliosides, and sphingomyelins
are synthesized from ceramide in
animals.
Gangliosides
1.
UDP-glucose
2.
UDP-galactose
3.
UDP-N-acetylgalactosamine
4.
CMP-sialic acid
(N-acetylneuraminidate)
Fig. 8-10, p. 243
24.3 – How Are Eicosanoid Synthesized
and What Are Their Functions?
•
•
Eicosanoid are all derived from 20-carbon
fatty acid (arachidonic acid)
Eicosanoids are local hormones
1. Exert their effect at very low concentration
2. Usually act near their sites of synthesis
•
PLA2 releases arachidonic acid from
phospholipids (phosphatidylcholine)
–
May also be released by PLC & DAG lipase
Figure 24.27 Arachidonic acid,
derived from breakdown of
phospholipids (PL), is the
precursor of prostaglandins (PG),
thromboxanes (Tx), and
leukotrienes.
Specificities of phospholipases A1,
A2, C, and D. (Figure 8.18, p. 251)
Eicosanoids are local hormones
• Eicosanoids include
–
–
–
–
Prostaglandins (PG)
Thromboxanes (Tx)
Leukotrienes
Other hydroxyeicosanoic acid
• Tissue injury and inflammation triggers
arachidonate release and eicosanoid
synthesis
• Most PGs are cyclopentanoic acids
• Initiated by PGH synthase associated with
the ER
• PGH synthase (Prostaglandin
endoperoxide H synthase)
• Known as Cyclooxygenase
• The enzyme has two different
activities:
• Cyclooxygenase (COX)
• Peroxidase (POX)
Aspirin & NSAIDs
• Aspirin and other nonsteroid anti-inflammatory
drugs (NSAIDs) inhibit the cyclooxygenase
– Aspirin covalently
– Others noncovalently
(Tylenol)
(Advil)
Two isoforms in animals
(a) COX-1: normal, physiological production of PG
(b) COX-2: induced by cytokines, mitogens, and
endotoxins in inflammatory cells
Aspirin and bromoaspirin bind covalently to COX1 and COX-2.
Ibuprofen binds noncovalently to both.
Celebrex and Deramaxx bind selectively to COX-2.
COX-2 has Val
(blue) at position
523, whereas
COX-1 has Ile
(red).
24.4 – How Is Cholesterol Synthesized?
Occurs primarily in the liver
• The most prevalent steroid in animal cells is
cholesterol
• Serve as cell membranes, precursor of bile
acids and steroid hormones, and vitamin D3
Biosynthesis begins in the cytosol with the
synthesis of mevalonate from acetyl-CoA
1. First step is a thiolase reaction
2. Second step makes HMG-CoA
3. Third step produces 3R-mevalonate
• HMG-CoA reductase
• The rate-limiting step in cholesterol biosynthesis
• HMG-CoA reductase is site of regulation in
cholesterol synthesis → the action site of
cholesterol-lowering drugs
Figure 24.32 A reaction mechanism
for HMG-CoA reductase. Two
successive NADPH-dependent
reductions convert the thioester,
HMG-CoA, to a primary alcohol.
Regulation of HMG-CoA Reductase
As rate-limiting step, it is the principal site
of regulation in cholesterol synthesis
1. Phosphorylation by cAMP-dependent kinases
inactivates the reductase
2. Degradation of HMG-CoA reductase
• Half-life is 3 hrs and depends on cholesterol level
• High [cholesterol] means a short half-time
3. Gene expression (mRNA production) is
controlled by cholesterol levels
• If [cholesterol] is low, more mRNA is made
cAMP-dependent
protein kinase
Figure 24.33 HMG-CoA reductase activity
is modulated by a cycle of phosphorylation
and dephosphorylation.
Squalene is synthesized from Mevalonate
• Six-carbon mevalonate makes 5-carbon
isopentenyl pyrophosphate and dimethylallyl
pyrophosphate
• Condensation of these two 5-carbon
intermediates produces geranyl pyrophosphate
(10-carbon)
• Addition of another isopentenyl pyrophosphate
yields farnesyl pyrophosphate (15-carbon)
• Two farnesyl pyrophosphates link to form
squalene
Figure 24.34
The conversion of
mevalonate to squalene.
Geranyl
pyrophosphate
Cholesterol from Squalene
At the endoplasmic reticulum membrane
• Squalene monooxygenase converts squalene
to squalene-2,3-epoxide
• 2,3-oxidosqualene:lanosterol cyclase
converts the epoxide to lanosterol
• Though lanosterol looks like cholesterol, 20
more steps are required to form cholesterol
• All at/in the endoplasmic reticulum
membrane
Figure 24.35 Cholesterol is synthesized
from squalene via lanosterol.
The primary route from lanosterol involves
20 steps, the last of which converts 7dehydrocholesterol to cholesterol. An
alternative route produces desmosterol as
the penultimate intermediate.
Inhibiting Cholesterol Synthesis
• HMG-CoA reductase is the key - the ratelimiting step in cholesterol biosynthesis
• Lovastatin (mevinolin) blocks HMG-CoA
reductase and prevents synthesis of
cholesterol
• Lovastatin is an inactive lactone
• In the body, the lactone is hydrolyzed to
mevinolinic acid, a competitive inhibitor of
the reductase, Ki = 0.6 nM
For several
years, Lipitor
has been the
best-selling
drug in the
world, with
annual sales
exceeding $10
Billion.
The structures of (inactive lactone) lovastatin, (active)
mevinolinic acid, mevalonate, and synvinolin.
24.5 – How Are Lipids Transported
Throughout the Body?
Lipoproteins are the carriers of most lipids in the
body
• Lipoprotein - a cluster of lipids, often with a
monolayer membrane (phospholipids), together with
an apolipoprotein
• Classify lipoproteins according to their densities
– The densities are related to the relative
amounts of lipid and protein
– Proteins have densities of about 1.3 to
1.4 g/ml
– Lipids have densities of about 0.8 g/ml
The various apoproteins have an abundance of hydrophobic amino
acid residues, as is appropriate for interactions with lipids.
• HDL (high density lipoprotein) & VLDL (very low
density lipoprotein) are assembled primarily in the ER
of liver cells (some in intestines)
• LDL (low density lipoprotein) not made directly, but is
made from VLDL
– LDL appears to be the major circulatory complex for
cholesterol & cholesterol esters
• Chylomicrons form in the intestines
• Chylomicrons carry TAG & cholesterol esters
from intestine to other tissues
• VLDLs carry lipid from liver to other tissues
– In the capillaries of muscle and adipose cells,
lipoprotein lipases hydrolyze triglycerides from
lipoproteins, making the lipoproteins smaller and
raising their density
– Thus chylomicrons and VLDLs are progressively
converted to IDL (intermediate-density
lipoprotein) and then LDL, which either return to
the liver for reprocessing or are redirected to
adipose tissues and adrenal glands
Figure 24.38 Lipoprotein components are synthesized predominantly in the ER of liver
cells. Following assembly of lipoprotein particles (red dots) in the ER and processing in
the Golgi, lipoproteins are packaged in secretory vesicles for export from the cell (via
exocytosis) and released into the circulatory system.
ACAT is acyl-CoA cholesterol acyltransferase.
Figure 24.39 Endocytosis and degradation of lipoprotein particles.
Lipoproteins
• LDLs are main carriers of cholesterol and
cholesterol esters
• Newly formed HDL contains virtually no
cholesterol esters
– Cholesterol esters are accumulated through the
action lecithin:cholesterol acyltransferase
(LCAT)
• HDLs function to return cholesterol and
cholesterol esters to the liver
The LDL Receptor
The LDL receptor in plasma
membranes consists of 839
amino acid residues and is
composed of five domains
D1: LDL binding domain on N-terminus
D2 & D3: N-linked and O-linked
oligosaccharide domains
D4: A single TMS
D5: A cytosolic domain essential to
aggregation of receptors in the
membrane during endocytosis
Dysfunctions in or absence of LDL
receptors lead to familial
hypercholesterolemia
24.6 – How Are Bile Acids
Biosynthesized?
Carboxylic acid derivatives of cholesterol
• Essential for the digestion of food, especially for
solubilization of ingested fats
• Synthesized from cholesterol in the liver, stored
in the gallbladder, and secreted as need into the
intestine
• Cholic acid conjugates with taurine and glycine
to form taurocholic and glycocholic acids
• First step is oxidation of cholesterol by a mixedfunction oxidase (p-450, 7a-hydroxylase)
24.7 – How Are Steroid Hormones
Synthesized and Utilized?
• Steroid hormones are crucial signal molecules
• Biosynthesis begins with the desmolase
reaction (in mitochondria), which converts
cholesterol to pregnenolone, precursor to all
others
• Pregnenolone migrates from mitochondria to
ER where progesterone is formed
• Progesterone is a branch point - it produces sex
steroids (testosterone and estradiol), and
corticosteroids (cortisol and aldosterone)
(27C)
(21C)
(19C)
aromatase
(18C)
Figure 24.44
The steroid hormones are
synthesized from cholesterol
• Anabolic steroids are illegal and dangerous