Organization of Fatty Acid Synthase from Animal Liver

1
BIOCHEMISTRY 441 Winter 2006
Dept. of Biochemistry
Bill Parson
9. Biosynthesis of fatty acids 10.
Phospholipids, triacylglycerols complex
lipids 11. Cholesterol lipoproteins 12.
Photosynthesis antenna reaction center
complexes 13. Photosynthesis photophosphorylation
14. Photosynthesis carbon fixation 15. Amino
acid catabolism transamination 16. Amino acid
catabolism urea cycle 17. N2 fixation amino
acid biosynthesis 18. Aromatic amino acids
neurotransmitters 19. One-carbon metabolism 20.
Biosynthesis of pyrimidines purines 21.
Deoxyribonucleotide biosynthesis nucleotide
catabolism
2
Fatty acids have extended hydrocarbon chains
Most natural fatty acids have an even number of
carbons. Unsaturated fatty acids usually have
cis double bonds.
3
Fatty acids are components of phospholipids and
triacylglycerols
triacylglycerols
glycerophospholipids
sphingolipids
Triacylglycerols are stored as energy reserves.
Sphingolipids on cell surfaces are sites of cell
recognition. Inositol phospholipids participate
in intracellular signaling.
Phospholipid bilayers are the central structural
elements of biological membranes.
Fatty acids also are found as cholesterol esters
in lipoproteins, and are attached covalently to
some proteins.
4
Triacylglycerols are stored as energy reserves in
adipose tissue and other tissues
capillary
lipid droplets
Cross section of four adipocytes from a guinea
pig. Lipid droplets, consisting mainly of
triacylglycerols, fill most of the volume of the
cells.
5
Fatty acids are synthesized from acetyl-CoA in
adipose tissue the liver
Why is CO2 needed?
The labeling patterns suggest that the fatty acid
chain forms by successive addition of two-carbon
units
6
MalonylCoA serves as the donor of two-carbon units
MalonylCoA is formed from acetylCoA and CO2 by a
multifunctional enzyme, acetylCoA carboxylase
(biotin carboxylase-transcarboxylase). Biotin is
attached covalently to a Lys residue of the
enzyme. In bacteria, the three domains are in
separate subunits in animals, they are on a
single, multifunctional polypeptide.
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The growing fatty acid chain is attached to
acyl-carrier protein (ACP)
ACP has 4-phosphopantetheine linked to a Ser
residue. Malonyl and acetyl groups are
transferred from CoA to the sulfur atom of the
4-phosphopantetheine.
Pantetheine is vitamin B5
8
Acyl-carrier protein from Bacillus subtilis
4-phosphopantetheine
www.rcsb.org/pdb
from pdb file 1f80.pdb K. D. Parris et al.
Struct. Fold. Design 8 883 (2000).
9
Malonyl-/Acetyl transferase (MAT) catalyzes
transfer of malonyl units from CoA to ACP
10
Malonyl-/Acetyl transferase (MAT) also transfers
acetyl units to ACP
Bacteria have separate malonyl acetyl
transferases
Acetyl units then move from ACP to the
keto-synthase (KS), which catalyzes the
condensation reaction
11
The condensation reaction
3-ketoacyl-ACP
malonyl-ACP
S
The acetyl group first is transferred from ACP to
a Cys residue of the synthase. Then it combines
with malonyl-ACP to give a 3-ketoacyl-ACP.
Release of bicarbonate is exothermic and pulls
the reaction in the direction of condensation.
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Reduction by NADPH, dehydration, and a second
reduction generates butyryl-ACP
dehydrase
13
To continue the cycle, the fatty acid chain must
move back to the ketoacyl synthase
b-ketoacyl synthase
CO2
H2O
b-ketoacyl reductase
enoyl reductase
dehydrase
HOH
14
A second turn of the cycle generates hexanoyl-ACP
b-ketoacyl synthase
CO2
b-ketoacyl reductase
enoyl reductase
dehydrase
HOH
15
The cycle stops when the fatty acid chain reaches
16 carbons
Thioesterase hydrolyzes palmitoyl-ACP, releasing
palmitate (C160)
C140
b-ketoacyl synthase
CO2
thioesterase
C160
HOH
16
Chain-length specificity of the
substrate-loading, chain-elongation and
chain-termination activities of mammalian
fatty-acid synthase
S. Smith et al. Prog. Lipid Res. 42 289-317
(2003)
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The enzymes of the fatty acid synthase system
have fused into a single protein during evolution
ACP
CO2-
H3N
O-phosphopantetheine
E. coli eight separate proteins
OH
18
Mammalian fatty-acid synthase is active only as a
dimer
Do the two subunits work independently or
cooperatively?
19
Testing models of the fatty-acid synthase dimer
monomers with inactivating mutations in two
different domains
Is the heterodimer active? The answer depends on
which domains are mutated.
20
Suggested model of mammalian fatty-acid synthase
dimer
solid arrows substrate loading condensation.
dotted arrows b-carbon processing
chain-termination
S. Smith et al. Prog. Lipid Res. 42 289-317
(2003)
21
Differences between fatty acid synthesis and
oxidation
oxidation
synthesis
H2O
cytosol
22
Fatty acids with longer chains (C180 C200)
are synthesized from palmitoylCoA malonylCoA
by an elongation mechanism






These reactions occur in mitochondria the
smooth ER.
NADPH
Different enzymes are involved, and CoA is used
in place of ACP, but the reactions are otherwise
formally the same as in synthesis of palmitate.
NADP
H2O
23
Citrate carries 2-carbon units from mitochondria
to the cytosol
Cytosol
citrate synthase
Mitrochondrion
acetyl-CoA
CoA-SH
CH3CO-S-CoA
oxaloacetate
citrate transporter
CoA-SH ATP
citrate lyase
citrate
ADP Pi
CH3CO-S-CoA
Citrate lyase uses ATP to drive the breakdown of
citrate to acetylCoA oxaloacetate in the
cytosol
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Integration of fatty acid synthesis with
carbohydrate metabolism
fatty acids
citrate
citrate
NADPH
CoA-SH
CoA-SH
ATP
pyruvate
ATP
citrate lyase
acetylCoA
acetylCoA
ADP Pi
TCA cycle
amino acids
oxaloacetate
oxaloacetate
NADH
malate dehydro-genase
NADH
ox. phos.
NAD
ATP
NAD
malate
malate
NADP
pyruvate carboxylase
malic enzyme
NADPH CO2
pyruvate
pyruvate
glucose
25
AcetylCoA carboxylase (biotin carboxylase/transcar
boxylase) is the main control point for fatty
acid synthesis in animals
the enzyme is regulated by both allosteric
effects and phosphorylation
citrate lyase
insulin stimulates dephosphorylation (activation)
the phosphorylated enzyme is inactive
acetyl-CoA carboxylase
-O--P
cAMP-dependent protein kinase
glucagon, epinephrine, and adiponectin stimulate
phosphorylation (inactivation)
malonylCoA inhibits carnitine-acyltransferase I,
blocking transport of palmitoylCoA into
mitochondria for oxidation
carnitine-acyltransferase I
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The active (unphosphorylated) form of acetylCoA
carboxylase forms long filaments
27
An imbalance between energy input and output can
lead to obesity
65 of the adult U.S. population are considered
to be overweight (BMI gt 25) 35 are obese (BMI
gt 30). More than 10 of U.S. children aged 2 to
5 are overweight. Obesity raises the risk of
heart disease, stroke, type-II diabetes and
cancer.
BMI (weight in kg)/(height in m)2
703x(weight in pounds)/(height in inches)2
28
Leptin and adiponectin convey signals of
nutritional excess
blood
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Defects in leptin or its receptor can cause
obesity
weight 35 g
weight 67 g
These mice are the same age. Both are homozygous
for a defective variant of leptin. The mouse on
the right received daily injections of purified
leptin the mouse on the left was not
treated. But most obese humans do not have a
deficiency in leptin.
30
Ghrelin and other hormones convey signals of
short-term hunger or satiety
Youre full! Stop eating!
Youre hungry! Eat!
J. Marx, Cellular Warriers at the Battle of the
Bulge Science 299 846 (2003)