Transcript Slide 1

LIPID MAPS Lipid Metabolomics Tutorial
Fatty Acid Oxidation
Professor Edward A. Dennis
Department of Chemistry and Biochemistry
Department of Pharmacology, School of Medicine
University of California, San Diego
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Attribution: Edward A. Dennis (2010) “LIPID MAPS Lipid Metabolomics Tutorial” www.lipidmaps.org
E.A. DENNIS 2010 ©
Metabolism and Energy Overview
Proteins
Amino
Acids
Carbohydrates
Simple
Sugars
Lipids
Fatty
Acids
Pyruvate
Acetyl CoA
• Triglycerides (TG’s) are
decomposed into Fatty
Acids (FA’s).
• FA’s are b-oxidized to
release energy and create
acetyl CoA fragments.
• Acetyl CoA enters the
power-producing Krebs
cycle and electron
transport chain.
Energy
(CO2, H2O)
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Naming Conventions: Palmitic Acid
b
d
16
6
e

5
4
g
3
2
1
a
Carbonyl carbon

e
d
g
b
a
omega, always the last alkyl carbon
epsilon, fifth carbon after the carbonyl
delta, fourth carbon after the carbonyl
gamma, third carbon after the carbonyl
beta, second carbon after the carbonyl
alpha, first carbon after the carbonyl
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Beta Oxidation In a Nutshell...
Fatty Acid
One iteration of b-Oxidation:
Make fatty acyl CoA.
Acyl CoA
(1)
Step 1: Oxidize the b-carbon (C3)
Enoyl CoA
(2)
L-hydroxyacyl CoA
(3)
Ketoacyl CoA
(4)
Step 2: Hydrate the b-carbon
Step 3: Oxidize the b-carbon, again!
Step 4: Thiolyze a-b bond, releasing acetyl CoA
REPEAT from step 1, w/ 2 fewer carbons
Acyl CoA
(shorter)
Acetyl CoA
TCA Cycle
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Make activated acyl CoA
Triglycerides
Hormone sensitive
lipase
Free FA’s
adipocyte
Albumin
– carries FA’s to target tissue
bloodstream
Free FA’s
– decomposes triglycerides
– removes FA groups
– free FA’s diffuse through
membrane
• Albumin
Free FA’s
Fatty acyl CoA
synthase
• Hormone-sensitive lipase
– FA’s diffuse into tissue cells
Acyl CoA
• Fatty acyl CoA synthase
– adds CoA to the free FA
• Carnitine shuttle
mitochondria
Acyl CoA
Carnitine
shuttle
– moves fatty acyl CoA into
the mitochondria
target tissue cell
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Regulation
• Hormone Sensitive Lipase is activated by cAMP
• Cyclic AMP also turns off acetyl CoA
carboxylase, stopping FA synthesis
• Hormones like glucagon and epinephrine
increase cAMP
– FA synthesis slows
– Triglycerides are broken down
– FA’s enter b-oxidation faster
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Location, Location, Location
• Fatty Acyl CoA is oxidized inside the
mitochondrial matrix
• The carnitine shuttle moves it into
the matrix
– Free carnitine is exchanged for acyl
carnitine
• Carnitine acyl transferases catalyze
reactions on both sides of membrane
– driven by concentration gradient
• CAT-1 is inhibited by high levels of
malonyl CoA generated by acetyl CoA
carboxylase in the pathway to Fatty
Acid Synthesis.
Figure: Voet, D, Voet JG, Pratt CW (2006), Fundamentals of Biochemistry: Lif e at the Molecular Level, 2 nd ed. Reprinted
with permission of John Wiley & Sons, Inc.
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Step 1: Oxidize the b-carbon
• Acyl CoA dehydrogenase
oxidizes the b-carbon
Fatty acyl CoA
FAD
Acyl CoA
dehydrogenase
FADH2
– 3 versions of the enzyme
exist
– Specific to short, medium
and long chain substrates
• One FADH2 is generated
– makes 2 ATP’s
• A trans double bond is
created at the 2-carbon
• Product is an Enoyl CoA
D 2-trans-enoyl CoA
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Medium-chain dehydrogenase deficiency
• Incidence: 1 in 10,000 live births
Fatty Acid
Acyl CoA
(1)
Enoyl CoA
(2)
L-hydroxyacyl CoA
(3)
Ketoacyl CoA
(4)
Acyl CoA
(shorter)
– More common than phenylketonuria!
• Symptoms:
– severe hypoglycemia >> lethargy, coma
• little energy from FA’s
• glucose reserves are immediately burned
– contributes to sudden infant death syndrome (10%
of cases)
• Mechanism:
– Normally, there are 3 separate fatty acyl CoA
dehydrogenase enzymes for STEP 1 of b-oxidation
• Specific for short, medium and long acyl chains, respectively
– Autosomal recessive lack of medium chain enzyme
• Treatment: Special diet and supportive care
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Step 2: Hydrate the b-carbon
• Enoyl CoA hydratase
adds a water molecule
across the double bond
• Product is b-hydroxyacyl
CoA
– S- stereoisomer
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Step 3: Oxidize the b-carbon, Again!
• Beta-hydroxyacyl CoA
dehydrogenase
oxidizes the b-carbon
again
• One NADH is created
– makes 3 ATP’s
• A ketoacyl CoA is
produced
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Step 4: Thiolyze off acetyl CoA
CoA-SH
Acyl CoA:
Acyltransferase
(“thiolase”)
• Thiolase (aka: Acyl
CoA acyltranferase)
splits the ketoacyl
• A new CoASH is
consumed
• Acetyl CoA is released
• A new, shorter Acyl
CoA remains and reenters the cycle
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REPEAT with a Shorter Acyl CoA
• Palmitoyl CoA (16 carbons) becomes
myristoyl CoA (14 carbons).
• Each iteration releases another acetyl CoA.
• 7 iterations will release 8 acetyl CoA
fragments from the original palmitoyl CoA.
• Net equation:
Palmitoyl CoA + 7CoASH + 7FAD + + 7NAD+ + 7H20
yields: 8 Acetyl CoA + 7 FADH 2 + 7NADH + 7H+
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b-Oxidation Cycle
Removal of 2 carbons
from acyl chain
Fatty acyl-CoA
[1]
Acyl CoA
dehydrogenase
[4]
Acyl CoA:
acyltransferase
D 2-trans-enoyl CoA
b-ketoacyl CoA
Acetyl-CoA
[3]
b-hydroxyacyl CoA
dehydrogenase
enoyl CoA
hydratase
[2]
TCA cycle
b-hydroxyacyl CoA
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How Much Energy?
Each palmitoyl CoA group released from a triglyceride:
– directly produces 7 NADH & 7 FADH2
• which generate 21 and 14 ATP’s, respectively
– releases 8 acetyl CoA molecules for TCA Cycle
• which each generate 12 ATP
Grand total is 131 ATP per fully oxidized palmitoyl CoA
Efficiency of Energy Recovery = 40%
** Recently, some have calculated energetically less ATP
equivalents per NADH/FADH2 [2.5 and 1.5 instead of 3 and 2]
which lowers the numbers somewhat (Berg).
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Efficiency of Fat Storage
Fat has 9 kcal/gram = 9 kcal/cc
Both Carbohydrate and Protein have 4
kcal/gram = 4 kcal/3cc or 1.33 kcal/cc
Fat/(Carbohydrate or Protein) = 6x/cc!!!
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Problem: Oxidizing Unsaturated FA’s
Linoleic Acid
C12 (Even # Unsat) C9 (Odd # Unsat)
3 NAD+ + 3FAD + CoA-SH
3 rounds of
b- oxidation
3 NADH + 3FADH2 + 3Acetyl-CoA
Problem: b-g cis
double bond
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Special Case 1: Odd Unsaturations
The GOOD News:
• Enzyme enoyl-CoA
isomerase moves the double
bond over 1 position.
• Oxidation process resumes
The BAD News:
• The first oxidation step is
skipped
• One less FADH2 is made
– 2 fewer ATPs
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Special Case 2: Even Unsaturations
4
2
The GOOD News:
• Enzymes 2,4 dienoyl-CoA
reductase and 3,2 dienoyl-CoA
isomerase convert the g-d and
a-b double bonds into a single
a-b unsaturation.
• Oxidation process resumes
3
H+ + NADPH
NADP+
3
4
2
3
4
The BAD News:
• The conversion costs one
NADPH directly.
• Net effect is the loss of one
NADH
– 3 fewer ATPs
2
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FA Synthesis vs. FA Oxidation
Synthesis
Oxidation
Cytosol
Mitochondria
ACP
CoA
Malonyl CoA
Acetyl CoA
b-hydroxyl acyl step
R-config
S-config
Electron carriers
NADPH
NADH, FADH
Liver
Muscle, liver
Cell Location
Acyl carrier
2-Carbon Piece
Primary tissue site
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FA Synthesis vs. FA Oxidation (cont)
1. They are not the reverse of one another.
– Different subcellular locations
– R and S isomers of b-hydroxyacyl intermediates
cannot easily jump to the other pathway
– Electrons donated from oxidation (NADH, FADH2)
cannot directly enter synthesis, which uses NADPH.
2. Separate, semi-independent pathways allow
more sophisticated regulation.
– Accelerating one pathway does not mean slowing
the other.
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Acknowledgement
This tutorial is based on an evolving subset of lectures and
accompanying slides presented to medical students in the Cell
Biology and Biochemistry course at the School of Medicine of the
University of California, San Diego.
I wish to thank Dr. Bridget Quinn and Dr. Keith Cross for aid in
developing many of the original slides, Dr. Eoin Fahy for advice in
applying the LIPID MAPS nomenclature and structural drawing
conventions [Fahy et al (2005) J Lipid Res, 46, 839-61; Fahy et al
(2009) J Lipid Res, 50, S9-14] and Masada Disenhouse for help in
adopting to the tutorial format.
Edward A. Dennis
September, 2010
La Jolla, California
E.A. DENNIS 2010 ©