Lipid Metabolism

1
Lipid Metabolism

• Hanley N. Abramson, Ph.D.
• Professor, Department of Pharmaceutical Sciences
• Wayne State University
• October 2009

2
Fatty Acids
Ester
Thioester
3
Fatty Acids as Stored Energy

• Fatty acids are the bodys principal form of
  stored energy
• Carbon almost completely reduced as CH2
• Very closely packed in storage tissues - not
  hydrated as sugars are

4
Dietary Fatty Acids

• Comprise 30-60 of caloric intake in average
  American diet
• Triacylglycerols, phospholipids, sterol esters
• Principal sources dairy products, meats

5
Digestion of Dietary Triacylglycerols

• Occurs in duodenum
• Facilitated by
• Bile salts (emulsification)
• Alkaline medium (pancreatic juice)

Blocked by Orlistat (Fat Blocker) - Xenical/Alli
Pancreatic lipases
OH
Intestinal lipases
Glycerol Fatty Acids
OH
TAG MAG
6
Epithelial Cell (Intestinal Wall)
MAG Glycerol Fatty Acids
Intestinal lumen
Lipoprotein
TAG
Lymphatics
Chylomicrons
Adipose Tissue And Muscle
Blood (bound to albumin)
7
Adipocytes
8
Fat Storage

• Mainly as triacylglycerols (triglycerides) in
  adipose cells
• Constitute 84 of stored energy
• Protein - 15
• Carbohydrate (glucose or glycogen) - lt1

9
Processing of Lipid Reserves Overview

• 1. Lipid Mobilization
• In adipose tissue TAGs hydrolyzed to fatty
  acids plus glycerol
• 2. Transport of Fatty Acids in Blood
• To Tissues
• Activation of Fatty Acids as CoA Ester
• Transport into Mitochondria
• 5. Metabolism to Acetyl CoA

10
Release of Fatty Acids from Triacylglycerols
11
Lipolysis
Hormone (Adrenalin, Glucagon, ACTH)
Receptor (7TM)
Activates
Adenylyl Cyclase
ATP c-AMP
Insulin blocks this step
Activates lipase
Triacylglycerols Glycerol
Fatty acids
Blood
Adipose Cell
12
Adenylyl cyclase Phosphodiesterase
ATP c-AMP AMP
Enhanced by insulin
Enhanced by glucagon
Inactive Kinase Activated Kinase
P
Inactive Lipase Activated Lipase
(Hormone-sensitive Lipase)
Phosphatase
Insulin favors formation of the inactive lipase
Glycerol Fatty Acids
Triacyl- glycerol
13
Acylglycerol Lipases

Diacylglycerol (DAG)
Triacylglycerol Lipase
OH
Diacylglycerol Lipase
Triacylglycerol (TAG)
OH
OH
Monoacylglycerol Lipase
OH
Monoacylglycerol (MAG)
OH
OH
Glycerol
14

Fate of Glycerol
Pyruvate
In Liver
Glycolysis
OH
Dihydroxyacetone Phosphate
OH
Gluconeogenesis
OH

Glycerol
Glucose

15
Beta Oxidation

• Cleavage of fatty acids to acetate in tissues
• Occurs in mitochondria

16
Steps in Beta Oxidation

• Fatty Acid Activation by Esterification with
  CoASH
• Membrane Transport of Fatty Acyl CoA Esters
• Carbon Backbone Reaction Sequence
• Dehydrogenation
• Hydration
• Dehydrogenation
• Carbon-Carbon Cleavage (Thiolase Reaction)

17
Fatty Acid Activation by Esterification with CoASH
AcylCoA
CoASH RCO2H ATP RCOSCoA AMP
PPi
Synthetase
Pyrophos- phatase
Occurs in outer mitochondrial membrane for long
chain fatty acids
2 Pi
?G0(KJ/mole)
ATP AMP PPi
-32.3 CoASH RCO2H RCOSCoA
31.5 PPi 2 Pi
-33.6
-34.4
18
Membrane Transport of Fatty Acyl CoA Esters
Transported across inner mitochondrial membrane
by translocase
19
Translocase
Carnitine acyltransferase I Carnitine
acyltransferase II
Source http//cellbio.utmb.edu/cellbio/mitochondr
ia_1.htm
20
Beta Oxidation Reaction Sequence
Occurs in Mitochondria
(?-ketothiolase)
Repeat Sequence
21
Complete Beta Oxidation of Palmitoyl CoA
7 Cycles
8 CH3COSCoA 7 FADH2 7 NADH 7 H
22
Energetics of Complete Oxidation of Fatty Acids
High Energy Phosphate Bonds Generated
Palmitic Acid Palmitoyl CoA
-2 CH3COSCoA CO2 H2O
108
TCA Cycle
Net 106
?G0 3,233 KJ/Mole
106 High Energy Phosphate Bonds
For Palmitic Acid CO2 ?G0 - 9,790 KJ/Mole
Efficiency of ?-Oxidation 33
23
Complete Oxidation Fatty Acids 9
kcal/g Carbohydrates 4 kcal/g Protein 4
kcal/g
24
American Golden Plover
25
Arctic Tern
26
Camel
27
Beta Oxidation of Odd Carbon Fatty Acids
5 Cycles
5 CH3COSCoA CH3CH2COSCoA
Propionyl CoA Carboxylase ATP/CO2
Propionyl CoA
TCA Cycle
Epimerase
Mutase Vit. B12
Succinyl CoA
L-Methylmalonyl CoA
D-Methylmalonyl CoA
28
Beta Oxidation of Unsaturated Fatty Acids
Oleoyl CoA
Beta Oxidation (3 Cycles)
Isomerase
trans-?2
cis-?3
Continuation of Beta Oxidation
29
Ketogenesis Formation of Ketone Bodies
Thiolase
CH3COSCoA
2 CH3COSCoA CH3COCH2COSCoA
Acetoacetyl CoA
HMG CoA Synthase
Several steps
Cholesterol (in cytosol)
OH
HO2C-CH2-C-CH2COSCoA
See Slide 78
CH3
(in liver mitochon- drial matrix)
?-Hydroxy-?-methylglutaryl CoA (HMG CoA)
Ketogenesis
30
Ketogenesis Formation of Ketone Bodies (Contd.)
OH
HMG CoA lyase
HO2C-CH2-C-CH2COSCoA
CH3COCH2CO2
- CH3COSCoA
Acetoacetate
CH3
HMG CoA
NADH H
- CO2
Dehydrogenase
NAD
OH
CH3COCH3
CH3CHCH2CO2
Acetone (volatile)
?-Hydroxybutyrate
Ketone bodies are important sources of energy,
especially in starvation
31
Ketone Bodies As Energy Sources
In liver
?-Hydroxybutyrate Acetoacetate
Succinyl CoA
Acetoacetate is major energy source in cardiac
muscle and renal cortex also in brain
in starvation and diabetes
?-Ketoacyl CoA transferase
Not found in liver
Thiolase
Succinate
Acetoacetyl CoA
2 Acetyl CoA
Combines with oxaloacetate
TCA Cycle
32
Ketones in Diabetes Mellitus

• In presence of insulin
• Enhanced glucose uptake by tissues
• Decreased mobilization of lipids by
  adipocytes
• In absence of insulin
• Decreased glucose uptake by tissues
• Increased mobilization of lipids by adipocytes

33
Ketones in Diabetes Mellitus

• Biochemical consequences of decreased
• insulin production
• Glucose not taken up by liver
• Decreased oxaloacetate to combine with
• acetyl CoA to enter TCA
• Adipocytes release fatty acids into blood
• Increased production of ketone bodies in
  liver

34
Metabolic Acidosis in Untreated Diabetes Mellitus
CH3COCH2CO2H pKa 3.6 Acetoacetic
Acid
OH
CH3CHCH2CO2H pKa 4.7
?-Hydroxybutyric acid
Concentration of acetoacetic acid can result in
metabolic acidosis (pH 7.1) affinity of
Hb for O2.
35
Fatty Acid Biosynthesis
36
Fatty Acid Synthesis vs. Degradation
Synthesis Degradation
Linked to SH in Linked to
CoASH Proteins (Acyl Carrier Proteins) Cytosol

Mitochondria Components of Separate
Polypeptides Single Peptide NADP / NADPH
NAD / NADH
Intermediates Site Enzymes Redox Coenzymes
37
Fatty Acid Biosynthesis

• Occurs in cytosol
• Starts with acetyl CoA
• Problem
• Most acetyl CoA produced in mitochondria
• Acetyl CoA unable to traverse mitochondrial
  membrane

38
Citrate As Carrier of Acetate Groups
Cytosol Mitochondria
Glucose Pyruvate
Pyruvate Acetyl CoA
Pyruvate Dehydrogenase
Malic enzyme
Malate
Oxalo- acetate
Malate dehydrogenase
Citrate
Acetyl CoA
Oxaloacetate
ATP-Citrate Lyase
Note Acetyl CoA cannot be converted to glucose
Citrate
Mitochondrial membrane
39
Fatty Acid Biosynthesis Formation of Malonyl CoA
Acetyl CoA Carboxylase
Malonyl CoA
CH3COSCoA ATP HCO3-
-O2CCH2COSCoA
ADP Pi H

• Committed step in fatty acid synthesis
• Reaction is irreversible
• Regulation of acetyl CoA carboxylase activity
• by palmitoyl CoA
• by citrate
• by insulin
• by epinephrine and glucagon
• Malonyl CoA inhibits carnitine acyl transferase
  I
• Blocks beta oxidation

40
Fatty Acid BiosynthesisRole of Acyl Carrier
Proteins
Acetyl Transferase
CH3COSCoA CH3CO-S-ACP
Acetyl ACP
Malonyl Transferase
-O2CCH2COSCoA -O2CCH2CO-S-ACP
Malonyl ACP
ACP Acyl carrier protein
41
Fatty Acid BiosynthesisFormation of Acetoacetyl
ACP
CH3CO-S-ACP -O2CCH2CO-S-ACP
?-Ketoacyl ACP Synthetase
CH3COCH2CO-S-ACP CO2
Acetoacetyl ACP
42
Fatty Acid BiosynthesisFormation of Butyryl ACP
?-Ketoacyl ACP reductase
OH
CH3COCH2CO-S-ACP
CH3CCH2CO-S-ACP
H
Acetoacetyl ACP
NADPH H
NADP
?-D-Hydroxybutyryl ACP
?-Hydroxyacyl ACP dehydratase
- H2O
H
NADPH H
NADP
CH3CC-CO-S-ACP
CH3CH2CH2CO-S-ACP
2,3-trans- Enoyl ACP reductase
Crotonyl ACP
Butyryl ACP
H
43
Fatty Acid BiosynthesisSources of NADPH
Pentose Phosphate Pathway
NADPH H
CO2-
NADPH H
CHO
OH
NADP
NADP
OH
OH
O
HO
OH
HO
OH
OH
OH
OH
OP
OH
OP
OP
CO2
Ribulose-5- phosphate
6-Phospho- gluconate
Glucose-6- phosphate
NADPH H
Malic Enzyme
NADP
O
HO-CH-CO2-
CH3CCO2-
Malic Enzyme
Pyruvate
Malate
CH2CO2-
CO2
44
Fatty Acid BiosynthesisChain Elongation
CH3CH2CH2CO-S-ACP
-O2CCH2CO-S-ACP

CH3CH2CH2COCH2CO-S-ACP
H
OH
CH2CH2CH2CHCH2CO-S-ACP
CH3CH2CH2CCCO-S-ACP
H
45
Fatty Acid BiosynthesisChain Elongation (Contd)
H
NADPH H
NADP
CH3(CH2)3CH2CO-S-ACP
CH3CH2CH2CCCO-S-ACP
H
5 Cycles
CH3(CH2)13CH2CO-S-ACP
CH3(CH2)13CH2CO2-
Thioesterase
Palmitoyl ACP
Palmitate
46
Fatty Acid BiosynthesisFatty Acid Synthase in
Animals

• Consists of a single polypeptide containing
• three distinct domains
• Conducts all steps in fatty acid synthesis
• except function of acyl CoA carboxylase

47
Orlistat A Fatty Acid Synthase (FAS) Inhibitor
Anti-obesity (Inhibits pancreatic lipase in
git) Inhibits thioesterase domain of
FAS Anti-cancer (experimental) FAS
overexpressed in several tumor types inhibition
induces apoptosis
48
The Crystal Structure of a Mammalian Fatty Acid
Synthase Timm Maier, Marc Leibundgut, Nenad Ban
Sept. 5, 2008
49
Further Processing of Fatty Acids Elongation
In mitochondria and at surface of endoplasmic
reticulum
CH3(CH2)13CH2COSCoA
Palmitoyl CoA
CH3COSCoA
Thiolase
CH3(CH2)13CH2COCH2COSCoA
NADH H
Dehydrogenase
OH
NAD
L-? Configuration
CH3(CH2)13CH2CCH2COSCoA
H
50
Further Processing of Fatty Acids Elongation
(Contd)
OH
CH3(CH2)13CH2CCH2COSCoA
H
- H2O Hydratase
H
CH3(CH2)13CH2CCCOSCoA
NADPH H
H
Dehydrogenase
NADP
CH3(CH2)13CH2CH2CH2COSCoA
Stearoyl CoA
51
Further Processing of Fatty Acids Unsaturation
CH3(CH2)13CH2CH2CH2COSCoA
Stearoyl CoA
Stearoyl CoA Desaturase
O2
CH3(CH2)7CC(CH2)7COSCoA H2O
Oleoyl CoA
H
H
This reaction occurs in eukaryotes Endoplasmic
reticulum membrane
52
Further Processing of Fatty Acids
Polyunsaturation
9
CH3(CH2)7CC(CH2)7CO2H
Oleic acid
H
H
(181?9)
Plants Further unsaturation occurs primarily in
this region
Animals Further unsaturation occurs primarily in
this region
12 9
Linoleic acid (182?9, 12)
Essential dietary fatty acids in mammals
15 12 9
Linolenic acid (183?9, 12, 15)
53
Formation of Arachidonate in Mammals
Linoleic acid
As CoA ester 1) Elongation 2) Desaturation x 2
14 11 8
5
Arachidonic acid (204?5, 8, 11,
14) (Eicosa-5,-8,11,14-tetraenoic acid)
Prostaglandins
54
Omega-3 Fatty Acids
Eicosapentaenoic acid (205?5, 8, 11, 14, 17)
?-3 double bond
Docahexaenoic acid (226?4, 7, 10, 13, 16, 19)

• Found in fish oils, esp. cold water fish
• Important in
• Growth regulation
• Modulation of inflammation
• Platelet activation
• Lipoprotein metabolism

55
Metabolite Regulation of Fatty Acid Synthesis and
Breakdown
Citrate
Glucose
Stimulates
Blocks
Beta Oxidation
Pyruvate Acetyl CoA
Malonyl CoA Palmitoyl
CoA
Inhibits
56
Hormonal Regulation of Fatty Acid Synthesis and
Breakdown
Phosphodiesterase
Adenylyl cyclase
ATP cAMP
AMP
Stimulates
Stimulates
Insulin
Glucagon and epinephrine
Inactivates lipase
Activates Protein Kinase
Inhibition of fatty acid synthesis
Activates triacyl- glycerollipase
Inactivates ACC by phosphorylation
57
Synthesis of Phosphatidate
Dihydroxyacetone Phosphate (from glycolysis)
Glycerol
Phosphatidate (formed in endoplasmic reticulum)
Diacylglycerol (important in cell signaling)
R3COSCoA
Triacylglycerol (transported to adipocytes
and muscle)
Diacylglycerol acyltransferase (liver)
58
Synthesis of Glycerophospholipids

Transferase
RH CDP ethanolamine RCH3 CDP choline CDP
cytidine diphosphate

Diacylglycerol
R3NH3 Phosphatidylethanolamine R3N(CH3)3
Phosphatidylcholine

Serine
Ethanolamine
Phosphatidylserine
59
Respiratory Distress Syndrome
Most frequently seen in premature infants Also
called hyaline membrane disease Failure to
produce sufficient dipalmitoyl phosphatidylcholine
, which normally is found in the extracellular
fluid surrounding alveoli decreases surface
tension of fluid to prevent lung collapse Treatme
nt in infants born before 30 weeks includes
administration of artificial lung surfactant
(e.g., Exosurf or Pumactant)
60
Synthesis of Glycero-phospholipids (Contd)
Phosphatidyl- inositol
Phosphorylation of 4 5 OH groups
Phosphatidate
Cytidine diphosphate (CDP) diacylglycerol
Phospholipase C (plasma membrane)
Both IP3 and DAG are important second
messengers in cell signaling pathways

Diacylglycerol (DAG)
Inositol-1,4,5- triphosphate (IP3)
61
Synthesis of Glycero-phospholipids (Contd)
Cytidine diphosphate (CDP) diacylglycerol
Cardiolipin formed in inner mitochondrial
membrane plays role in oxidative phosphorylation
62
Synthesis of Glycero-phospholipids (Contd)
Dihydroxyacetone Phosphate (from glycolysis)
Plasmalogens (Abundant in cardiac tissue and CNS)
63
Synthesis of Sphingolipids
Serine
CH3(CH2)14COSCoA
Palmitoyl CoA
3-Ketosphingosine synthase
CoASH
HCO3-2
CH3(CH2)14CO-CHCH2OH
2S,3-Ketosphinganine
NH3
3 Steps
OH
trans
CH3(CH2)12CHCH-CH-CH-CH2OH
Ceramide
CH3(CH2)nCONH
64
Synthesis of Sphingolipids(Contd)
OH
trans
CH3(CH2)12CHCH-CH-CH-CH2OH
CH3(CH2)nCONH
Ceramide
Phosphatidylcholine
Diacylglycerol
OH
O

trans
CH3(CH2)12CHCH-CH-CH-CH2O-P-OCH2CH2N(CH3)3
Sphingomyelin
O-
CH3(CH2)nCONH
Cerebrosides
Gangliosides
65
Synthesis of Gangliosides
OH
trans
CH3(CH2)12CHCH-CH-CH-CH2OH
Ceramide
CH3(CH2)nCONH
OH
Glucose or galactose
trans
CH3(CH2)12CHCH-CH-CH-CH2O-Sugar
CH3(CH2)nCONH
Cerebroside
Ceramide - Sugar - Sugar - GalNAc - Gal
Ganglioside
NAN N-acetylneuraminate GalNAc
N-acetylgalactose
NAN
66
Lipid Storage Diseases(Gangliosidoses)
67
Tay-Sachs Disease
GM2 (a ganglioside)
Ceramide - O - Glucose - Galactose -
N-Acetylgalactose
Hexoseaminidase A catalyzes cleavage of this
glycoside linkage
Autosomal recessive disorder characterized by
deficiency of hexoseaminidase A accumulation of
gangliosides in brain Most prevalent in Jews from
Eastern Europe For further information see
http//www.marchofdimes.com/professionals/681_122
7.asp
68
Other Gangliosidoses
Gauchers disease Fabrys disease Nieman-P
ick disease
Ceramide - O - Glucose
?-glucosidase
Ceramide - O - Glucose - O - Galactose - O -
Galactose
?-galactosidase
Ceramide - Phosphate - Choline
sphingomyelinase
69
Synthesis of Eicosanoids
In cell membrane
R H or CH3
Hydrolysis of sn-2 ester bond by phospholipase A2
(PLA2)
Arachidonate
70
Synthesis of EicosanoidsPLA2 Activation
Various stimuli
Activation of Hormones, autacoids, etc.
Membrane-bound Receptors
PLA2 Activity
Ca2
Arachidonate release and eicosanoid synthesis are
important mediators of tissue injury and
inflammation
71
Synthesis of EicosanoidsProstaglandin Synthesis
Prostaglandin endoperoxide synthetase
(Cyclooxygenase)
Cyclooxygenase
Cyclic endoperoxide
Hydroperoxidase
PGG2
PGH2
Hydroperoxide
Prostaglandin endoperoxide synthetase (also
called cyclooxygenase) possesses both
cyclooxygenase and hydroperoxidase activity Two
forms of cyclooxygenase COX -1 - constitutively
expressed
COX -2 - inducible
72
Cyclooxygenase (COX) Inhibitors
Nonsteroidal antiinflammatory drugs
Ser-530
Acetylsalicylic acid (aspirin)
Irreversible inhibition of COX by acetylation of
the active site
Actions of Aspirin Antiinflammatory (COX-2
inhibition) GI injury (COX-1 inhibition)
73
COX-2 Selective Inhibitors
Rofecoxib (Vioxx)
Celecoxib (Celebrex)
Glucocorticoids block COX-2 expression
74
Prostaglandins
PGE2
PGH2
PGD2
Prostaglandins exhibit a variety of actions on
different tissues
PGF2a
75
Prostacyclin and Thromboxanes
Prostacyclin synthase
Prostacyclin (PGI2) Blocks platelet aggregation
PGH2
Thromboxane synthase
Non-Enzymatic
Thromboxane A2 (TxA2) Promotes platelet
aggregation (t1/2 30 sec.)
Thromboxane B2 (TxB2) inactive
76
Leukotriene Biosynthesis
Leukotrienes are important mediators of
inflammation
5-Lipoxygenase
Arachidonic acid
5-Hydroperoxyeicosa- 6,8,11,14-tetraenoic
acid (5-HPETE)
LTA Hydrolase
5-Lipoxygenase
Leukotriene A4 (LTA4)
Leukotriene B4 (LTB4)
Glutathione LTC4 synthase
- Glu - Gly
Leukotriene C4 (LTC4)
Leukotriene E4 (LTE4)
Cysteinyl leukotrienes
77
Leukotriene Biosynthesis (Contd)
12-Lipoxygenase
12-Hydroperoxyeicosa- 5,8,10,14-tetraenoic
acid (12-HPETE)
Arachidonic acid
12-Hydroxyeicosa- 5,8,10,14-tetraenoic
acid (12-HETE)
78
Leukotriene Biosynthesis Inhibition
Zileuton (Zyflo) An inhibitor of
5-lipoxygenase Used in the treatment of asthma
79
Cholesterol Biosynthesis Formation of Mevalonate
Liver is primary site of cholesterol biosynthesis
Thiolase
CH3COSCoA
2 CH3COSCoA CH3COCH2COSCoA
Acetoacetyl CoA
HMG CoA Synthase
OH
OH
HMGCoA reductase
HO2C-CH2-C-CH2COSCoA
HO2C-CH2-C-CH2CH2OH
CH3
CH3
CoASH
?-Hydroxy-?-methyl- glutaryl CoA (HMG CoA)
3R-Mevalonic acid
Key control step in cholesterol biosynthesis
80
Cholesterol Biosynthesis Processing of Mevalonate
OH
OH
-O2C-CH2-C-CH2CH2OH
-O2C-CH2-C-CH2CH2OPOP
2 Steps
CH3
CH3
ATP
Mevalonate
5-Pyrophospho- mevalonate
- CO2 - H2O
CH3
Isomerase
CH2C-CH2CH2OPOP
CH3-CCH2CH2OPOP
Isopentenyl pyrophosphate
Dimethylallyl pyrophosphate
CH3
81
Cholesterol BiosynthesisIsoprenoid Condensation
Tail
Dimethylallyl pyrophosphate
Head to tail Condensation
Head
Tail
Head
Tail
Geranyl transferase
Isopentenyl Pyrophosphate (IPP)
Geranyl Pyrophosphate (GPP)
Head
Isoprenes
Head to tail condensation of IPP and GPP
Geranyl transferase
Tail to tail condensation of 2 FPPs
Squalene synthase
Squalene
Head
Tail
Farnesyl Pyrophosphate (FPP)
82
Isoprenoids

• Widely distributed in nature
• Generally contain multiple of 5 carbons
• Monoterpene 10 carbons
• Sesquiterpene 15 carbons
• Diterpene 20 carbons

Menthol a monoterpene
Lycopene a tetraterpene
83
Conversion of Squalene to Cholesterol
Squalene- 2,3-epoxide
Squalene monooxygenase
O2
2,3-Oxidosqualene cyclase
Squalene
20 Steps
Lanosterol
Cholesterol
Acyl-CoA cholesterol acyltransferase
Cholesterol esters (principal transport form in
blood)
84
Inhibition of Cholesterol Biosynthesis
HMGCoA reductase
HMG CoA
Mevalonate
Intermediate
Atorvastatin (Lipitor) resembles intermediate
85
Transformations of Cholesterol Bile Salts
Cholesterol
Cholic acid
R CH2SO3- Taurocholate R CO2- Glycocholate
Detergents
86
Transformations of Cholesterol Steroid Hormones
Cortisol
Cholesterol
Progesterone
Testosterone
Estradiol
Vitamin D