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Chapter 5. Metabolism of Lipids. Lipids Insoluble or immiscible Triacylgerols store and supply energy for metabolism . Lipoids: phospholids, glycolipids, cholesterol and cholesterol ester membrane components. Metabolism of lipid. Fatty acids esterified to some backbone molecules

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chapter 5 metabolism of lipids
Chapter 5.Metabolism of Lipids

Lipids

Insoluble or immiscible

Triacylgerols

store and supply energy for metabolism.

Lipoids: phospholids, glycolipids, cholesterol and cholesterol ester

membrane components

metabolism of lipid
Metabolism of lipid
  • Fatty acids

esterified to some backbone molecules

glycerol sphingosine

cholesterol

metabolism of lipids
Metabolism of Lipids

Fats

store in adipose tissue

Essential fatty acids: formation of membrane, regulation of chollesterol metabolism, precursors of eicosanoids (protaglandins, thromboxanes and leukotrienes.

Necessary unsaturated fatty acids

fat facts
Fat Facts
  • Dietary lipids are 90% triacylglycerols; also include cholesterol esters, phospholipids, essential unsaturated fatty acids; fat soluble vitamins (A,D,E,K)
  • Fat is energy rich and provides 9 kcal/gm
  • Normally essentially all (98%) of the fat consumed is absorbed, and most is transported to adipose for storage.
slide5

SIX STEPS OF LIPID DIGESTION AND ABSORPTION

  • Minor digestion of triacylglycerols in mouth and stomach by lingual (acid- stable) lipase
  • Major digestion of all lipids in the lumen of the duodenum/jejunum by pancreatic lipolytic enzymes
  • Bile acid facilitated formation of mixed micelles that present the lipolytic products to the mucosal surface, followed later by enterohepatic bile acid recycling
  • Passive absorption of the lipolytic products from the mixed micelle into the intestinal epithelial cell
  • Reesterificationof 2-monoacylglycerol, lysolecithin, and cholesterol with free fatty acids inside the intestinal enterocyte
  • Assembly and exportfrom intestinal cells to the lymphatics of chylomicrons coated with Apo B48 and containing triacylglycerols, cholesterol esters and phospholipids
slide9

LIPOLYSIS

  • Mobilization of fats from triacylglycerols
  • Hormone sensitive lipase
    • Rate-determining step
    • Specific for removing first fatty acid
    • Phosphorylated form is active
slide10

Fatty acid +

Triacylglycerol

Diacylglycerol

ATP

HSL-a

OP

ADP

Insulin

+

protein

cyclic

AMP

phosphatase

+

ATP

HSL-b

Pi

OH

(inactive form)

AMP

caffeine

theophylline

-

cell

membrane

HORMONES

Epinephrine

Glucagon

Adenylyl

cyclase

active

protein

kinase A

RECEPTORS

inactive

phosphodiesterase

+

= activation

-

= inhibition

HSL = hormone-sensitive lipase

Figure 1. Hormonal activation of triacylglycerol (hormone-sensitive) lipase. Phosphorylation brings about activation to HSL-a.

lipolysis
lipolysis

Glycerols and fatty acids

diffuse out of adipose cells and enter into circulation

Free fatty acids (FFA)

form fatty acid-albumin complexes

Glycerols

to form dihydroxyacetone phosphate (DHAP)

Figure. Page 176

beta oxidation part i
Beta Oxidation Part I

The break down of a fatty acid to acetyl-CoA

units…the ‘glycolysis’ of fatty acids

STRICTLY AEROBIC

Occurs in the mitochondria

Acetyl-CoA is fed directly into the Krebs cycle

Overproduction causes KETOSIS

Exemplifies Aerobic Metabolism

at its most powerful phase

slide14

ATP

PPi

HS-CoA

AMP

CH3CH2CH2COOH

[CH3CH2CH2CO-AMP]

Acyl-CoA

synthetase

CH3CH2CH2CO~SCoA

Fatty acyl CoA

Prepares a Fatty Acid for transport and metabolism

slide16

Knoop’s Experiment

Phenylacetate

Benzoate

slide18

THE ENERGY STORY

Glucose

C6H12O6 + 6O2  6CO2 + 6H2O Ho = -2,813 kJ/mol

= - 672 Cal/mol = 3.74 Cal/gram

Stearic Acid

C18H36O2 + 26O2 18CO2 + 18 H2O Ho = -11,441 kJ/mol

= - 2,737 Cal/mol

= 9.64 Cal/gram

On a per mole basis a typical fatty acid is

4 times more energy rich that a typical hexose

slide19

Sample calculation of energy produced for the cell via b-oxidation of palmitate (a C16 fatty acid):

slide20

Palmitoyl-CoA

Palmitoyl-CoA + 7CoA + 7FAD + 7NAD+ + 7H2O

8 Acetyl-CoA 80 ATP

7 FADH2 10.5 ATP

7 NADH + 7H+ 17.5 ATP

108 ATP

-2 ATP

Total 106 ATP

slide21

Beta Oxidation Part II

3 Obstacles

Unsaturated fatty acid

Obstacle of cis double bonds

Polyunsaturated fatty acid

Obstacle of position of double bond

Odd number chain fatty acid

Obstacle of 3 carbons at the end

slide22

H

H

C=C

CH3CH2CH2CH2CH2C CH2CH2CH2CH2CH2CH2CH2CO~SCoA

Whoops!

4

2

3

4

1

1

2

3

5

H

H

H

H

C=C

C=C

CH3CH2CH2CH2 CH2 CH2CH2CH2CH2CH2CH2CH2CO~SCoA

Oleic Acid

C18:cis9

A cis D.B. will interfere

Linoleic

slide23

New  carbon

Cleavage here

9

H

H

C=C

CH3CH2CH2

CH2CH2CH2-CO~SCoA

New COO group

9

H

8

7

CH2C

CH3CH2CH2

C-CO~SCoA

H

Unsaturated and Polyunsaturated Require

Additional Enzymes

8

7

Enoyl CoA

Isomerase

Trans double bond

slide24

H

H

H

H 9

C=C

C=C

CH2-CH2

CH2

CH2-CH2-CH2-CH2-CH2-CH2CH2C~SCoA

O

CH3C~SCoA

CH3C~SCoA

CH3C~SCoA

CH2C~SCoA

CH2C~SCoA

CH2C~SCoA

O

O

O

O

O

O

6

5

4

3

2

1

4

1

3

2

Linoleic Acid C18 cis9,12

slide25

9

-CH2 CH2 CH2CO~SCoA

-CH2 CH2

-CH2 CH2 C-CO~SCoA

H

H

H

H

H

H

H

H

H

H

C=C

C=C

CH2CO~SCoA

C=C

C=C

C-C

H

Round 4

starter

Round 5

starter

Beta carbon to be

Poly Unsaturated (Continued)

Enoyl-CoA

isomerase

slide26

beta 6

-CH2 CH2

H

FAD

H

H

H

H

H

H

C=C

C=C

C-CO~SCoA

C=C

CH2CO~SCoA

H

FADH2

C

Round 5

starter

CO~SCoA

CH2

Dead end

Acyl-CoA

dehydrogenase

New Strategy

slide27

H

beta 6

H

CH2

C

CH2CO~SCoA

NADPH + H+

C

H

H

NADP+

H

C=C

C-CO~SCoA

H

beta 6

beta 6

H

-CH2

CH2

C-CO~SCoA

C

C

H

Reduce near (bond), Shift far (bond)

2,4 dienoyl-CoA

reductase

3,2 enoyl-CoA

isomerase

Continue Beta Oxidation

what is ketosis

CH3CCH2COO-

H

CH3CCH2COO-

O

O

OH

CH3-C-CH3

What is Ketosis?

An excessive production of ketones in the blood

3 derivatives of acetyl-CoA

Acetoacetate

-hydroxybutyrate

Acetone

what is the significance of ketosis
What is the Significance of ketosis

Acidosis

Excessive acid in the blood

Overflow

Excessive oxidation of fatty acids

Metabolic Problem

Faulty Carbohydrate Metabolism

metabolic fate of acetyl coa
Metabolic fate of Acetyl CoA

Pyruvate

minor

Acetyl-CoA

Fatty Acids

Ketone Bodies

major

Citrate

slide32

CH3C~SCoA

CH3C~SCoA

CH2C~SCoA

O

O

O

O

O

CH3CCH2C~SCoA

HS-CoA

CH3C

+

-Ketothiolase

rearrangement

OH

Acetoacetyl-CoA

slide33

HS-CoA

CH3CCH2C~SCoA

HO

OH

O

O

O

O

O

O

OOC-CH2-C-CH2-C~SCoA

CH3CCH2C~SCoA

CH2C-O-

CH3C~SCoA

CH3

HMG-CoA

Synthase

-hydroxy--methyl

glutaryl-CoA

(HMG-CoA)

slide34

CH3-C~SCoA

OH

OOC-CH2-C-CH2-C~SCoA

Acetoacetate

CH3

OH

O

O

O

O

O

OOC-CH2-C-CH3

NADH + H+

OOC-CH2-C-CH2-C~SCoA

CO2

NAD+

CH3

CH3-C-CH3

OOC-CH2-CH-CH3

OH

HMG-CoA

HMG-CoA

Lyase

+

Acetone

-hydroxybutyrate

utilization of ketone bodies
Utilization of ketone bodies

Acetoacetate/succinyl-CoA CoA transferase

Acetoacetyl-CoA thiokinase

Acetoacetyl-CoA thiolase

Page 180

pysiological significance of ketogenesis
Pysiological Significance of ketogenesis

Ketone bodies produced by the liver are excellent fuels for a variety of extrahepatic tissues, especially during times of prolonged starvation.

Reconversion of ketone bodies to acetyl-CoA inside the mitochondria provides metabolic energy.

regulation of ketogenesis
Regulation of Ketogenesis

Feeding status

In the hungry state, higher glucagon and other lipolytic hormones trigger the lipolytic process in adipose tissue with the result that free fatty acids pass into the plasma for uptake by liver and other tissues. This promotes fatty acid oxidation and ketogenesis in the liver.

regulation of ketogenesis1
Regulation of Ketogenesis

Metabolism of glycogen in the hepatic cells

once fats enter the liver, they have two distinct fates: activated to acyl-Co-A and oxidized, or esterified to glycerol in the production of triacylglycerols in cytoplasm. If the liver has sufficient supplies of glycerol-3 phosphate by glucose metabolism, most of the fats will be turned to the production of triacylglycerols. In contrast, glucose deficiency will cause a lower triacylglycerols and ATP generation, with the majority of the FAs entering beta-oxidation leading to a increased production of ketone bodies.

regulation of ketogenesis2
Regulation of Ketogenesis

The fall in malonyl-CoA concentration can terminate the inhibition on carnitine acyltransferase I, such that long-chain fatty acids can be transported through the inner mitochondrial membrane to the enzymes of fatty acid oxidation and ketogenesis. This may happen during a hungry state. In contrast, administration of food after a fast, or of insulin to the diabetic subject, reduces plasma free fatty acid concentrations and increases liver concentration of malonyl-CoA, this will inhibit carnitine acyltransferase I and thus reverses the ketogenic process.

fatty acid biosynthesis
Fatty Acid Biosynthesis

Not exactly the reverse of degradation

by a different set of enzymes , in a different part of the cell

Primarily in the cytoplasm of the following tissues: liver, kidney, adipose, central nervous system and lactating mammary gland

Liver is the major organ for fatty acid synthesis

lipid biosynthesis
LIPID BIOSYNTHESIS
  • Fatty acid biosynthesis-basic fundamentals
  • Fatty acid biosynthesis-elongation and desaturation
  • Triacylglycerols
  • Phospholipids
  • Cholesterol
  • Cholesterol metabolism
fatty acid biosynthesis1
Cytosol

Requires NADPH

Acyl carrier protein

D-isomer

CO2 activation

Keto  saturated

Mitochondria

NADH, FADH2

CoA

L-isomer

No CO2

Saturated  keto

Fatty Acid Biosynthesis

Synthesis

Beta Oxidation

slide43

Rule:

Fatty acid biosynthesis is a stepwise assembly

of acetyl-CoA units (mostly as malonyl-CoA)

ending with palmitate (C16 saturated)

3 Phases

Activation

Elongation

Termination

slide44

Cofactor

CH3C~SCoA

O

O

ATP

HCO3-

ADP + Pi

CO2

-OOC-CH2C~SCoA

active carbon

Biocytin

ACTIVATION

Biotin

Acetyl-CoA carboxylase

Carboxybiocytin

acetyl coa carboxylase the rate controlling enzyme of fa synthesis
Acetyl-CoA CarboxylaseThe rate-controlling enzyme of FA synthesis
  • In Bacteria -3 proteins (1) Carrier protein with Biotin (2) Biotin carboxylase (3) Transcarboxylase
  • In Eukaryotes - 1 protein (1) Single protein, 2 identical polypeptide chains
  • (2) Each chain Mwt = 230,000 (230 kDa) (3) Dimer inactive (4) Activated by citrate which forms filamentous form of protein that can be seen in the electron microscope
slide46

Yeast Fatty Acid Synthase Complex

2,500 kDa Multienzyme Complex

6 molecules of 2 peptide chains called A and B

(66)

A: (185,000)

Acyl Carrier protein

-ketoacyl-ACP synthase (condensing enzyme)

-ketoacyl-ACP reductase

B: (175,000)

-hydroxy-ACP dehydrase

enoyl-ACP reductase

palmitoyl thioesterase

Fatty Acid

Synthase

Complex

slide47

H

CH3

H

HO

O

ACP

HS-CH2-CH2-N-C-CH2-CH2-N-C-C-C-CH2-O-P-O-CH2-Ser-

O

O

O

H

H

H

CH3

H

HO

O

O

HS-CH2-CH2-N-C-CH2-CH2-N-C-C-C-CH2-O-P-O-P-O-CH2

Adenine

O

O

O

O

O

H

H

H

O

OH

O-P-O

OH

Acyl Carrier Protein

Phosphopantetheine

Cysteamine

Acyl carrier protein

10 kDa

Coenzyme A

slide48

Initiation

CH3C~SCoA

CH3C-

ACP

ACP

+ HS-CoA

O

O

O

O

-OOC-CH2C~S-

CH2C~S-

Overall Reaction

Malonyl-CoA + ACP

Acyl Carrier

Protein

CO2

HS-CoA

NOTE:

Malonyl-CoA carbons become new COOH end

Nascent chain remains tethered to ACP

CO2, HS-CoA are released at each condensation

slide49

-Carbon

CH3C-

ACP

ACP

ACP

O

O

O

O

O

H

D isomer

CH3C-

HO

H

CH2C~S-

= C- C~S-

CH2C~S-

CH3C-

H

CH3CH2CH2C~S-

ACP

Elongation

Reduction

NADPH

-Ketoacyl-ACP reductase

Dehydration

-H2O

 -Hydroxyacyl-ACP dehydrase

NADPH

Reduction

Enoyl-ACP reductase

slide50

O

Free to bind

Malonyl-CoA

-CH2CH2CH2C~S-

ACP

TERMINATION

Ketoacyl ACP

Synthase

-KS

Transfer to Malonyl-CoA

Transfer to KS

-S-ACP

Split out CO2

CO2

When C16 stage is reached, instead of transferring to KS,

the transfer is to H2O and the fatty acid is released

slide51

O

O

O

CH3-CH -CH2-C-S

CH3-CH2-CH2-C-S

S-C-CH2-CH2-CH3

Acetyl-CoA

-SH

-SH

S

HS

CoA-SH

C=O

KS

KS

KS

KS

KS

CH2

NADP+

O

C=O

NADPH

H+

CH3-CH=CH-C-S

CH3

O

O

OH

-C-CH3

SH

Malonyl-CoA

S-C-CH3

ACP

O

CoA-SH

-C-CH2-COO-

S

NADP+

NADPH

H+

S

Fatty Acid Synthase

-Ketoacyl

-ACP synthase

Acetyl-CoA-

ACP transacylase

Initiation or

priming

Enoyl-ACP

reductase

-Hydroxyacyl-ACP

dehydrase

H2O

Malonyl-CoA-

ACP transacylase

-Keto-ACP

synthase (condensing enzyme)

CO2

-Ketoacyl

-ACP reductase

Elongation

overall reactions

Acetyl-CoA + 7 malonyl-CoA + 14NADPH + 14H+

Palmitate + 7CO2 + 14NADP+ + 8 HSCoA + 6H2O

7 Acetyl-CoA + 7CO2 + 7ATP

7 malonyl-CoA +7ADP + 7Pi + 7H+

8 Acetyl-CoA + 14NADPH + 7H+ + 7ATP

Palmitate + 14NADP+ + 8 HSCoA + 6H2O + 7ADP + 7Pi

Overall Reactions

7H+

slide53

acetyl-CoA

PROBLEM:

Fatty acid biosynthesis takes place in the

cytosol. Acetyl-CoA is mainly in the

Mitochondria

How is acetyl-CoA made available to the cytosolic

fatty acyl synthase?

SOLUTION:

Acetyl-CoA is delivered to cytosol from the

mitochondria as CITRATE

slide54

HS-CoA

COO

COO

CH2

CH2

HO-C-COO

HO-C-COO

COO

C=O

CH2

CH2

COO

COO

CH2

COO

Acetyl-CoA

COO

CO2

HO-C-H

CH2

CO2

COO

COO

C=O

NADP+

CH3

NADPH + H+

Acetyl-CoA

mitochondria

Citrate lyase

OAA

Malate

dehydrogenase

NADH

L-malate

L-malate

Malic enzyme

OAA

Cytosol

Pyr

Pyruvate

post synthesis modifications
Post-Synthesis Modifications
  • C16 satd fatty acid (Palmitate) is the product
  • Elongation
  • Unsaturation
  • Incorporation into triacylglycerols
  • Incorporation into acylglycerol phosphates
slide56

HS-CoA

R-CH2CH2CH2C~SCoA

OOC-CH2C~SCoA

O

O

CO2

CH3C~SCoA

O

3

2

1

R-CH2CH2CH2CCH2C~SCoA

O

O

NADPH

NADH

- H2O

NADPH

R-CH2CH2CH2CH2CH2C~SCoA

O

Elongation of Chain (two systems)

Malonyl-CoA*

(cytosol)

Acetyl-CoA

(mitochondria)

Elongation systems are

found in smooth ER and

mitochondria

slide57

Desaturation

Rules:

The fatty acid desaturation system is

in the smooth membranes of the endoplasmic

reticulum

There are 4 fatty acyl desaturase enzymes in

mammals designated 9 , 6, 5, and 4 fatty

acyl-CoA desaturase

Mammals cannot incorporate a double bond

beyond 9; plants can.

Mammals can synthesize long chain unsaturated

fatty acids using desaturation and elongation

triacylglycerol synthesis

O

O

O

R-C-O

O-C-R

O-C-R

Triacylglycerol Synthesis
  • Fatty acyl-CoA
  • DHAP reduction to glycerol-PO4 or
  • Glycerol kinase to glycerol-PO4
  • Two esterifications
  • Diacylglycerol-PO4 intermediate
  • Triacylglycerol
slide59

glycolysis

NADH + NAD+

ADP ATP

CH2OH

C=O

CH2OP

CH2OH

CH2OH

O

O

Not in adipose

tissue

O

O

O

2

HO-C-H

HO-C-H

R-C~CoA

R-C~CoA

CH2O-C-R

CH2O-C-R

CH2O-C-R

O

O

O

CH2OH

CH2OP

R-C-O-C-H

R-C-O-C-H

R-C-O-C-H

CH2OH

CH2OP

CH2O-C-R

H2O

PO4

O

Triacylglycerol Biosynthesis

glycerol kinase

glycerol-PO4

dehydrogenase

DHAP

Glycerol-PO4

Phosphatidic

acid

Phospholipid

biosynthesis

1,2 Diacylglycerol

(DAG)

question
Question
  • Can a triacylglycerol (triglyceride) storage fat be synthesized entirely from glucose, i.e., every carbon in the fat comes from a sugar?
  • Answer: YES
metabolism of phospholipids
Metabolism of Phospholipids

Phospholipid

phosphorous-containing lipids

fatty acids, a phosphate group, and a simple organic molecule

Glycerolphospholipids (phosphoglycerides)

glycerol

Sphingolipid

sphingosine

classification of structural features of glycerolphospholipids table 8 2
Classification of structural features of glycerolphospholipidsTable 8-2
  • Phospholipids

hydrophilic head , hydrophobic tail

  • Membrane

phospholipid bilayer

slide64

Phosphatidic

Acid

O

- or +

- or +

O

O

O

Ester linkage

O

O

CH2O-C-R

CH2O-C-R

O

O

R-C-O-C-H

R-C-O-C-H

CH2OPO-CH2CH2-N(CH3)3

CH2OP

O

O

Phospholipid

Biosynthesis

(smooth ER)

Polar component

= choline, serine, ethanolamine, etc

Glycerophospholipids

+

Phosphatidylcholine or lecithin

strategy of glycerophospholipid biosynthesis

PPP-

Ribose

CTP

Strategy of Glycerophospholipid Biosynthesis
  • Activate diacylglycerol
  • Activate appending moiety (salvage)
slide66

1

2

3

O

O

CH2O-C-R

CH2O-C-R

O

O

R-C-O-C-H

R-C-O-C-H

CH2OP

CH2OH

Pi

Eukaryotes

ATP

Glycerol

Glycerol-3-PO4

DHAP

FA-CoA

1-Acyl-DHAP

Phosphatidic acid

NADPH

DAG

1-Acyl-glycerol-3-PO4

ATP

CTP

CDP-diacylglycerol

ethanolamine (CDP-ethanolamine)

choline (CDP-choline)

Serine (phosphatidylethanolamine)

Glycerol (CDP-diacylglycerol)

Inositol (CDP-diacylglycerol)

1,2 DAG

Cardiolipin (phosphatidylglycerol)

slide67

N

H

2

N

O

O

O

N

+

H

O

P

N

C

H

C

H

O

P

O

C

H

2

2

2

3

O

O

O

H

O

ethanolamine

O

H

C

y

t

i

d

i

n

e

d

i

p

h

o

s

p

h

a

t

e

(

C

D

P

)

regulation of triacylglecerol metabolism
Regulation of Triacylglecerol Metabolism

Pancreas

primary organ involved in sensing the organism’s dietary and energetic states.

monitoring glucose concentrations in the blood.

Low blood glucose stimulates the secretion of glucagon

Elevated blood glucose calls for the secretion of insulin

acetaly coa carboxylase acc
Acetaly-CoA carboxylase (ACC)

Committed enzyme in fatty acid synthesis

activated by citrate

inhibited by palmitoyl-CoA, long-chain fatty acyl-CoAs

Affected by phosphorylation

glucagon or epinephrine

decreased activity of ACC by phosphorylation

insulin

increases the synthesis of triacylglycerols

slide71

EICOSANOID FACTS

  • 20-carbon compounds
  • Include prostaglandins, prostacyclins, thromboxanes, leukotrienes
  • Physiological effects at very low concentrations
  • Many of their effects mediated by cyclic AMP or calcium second messengers
  • Unlike hormones, not transported in the blood
  • Local mediators that act where synthesized or in adjacent cells
slide72

The Actions of Prostaglandins and Leukotrienes

  • the inflammatory response involving primarily the joints (rheumatoid arthritis) and skin (psoriasis);
  • b. the production of pain and fever;
  • c. the regulation of blood pressure (vaso-constrictors/dilators) and blood clotting (platelet function);
  • d. decreased gastric acid secretion (prostacyclins may be an ideal way to control the symptoms of peptic ulcer, but prostanoid synthesis inhibitors, like aspirin, increase acid secretion causing peptic ulcer);
  • e. the control of several reproductive functions such as the induction of laborand delivery - this has led to the use of PGF2 as a mid-trimester abortifacient drug or as a labor-inducing agent;
  • f. the regulation of the sleep/wakecycle;
  • g. hypersensitivity allergic reactions (a primary action of leukotrienes).
slide73

Dietary linoleic acid

metabolism

Arachidonic acid

esterification

Membrane phospholipids

Cell Activation Events:

mechanical trauma,

cytokines

growth factors

Zyflo

Arachidonic acid

Anti-inflammatory glucocorticoids

Lipooxygenase

(LOX)

Cyclooxygenase

(COX)

Aspirin, Indomethacin, Ibuprofen

NSAIDs

Phospholipase A2 (PLA2)

GC induce lipocortin that inhibits PLA2

Prostaglandins

and thromboxanes

(Cyclic/ring product)

Leukotrienes

(Linear product)

Aspirin inhibits irreversibly

Indomethacin forms a salt bridge in the binding site

Ibuprofen competes for substrate binding

Zyflo competes with AA for binding

Figure1. Liberation of arachidonic acid and its metabolism to prostaglandins/ thromboxanes or to leukotrienes

slide74

LEUKOTRIENE FACTS

leukotriene synthesis inhibited by Zyflo, a lipooxygenase inhibitor

leukotriene action blocked by accolate, a receptor antagonist

peptidoleukotrienes:

leukotrienes with short peptides added

components of slow reacting substances of anaphylaxis (SRS-A)

anaphylaxis violent (potentially fatal) allergic reaction

10,000 times more potent than histamine

SRS-A released from lung following immunological stress

SRS-A contract smooth muscle causing constriction of bronchi

implicated in hypersensitivity reaction – such as insect sting

slide75

aspirin

indomethacin

ibuprofen

O2

Cyclooxygenase

PGG2

2GSH

Hydroperoxidase

GSSG

Arachidonic Acid (6)

derived from membrane phospholipids

X

Prostaglandin

endoperoxide

synthase

PGH2

central intermediate

(Head of pathway)

Figure3. Conversion of arachidonic acid to PGH2

slide76

COOH

O

O

O

O

O

O

COOH

OH

O

C

C

C

C

C

C

CH3

CH3

CH3

CH3

CH3

CH3

O

CH3

CH2

O

C

Ser

Acetylated

Cyclooxygenase

(inactive)

Figure4. Structure and

mechanism of action of aspirin

CH2

OH

Ser

Cyclooxygenase

(active)

slide77

COX-1 VS COX-2 DRUG ACTION

  • Aspirin:
    • works on both isoforms
    • COX-1 effect reduces platelet aggregation (TXA2)
    • COX-2 effect reduces inflammation
    • Side effects due to COX-1 inhibition – stomach irritation
  • Specific COX-2 inhibitors
    • Celebrex/Vioxx
    • Target inflammatory response
    • No COX-1 inhibition to produce aspirin-induced side effects
biosynthesis of cholesterol introduction
Biosynthesis of Cholesterol Introduction
  • Functions of cholesterol.
    • Important cell membrane component.
    • Precursor for 3 biologically active compounds.
      • Bile.
      • Steroid hormones.
      • Vitamin D.
  • Disease implications.
    • Cardiovascular disease.
      • Diet control and synthesis manipulation = < heart disorders.
biosynthesis of cholesterol introduction1
Biosynthesis of Cholesterol Introduction
  • Disease implications.
    • Gall stones.
    • Steroidogenic enzyme deficiency.
  • Source of cholesterol.
    • Meat.
    • Eggs.
    • Dairy products.
    • De novo liver synthesis.
slide86
Hydroxymethylglutaryl-coenzyme A (HMG-CoA) is the precursor for cholesterol synthesis.

HMG-CoA is also an intermediate on the pathway for synthesis of ketone bodies from acetyl-CoA.

The enzymes for ketone body production are located in the mitochondrial matrix.

HMG-CoA destined for cholesterol synthesis is made by equivalent, but different, enzymes in the cytosol.

slide87
HMG-CoAis formed by condensation of acetyl-CoA & acetoacetyl-CoA, catalyzed by HMG-CoA Synthase.
  • HMG-CoA Reductase catalyzes production of mevalonate from HMG-CoA.
slide88
The carboxyl of HMG that is in ester linkage to the CoA thiol is reduced to an aldehyde, and then to an alcohol.

NADPH serves as reductant in the 2-step reaction.

Mevaldehyde is thought to be an active site intermediate, following the first reduction and release of CoA.

slide89
HMG-CoA Reductase is an integral protein of endoplasmic reticulum membranes.

The catalytic domain of this enzyme remains active following cleavage from the transmembrane portion of the enzyme.

The HMG-CoA Reductase reaction, in which mevalonate is formed from HMG-CoA, is rate-limiting for cholesterol synthesis.

This enzyme is highly regulated and the target of pharmaceutical intervention.

slide90
Mevalonate is phosphorylated by 2 sequential Pi transfers from ATP, yielding the pyrophosphate derivative.

ATP-dependent decarboxylation, with dehydration, yields isopentenyl pyrophosphate.

slide91
Isopentenyl pyrophosphate is the first of several compounds in the pathway that are referred to as isoprenoids, by reference to the compound isoprene.
slide92
Isopentenyl Pyrophosphate Isomeraseinter-converts isopentenyl pyrophosphate & dimethylallyl pyrophosphate.

Mechanism: protonation followed by deprotonation.

condensation reactions
Condensation Reactions

Prenyl Transferase catalyzes head-to-tail condensations:

  • Dimethylallyl pyrophosphate & isopentenyl pyrophosphate react to form geranyl pyrophosphate.
  • Condensation with another isopentenyl pyrophosphate yields farnesyl pyrophosphate.
  • Each condensation reaction is thought to involve a reactive carbocation formed as PPiis eliminated.
slide95
Squalene Synthase: Head-to-head condensation of 2 farnesyl pyrophosphate, with reduction by NADPH, yields squalene.
slide96
Squaline epoxidase catalyzes conversion of squalene to 2,3-oxidosqualene.

This mixed function oxidation requires NADPH as reductant & O2 as oxidant. One O atom is incorporated into substrate (as the epoxide) & the other O is reduced to water.

slide97

Squalene Oxidocyclase catalyzes a series of electron shifts, initiated by protonation of the epoxide, resulting in cyclization.

Structural studies of a related bacterial enzyme have confirmed that the substrate binds at the active site in a conformation that permits cyclization with only modest changes in position as the reaction proceeds.

The product is the sterol lanosterol.

slide98
Conversion of lanosterol to cholesterol involves 19 reactions, catalyzed by enzymes in ER membranes.

Additional modifications yield the various steroid hormones or vitamin D.

Many of the reactions involved in converting lanosterol to cholesterol and other steroids are catalyzed by members of the cytochrome P450 enzyme superfamily.

slide99
Regulation of cholesterol synthesis

HMG-CoA Reductase, the rate-limiting step on the pathway for synthesis of cholesterol, is a major control point.

Short-term regulation:

HMG-CoA Reductase is inhibited by phosphorylation,catalyzed by AMP-Dependent Protein Kinase (which also regulates fatty acid synthesis and catabolism).

This kinase is active when cellular AMP is high, corresponding to when ATP is low.

Thus, when cellular ATP is low, energy is not expended in synthesizing cholesterol.

slide100
Long-term regulationis by varied formation and degradation of HMG-CoA Reductase and other enzymes of the pathway for synthesis of cholesterol.
  • Regulatedproteolysis of HMG-CoA Reductase:
    • Degradation of HMG-CoA Reductaseis stimulated by cholesterol, oxidized derivatives of cholesterol, mevalonate, & farnesol (dephosphorylated farnesyl pyrophosphate).
    • HMG-CoA Reductase includes a transmembrane sterol-sensing domain that has a role in activating degradation of the enzyme via the proteasome (proteasome to be discussed later).
slide101
Long-term regulationis by varied formation and degradation of HMG-CoA Reductase and other enzymes of the pathway for synthesis of cholesterol.
  • Regulatedproteolysis of HMG-CoA Reductase:
    • Degradation of HMG-CoA Reductaseis stimulated by cholesterol, oxidized derivatives of cholesterol, mevalonate, & farnesol (dephosphorylated farnesyl pyrophosphate).
    • HMG-CoA Reductase includes a transmembrane sterol-sensing domain that has a role in activating degradation of the enzyme via the proteasome (proteasome to be discussed later).
slide104

Lipid transport

  • triacylglycerides, cholesterol, phospholipids
  • dietary lipid transport –chylomicron
  • endogenous lipid transport (VLDL, IDL, LDL, HDL)
slide107

Dietary uptake and distribution of fatty acids

pancreatic

lipases

intestinal lumen

triacylglycerols

FFA + monoacylglycedrols

bile acids

cholesterol

absorbed by intestinal

epithelial cells and

reconverted to

triacylglycerols

epithelial cells

triacylglycerols

micelles

micelles

  • Packaged into chylomicron
  • Released into lymphatic system and then via
  • capillaries to blood stream

chylomicron

  • acted upon by lipases on cell walls of
  • capillaries in tissues

energy production

FFA

  • taken up by
  • tissues

reconversion to TAGs

in adipocytes for storage

hormone sensitive lipases

FFA

released to circulatory system

and combine with albumin for

delivery to tissues

why do we need lipoproteins
Why do we need lipoproteins?
  • Triacylglycerides (TAGs) + cholesterol (Chol) are nonpolar molecules → insoluble in H2O
  • TAG + Chol must be packaged within a polar shell in order to be transported through the blood to the various tissues
  • This is accomplished by combining nonpolar lipids w/ amphipathic lipids → (a polar water-soluble terminal group attached to an H2O -insoluble hydrocarbon chain)
lipoproteins apolipoproteins
Lipoproteins & Apolipoproteins

Lipoproteins (LP)

 function: transport of cholesterol + esterified lipids in blood

 structure:

1) polar shell ---single phospholipid (PL) layer: head groups directed outward -Chol -apolipoproteins

2) nonpolar lipid core

-hydrophobic TAG(triacylglycerol) -cholesteryl ester (CE)

apolipoproteins
apolipoproteins
  • Provide structural stability to Lp
  • Act as cofactors for enzymes involved in plasma lipid and Lp metabolism
  • Serve as ligands for interaction w/Lp receptors that help determine disposition of individual particles
slide114

Lipoproteins

  • hydrophobic core (TAGS, cholesterol esters)
  • hydrophilic surface (P-lipids, cholesterol, and
  • apolipoproteins)
  • Function
  • transport of lipids in blood
  • Types of lipoproteins
  • (classified according to density)
  • very low density (VLDL)
  • intermediate density (IDL)
  • low density (LDL)
  • high density (HDL)
  • Protein content increase, lipid decreases as density increases.

8%

85%

% TAGS

Chylomicron VLDL IDL LDL HDL

% Protein

2%

33%

slide117

Chylomicron:

  • 85% TAG, 4% chol., 8% protein
  • formed in intestinal epithelial cells
  • deliver exogenous TAGS to tissue
  • 80 -500nm
  • ApoCII activates lipases in capillary cell
  • walls releasing FFA to tissue
  • chylomicron remnants return to liver where
  • they bind to ApoE receptor and are taken up
  • 1/2 life in blood - 4-5 minutes

Lipoproteins

slide118

VLDL:

  • 50% TAGs, 22% choles., 10% protein
  • 30 -100 nm
  • formed in liver
  • deliver endogenous lipids to other tissues
  • (mainly muscle and fat cells)
  • ApoCII activates lipases in capillary cell
  • walls releasing FFA to tissue
  • converted to IDLs and LDL as lipids are
  • released
slide119

IDL: (31% TAGs, 29% choles., 18% protein)

  • formed from VLDLs as lipids removed
  • some IDLs return to liver
  • rest converted to LDLs by further removal
  • of lipids

Lipoproteins

slide120

LDL: “bad” cholesterol

  • 10% TAGs, 45% choles., 25% protein
  • 25 - 30 nm
  • formed as lipids removed from VLDLs
  • and IDLs.
  • all apolipoproteins lost except ApoB100
  • bind to LDL receptor via ApoB100 and
  • taken up by endocytosis by hepatic and other
  • tissues (50-75% taken up by liver).
  • Primary mode of cholesterol delivery to tissues.
  • Synthesis of LDL receptor is inhibited by
  • high levels of intracellular cholesterol and
  • stimulated by low levels of cholesterol.
  • Therefore, cholesterol uptake is closely
  • matched to intracellular cholesterol levels.
slide121

Lipoproteins

  • HDL: “good” cholesterol
  • 8% TAGs, 30% choles., 33% protein
  • 7.5 - 10 nm
  • formed in liver
  • scavenge cholesterol from cell surfaces
  • and other lipoproteins and deliver it to liver.
  • Convert cholesterol to cholesterol ester
  • bind to “scavenger receptor” on liver cell
  • surface - cholesterol esters taken up and
  • HDLs released and reenter circulation.
slide122

Dietary lipids

Dietary lipids

chylomicron

chylomicron

Liver

Intestine

Triacylglycerols

cholesterol

Cholesterol esters

HDL

VLDLs

HDLs

LDLs

Cholesterol

Cholesterol esters

Triacylglycerols

FFA

monoacylglycerols

Peripheral tissues

slide123

Dietary lipids

Dietary lipids

ApoE/LDLR

mediated

uptake

chylomicron

chylomicron

Chylomicron

remnants

acquire

ApoE, CII

and others

LPLs activated by ApoCII

Distribution of endogenous lipidsThe Exogenous Pathway

Liver

Intestine

Cholesterol

Cholesterol esters

Triacylglycerols

FFA

monoacylglycerols

Peripheral tissues

slide124

acquire

ApoE, CII

and others

IDLs

LDLR/ApoB100

LPLs activated

by ApoCII

Distribution of endogenous lipidsThe Endogenous Pathway

Liver

Triacylglycerols

cholesterol

Cholesterol esters

VLDLs

LDLR/ApoE

LDLs

Cholesterol Ester

Cholesterol

Triacylglycerols

FFA

monoacylglycerols

Peripheral

tissues

slide125

Distribution of endogenous lipidsThe HDL Pathways

Transport of excess cholesterol from peripheral tissues back

to liver for excretion in bile

HDLs act as acceptors for excess chol, Apo, PL derived from

CM, VLDL and LDL

HDLs synthesized by both liver and intestine

slide126

scavenger receptor

uptake of cholesterol

IDLs

CEs

TAGs

HDLs

Distribution of endogenous lipidsThe HDL Pathways

Liver

Triacylglycerols

cholesterol

Cholesterol esters

HDL

VLDLs

LDLs

VLDL

Choles.

Cholesterol Ester

Cholesterol

Triacylglycerols

FFA

monoacylglycerols

Peripheral

tissues

abnormal metabolism of lipoprotein
Abnormal Metabolism of Lipoprotein

Hyperlipoproteinemia

Genetic diseases