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21| Lipid Biosynthesis

© 2013 W. H. Freeman and Company

CHAPTER 21 Lipid Biosynthesis

CHAPTER 21 Lipid Biosynthesis Key topics : – Biosynthesis of fatty acids and eicosanoids – Assembly

Key topics:

– Biosynthesis of fatty acids and eicosanoids

– Assembly of fatty acids and glycerol into triacylglycerols

– Biosynthesis of cholesterol

Lipids fulfill a variety of biological functions

Lipids fulfill a variety of biological functions • Energy storage • Constituents of membranes • Anchors

• Energy storage • Constituents of membranes • Anchors for membrane proteins • Cofactors for enzymes • Signaling molecules • Pigments • Detergents • Transporters • Antioxidants

Catabolism and anabolism of fatty acids proceed via different pathways

Catabolism and anabolism of fatty acids proceed via different pathways • Catabolism of fatty acids –

• Catabolism of fatty acids

produces acetyl-CoA produces reducing power (NADH) takes place in the mitochondria

Anabolism of fatty acids

requires acetyl-CoA and malonyl-CoA requires reducing power from NADPH

takes place in cytosol in animals, (chloroplast in plants)

21.1 Biosynthesis of fatty acids and Eicosanoids

- Fatty acid biosynthesis and breakdown occur by different pathways,

are catalyzed by different sets of enzymes,

and take place in different parts of cell - The biosynthesis requires the participation of a three-carbon intermediate,

malonyl CoA

Malonyl-CoA is formed from acetyl-CoA and bicarbonate

- The formation of malonyl-CoA from acetyl-CoA is an irreversible process catalyzed by acetyl-CoA carboxylase, containing biotin prosthetic group

(Fig21-1)

• 21.1 Biosynthesis of fatty acids and Eicosanoids • - Fatty acid biosynthesis and breakdown occur

Malonyl-CoA is formed from acetyl-CoA and bicarbonate

Malonyl-CoA is formed from acetyl-CoA and bicarbonate • Rx carboxylates acetyl CoA • Catalyzed by acetyl-CoA

• Rx carboxylates acetyl CoA • Catalyzed by acetyl-CoA carboxylase

– Enz has three subunits:

• One unit has Biotin covalently linked to Lys • Biotin carries CO 2

• In animals, all three subunits are on one polypeptide chain

– HCO 3 (bicarbonate) is the source of CO 2

The Acetyl-CoA Carboxylase Reaction

The Acetyl-CoA Carboxylase Reaction Acetyl-CoA carboxylase has three functional regions: biotin carrier protein (gray); biotin carboxylase,

Acetyl-CoA carboxylase has three functional regions:

biotin carrier protein (gray); biotin carboxylase, and transcarboxylase, which transfers activated CO 2

(shaded green) from biotin to acetyl-CoA, producing malonyl-CoA.

The long, flexible biotin arm carries the activated CO 2 from

the biotin carboxylase region to the transcarboxylase active site.

The active enzyme in each step is shaded in blue.

The Acetyl-CoA Carboxylase Reaction Acetyl-CoA carboxylase has three functional regions: biotin carrier protein (gray); biotin carboxylase,

Fatty Acid Synthesis

Fatty Acid Synthesis

• Overall goal: attach two-C acetate unit from malonyl- CoA to a growing chain and then reduce it

• Reaction involves cycles of four enzyme-catalyzed steps

  • - Condensation of the growing chain with activated

acetate

  • - Reduction of carbonyl to hydroxyl

  • - Dehydration of alcohol to trans-alkene

  • - Reduction of alkene to alkane

• The growing chain is initially attached to the enzyme via a thioester linkage • During condensation, the growing chain is transferred to the acyl carrier protein • After the second reduction step, the elongated chain is transferred back to fatty acid synthase

Synthesis of fatty acids is catalyzed y fatty acid synthase (FAS)

Synthesis of fatty acids is catalyzed y fatty acid synthase (FAS) • catalyzes a repeating four-step

• catalyzes a repeating four-step sequence that elongates the fatty acyl chain by two carbons at each step

See Fig. 21-2 – Uses NADPH

as the electron donor

– Uses two enzyme-bound -SH groups

as activating groups

FAS I in vertebrates and fungi FAS II in plants and bacteria

FAS I vs. FAS II

FAS I vs. FAS II FAS I • Single polypeptide chain in vertebrates • Leads to

FAS I

Single polypeptide chain

in

vertebrates

• Leads to single product:

palmitate 16:0

• C-15 and C-16 are from the acetyl CoA used to prime the Rx

FAS II

• Made of separate, diffusible enzymes

• Makes many products (saturated, unsaturated, branched, many lengths, etc.)

Mostly in plants and bacteria

The structure of fatty acid synthase type I systems.

The structure of fatty acid synthase type I systems .

(a)All of the active sites in the mammalian system

are located

in different domains within a single large polypeptide chain.

The 7 different enzymatic activites are:

β-ketoacyl-ACP synthase (KS), malonyl/acetyl-CoA–ACP transferase (MAT), β-hydroxyacyl-ACP dehydratase (DH), enoyl-ACP reductase (ER), and β-ketoacyl-ACP reductase (KR).

ACP is the acyl carrier protein. The linear arrangement of the domains in the polypeptide is shown in the lower panel. The seventh domain (TE) is a thioesterase that releases the palmitate product from ACP when the synthesis is completed.

*The ACP and TE domains are disordered in the crystal and are therefore not shown in the structure.

Enzymes in Fatty Acid Synthase

Enzymes in Fatty Acid Synthase • Condensation with acetate  -ketoacyl-ACP synthase (KS) • Reduction of

• Condensation with acetate

-ketoacyl-ACP synthase (KS)

• Reduction of carbonyl to hydroxyl

-ketoacyl-ACP reductase (KR)

• Dehydration of alcohol to alkene

-hydroxyacyl-ACP dehydratase (DH) • Reduction of alkene to alkane – enoyl-ACP reductase (ER)

• Chain transfer/charging

– Malonyl/acetyl-CoA ACP transferase

Fatty Acid Synthase Type I Systems

Fatty Acid Synthase Type I Systems
Fatty Acid Synthase Type I Systems

Overall Palmitate Synthesis

Overall Palmitate Synthesis
Overall Palmitate Synthesis

The overall process of palmitate synthesis.

The overall process of palmitate synthesis. - The fatty acyl chain grows by two-carbon units donated
  • - The fatty acyl chain grows by two-carbon units donated by activated malonate, with loss of CO 2 at each step.

  • - The initial acetyl group is shaded yellow, C-1 and C-2 of malonate are shaded light red, and the carbon released as CO 2 is shaded green.

  • - After each two-carbon addition, reductions convert the growing chain to a saturated fatty acid of four, then six, then eight carbons, and so on. The final product is palmitate (16:0).

The synthase stop generally at this point. don’t why!!!

Acyl Carrier Protein (ACP) serves as a shuttle in fatty acid synthesis

Acyl Carrier Protein (ACP) serves as a shuttle in fatty acid synthesis • contains a covalently

• contains a covalently attached prosthetic group,

4’-phosphopantetheine

- Flexible arm to tether acyl chain while carrying intermediates

from one enzyme subunit to the next

• Delivers acetate (in the first step) or malonate (in all the next steps) to the fatty acid synthase

Acyl Carrier Protein (ACP) serves as a shuttle in fatty acid synthesis • contains a covalently

Charging ACP and FAS I with acyl groups activates them

Charging ACP and FAS I with acyl groups activates them • Two thiols must be charged

• Two thiols must be charged with the correct acyl groups before condensation rx can begin

Thiol from 4-phosphopantethine in ACP Thiol from Cys in fatty acid synthase

1)Acetyl group of acetyl-CoA is transferred to ACP

– Catalyzed by malonyl/aceyl-CoA transferase (MAT)

– ACP passes this acetate to the Cys of the -ketoacyl-ACP synthase (KS) domain of FAS 1

2) ACP –SH group is re-charged with malonyl from malonyl-CoA

Charging, Activation with ACP, and the Four-Step Sequence of Mammalian Fatty Acid Synthesis

Charging, Activation with ACP, and the Four-Step Sequence of Mammalian Fatty Acid Synthesis

Charging, Activation with ACP, and the Four-Step Sequence of Mammalian Fatty Acid Synthesis

Fatty Acid Synthesis in Detail:

Step 1:Condensation

Fatty Acid Synthesis in Detail: Step 1:Condensation • Activated acetyl and malonyl groups form acetoacetyl-ACP and

• Activated acetyl and malonyl groups form acetoacetyl-ACP and CO 2

– Claisen condensation rx • Catalyzed by -ketoacyl-ACP synthase • Coupling condensation to decarboxylation of malonyl-CoA makes the rx energetically favorable

Fatty Acid Synthesis in Detail:

Step 2: Reduction

Fatty Acid Synthesis in Detail: Step 2: Reduction • Carbonyl at C-3 is reduced to form

• Carbonyl at C-3 is reduced to form D--hydroxybutyryl- ACP

- NADPH is e donor

• Catalyzed by -ketoacyl-ACP reductase (KR)

Fatty Acid Synthesis in Detail:

Step 3: Dehydration

Fatty Acid Synthesis in Detail: Step 3: Dehydration • OH and H removed from C-2 and

• OH and H removed from C-2 and C-3 of -hydroxybutyryl-ACP to form trans-2 -butenoyl-ACP

• Catalyzed by -hydroxyacyl-ACP dehydratase (DH)

Fatty Acid Synthesis in Detail:

Step 4: Reduction (2 nd )

Fatty Acid Synthesis in Detail: Step 4: Reduction (2 ) • NADPH is the electron donor

• NADPH is the electron donor to reduce double bond of trans-2 -butenoyl-ACP

to form butyryl-ACP

• Catalyzed by

enoyl-ACP reductase (ER)

The Transferase and FAS rx’s are repeated in new rounds

The Transferase and FAS rx’s are repeated in new rounds • Product of first round is

• Product of first round is butyryl-ACP – (bound to phosphopantetheine-SH group of ACP) • Butyrul gp is transferred to the Cys of -ketoacyl-ACP synthase – In the first round, acetyl-CoA was bound here • New malonyl-CoA binds to ACP • After new round of four steps, six-C product is made (bound to ACP)

The fatty acid synthase reaction are repeated to form palmitate

• The fatty acid synthase reaction are repeated to form palmitate - Production of the four-carbon

- Production of the four-carbon, saturated fatty-acyl-ACP marks completion of one pass through the fatty acid synthase complex

- To start the next cycle of four reactions that lengthens the chain by two more carbons, another malonyl group is linked to the new unoccupied phosphopantetheins –SH group of ACP

7)

(Fig 21-

  • - Seven cycles of condensation and reduction produce the16-carbon saturated palmitoyl group.

The synthase stop generally at this point.

Beginning of the Second Round of Fatty Acid Synthesis

Beginning of the Second Round of Fatty Acid Synthesis
Beginning of the Second Round of Fatty Acid Synthesis

Stoichiometry of Synthesis of Palmitate (16:0)

Stoichiometry of Synthesis of Palmitate (16:0) 1) 7 acetyl-CoAs are carboxylated to make 7 malonyl-CoAs… using

1) 7 acetyl-CoAs are carboxylated to make 7 malonyl-CoAs… using ATP 7 AcCoA + 7 CO 2 + 7 ATP 7 malCoA + 7 ADP + 7 P i

2) Seven cycles of condensation, reduction, dehydration and reduction…using NADPH to reduce the -keto group and trans-double bond

AcCoA + 7 malCoA + 14 NADPH + 14 H + Palmitate + 7 CO 2 + 8 CoA + 14 NADP + + 6 H 2 O

(eq. 21-3)

Note: Eukaryotes have one additional energy cost. (Next slide)

Compartmentalization of Lipid Metabolism in Animal Cells vs. Plant Cells

Compartmentalization of Lipid Metabolism in Animal Cells vs. Plant Cells Fatty acid synthesis takes place in
Compartmentalization of Lipid Metabolism in Animal Cells vs. Plant Cells Fatty acid synthesis takes place in

Fatty acid synthesis takes place in the compartment in which NADPH is available for reductive synthesis; this is the cytosol in animals and yeast, and the chloroplast in

Acetyl-CoA is transported into the cytosol for fatty acid synthesis

Acetyl-CoA is transported into the cytosol for fatty acid synthesis In nonphotosynthetic eukaryotes… • Acetyl-CoA is

In nonphotosynthetic eukaryotes… Acetyl-CoA is made in the mitochondria But fatty acids are made in the cytosol So, Acetyl-CoA is transported into the cytosol with a cost

of 2 ATPs

•Therefore,

The cost of FA synthesis is 3 ATPs per 2-C

unit

Fatty acid synthesis occurs in cell compartments where NADPH levels are high

Fatty acid synthesis occurs in cell compartments where NADPH levels are high • Cytosol for animals,

Cytosol for animals, yeast Chloroplast for plants

Sources of NADPH:

– In adipocytes:

pentose phosphate pathway and malic enzyme (Fig.

21-9)

• NADPH made as malate converts to pyruvate + CO 2 – In hepatocytes and mammary gland:

pentose phosphate pathway

NADPH made as glucose-6-phosphate converts

to ribulose 6-

phosphate – In plants: photosynthesis

Pathways for NADPH Production

Pathways for NADPH Production
Pathways for NADPH Production

   
 

Acetate is shuttled out of mitochondria as citrate (1)

- All the acetyl CoA used in fatty acid synthesis is formed in mitochondria from pyruvate oxidation and from catabolism of the carbon skeletons of amino acids

- The mitochondria inner membrane

is impermeable to

acetyl-CoA, so, indirect shuttle transfers

acetyl group equivalents across the inner membrane

Acetyl-CoA, generated in the mitochondria, is shuttled to the cytosol as citrate (2)

Acetyl-CoA, generated in the mitochondria, is shuttled to the cytosol as citrate (2) • In most

• In most eukaryotes, the acetyl-CoA for lipid synthesis is made in the mitochondria – But lipid synthesis occurs in the cytosol

• And there is no way for acetyl-CoA to cross mitochondrial inner membrane to the cytosol

• So acetyl-CoA is converted to citrate Acetyl-CoA + oxaloacetate citrate • Same rx as occurs in CAC • Catalyzed by citrate synthase • Citrate passes through citrate transporter

Oxaloacetate cyt is converted to malate (3)

Oxaloacetate is converted to malate (3) • Malate dehydrogenase in cytosol reduces oxaloacetate to malate •

Malate dehydrogenase in cytosol reduces oxaloacetate to malate • Two potential fates for malate:

– can be converted to NADPH cyt and pyruvate cyt via the malic enzyme

• NADPH used for lipid synthesis • Pyruvate cyt sent back to mt via pyruvate transporter • Converted back to oxaloacetate mt by pyruvate carboxylase, requires ATP

– can be transported back to mt via malate --ketoglutarate transporter

Malate mt is reoxidized to oxaloacetate mt

Shuttle for transfer of acetyl groups from mitochondria to the cytosol.

• Shuttle for transfer of acetyl groups from mitochondria to the cytosol. • The outer mitochondrial

The outer mitochondrial membrane

 
 

is freely permeable to all these compounds.

Pyruvate derived from amino acid catabolism in the mitochondrial matrix, or from glucose by glycolysis in the cytosol, is converted to acetyl-CoA in the matrix.

Acetyl groups pass out of the mitochondrion as citrate; in the cytosol they are delivered as acetyl-CoA for fatty acid synthesis.

Oxaloacetate is reduced to malate, which can return to the mitochondrial matrix and is converted to oxaloacetate.

The major fate for cytosolic malate is oxidation by malic enzyme to generate cytosolic NADPH; the pyruvate produced returns to the mitochondrial matrix.

Shuttle for Transfer of Acetyl Groups from Mitochondria to Cytosol

Shuttle for Transfer of Acetyl Groups from Mitochondria to Cytosol
Shuttle for Transfer of Acetyl Groups from Mitochondria to Cytosol

Fatty acid synthesis is tightly regulated via ACC

Fatty acid synthesis is tightly regulated via ACC • Acetyl CoA carboxylase (ACC) catalyzes the rate-limiting

• Acetyl CoA carboxylase (ACC)

catalyzes the rate-limiting step

  • - ACC is feedback-inhibited by palmitoyl-CoA

  • - ACC is activated by citrate

• Remember citrate is made from acetyl-CoA mt

Citrate signals excess energy

fat

to be converted to

- When [acetyl-CoA] mt , converted to citrate…citrate exported to

cytosol

Regulation of Fatty Acid Synthesis in Vertebrates

Regulation of Fatty Acid Synthesis in Vertebrates Both allosteric regulation and hormone-dependent covalent modification influence the

Both allosteric regulation and hormone-dependent covalent modification

influence the flow of precursors into malonyl-CoA.

Regulation of Fatty Acid Synthesis in Vertebrates Both allosteric regulation and hormone-dependent covalent modification influence the

Long-chain saturated fatty acid are synthesized from palmitate

Long-chain saturated fatty acid are synthesized from palmitate - Palmitate , the principal product of the

- Palmitate, the principal product of the fatty acid synthase in animal cell,

is the precursor of other

long-chain fatty acid

(p842)

(Fig 21-12)

Desaturation of a Fatty Acid by Fatty Acyl-CoA desaturase

Desaturation of a Fatty Acid by Fatty Acyl-CoA desaturase . Blue arrows show the path of

. Blue arrows show the path of electrons as two substrates a fatty acyl–CoA and NADPH—undergo oxidation by O 2 .

Desaturation of a Fatty Acid by Fatty Acyl-CoA desaturase . Blue arrows show the path of

fatty acyl desaturase is a non-heme, iron-containing, mixed function oxidase

fatty acyl desaturase is a non-heme, iron-containing, mixed function oxidase • O accepts four electrons from

• O 2 accepts four electrons from two substrates • Two electrons come from saturated fatty acid

• Two electrons

come from ferrous state of

cytochrome b 5

Eicosanoids are formed

from 20-carbon polyunsaturated fatty acid

• Eicosanoids are formed from 20-carbon polyunsaturated fatty acid • - Eicosanoids are a family of

- Eicosanoids are a family of very potent biological signaling molecules that act as short-range messengers, affecting tissue near the cells

that produce them (Fig 21-15, 16)

(p845)

Eicosanoids are potent short-range hormones made from arachidonate

Eicosanoids are potent short-range hormones made from arachidonate • Eicosanoid hormones include prostaglandins, leukotrienes, thromboxanes •

Eicosanoid hormones

include prostaglandins, leukotrienes, thromboxanes

created from the arachidonate 20:4 that’s incorporated into the phospholipids of membranes

In response to stimuli (hormone, etc.), phospholipase A 2 is activated and attacks the

C-2

fatty acid, releasing arachidonate

Conversion of Arachidonate to Prostaglandin s and Other Eicosanoids

Conversion of Arachidonate to Prostaglandin s and Other Eicosanoids After arachidonate is released from phospholipids by

After arachidonate is released from phospholipids by the action of phospholipase A 2 ,

the cyclooxygenase and peroxidase activities of COX (also called prostaglandin H 2 synthase) catalyze the production of PGH 2 , the precursor of other prostaglandins and thromboxanes.

Conversion of Arachidonate to Prostaglandin s and Other Eicosanoids After arachidonate is released from phospholipids by

NSAIDs inhibit cyclooxygenase activity

NSAIDs inhibit cyclooxygenase activity • See Fig. 21-15b • Aspirin (Acetylsalicylate) is an irreversible inhibitor -

• See Fig. 21-15b Aspirin (Acetylsalicylate) is an irreversible inhibitor

  • - acetylates a Ser in active site

  • - blocks active site in both COX isozymes

Ibuprofen and Naproxen are competitive inhibitors

- resemble substrate, also block active site in both isozymes

A Few NSAIDs that Inhibit PGH 2

A Few NSAIDs that Inhibit PGH

Aspirin

- inhibits the first reaction by acetylating an essential Ser residue on the enzyme.

Ibuprofen and naproxen

  • - inhibit the same step, probably by mimicking

the structure of the substrate

or an intermediate in the reaction.

A Few NSAIDs that Inhibit PGH Aspirin - inhibits the first reaction by acetylating an essential

Leukotriene synthesis also begins with arachidonate

Leukotriene synthesis also begins with arachidonate • O is added to arachidonate via lipoxygenases (mixed-function oxidases)

O 2 is added to arachidonate via lipoxygenases (mixed-function oxidases) • creates species that differ in the position of the OOH group • See Fig. 21-16

Synthesis of Leukotrienes

Synthesis of Leukotrienes
Synthesis of Leukotrienes

21.2 Biosynthesis of Triacylglycerol

21.2 Biosynthesis of Triacylglycerol Triacylglycerols and glycerophospholipids are synthesized from the same precursors • Glycogen can

Triacylglycerols and glycerophospholipids are synthesized from the same precursors

Glycogen can be stored only a few hundred grams in liver and muscle, ~12 hr using.

Triacyglycerol can be stored ~20kg, supporting for 12 weeks.

•In animal tissue, triacyglycerol and glycerophospholipid share two precursor (fatty acyl-CoA and L-glycerol 3-phosphate) and several steps.

(Fig 12-17, 18)

Synthesis of Phosphatidic

Synthesis of Phosphatidic Acid - A fatty acyl group is activated by formation of the fatty

Acid

  • - A fatty acyl group is activated by formation of the fatty acyl– CoA, then transferred to ester linkage with L-glycerol 3- phosphate,

  • - Phosphatidic acid is shown here with the correct stereochemistry at C-2 of the glycerol molecule

Synthesis of Phosphatidic Acid - A fatty acyl group is activated by formation of the fatty

Conversion of Phosphatidic Acid into Triacylglycerol

Conversion of Phosphatidic Acid into Triacylglycerol - Phosphatidic acid is the precursor of both triacylglycerols and
  • - Phosphatidic acid

is the precursor of both

triacylglycerols

and

glycerophospholipids.

Conversion of Phosphatidic Acid into Triacylglycerol - Phosphatidic acid is the precursor of both triacylglycerols and

Triacylglycerol biosynthesis in animal

is

Triacylglycerol biosynthesis in animal is regulated by hormone • - Insulin promotes the conversion of carbohydrate

regulated by hormone

• - Insulin promotes the conversion of carbohydrate to triacylglycerol

(Fig 21-19)

Severe diabetes mellitus make people to fail to synthesize fatty acid from carbohydrate or amino acid

• - Glucagon and epinephrine

Regulation of Triacylglycerol Synthesis by Insulin

Regulation of Triacylglycerol Synthesis by Insulin - Insulin stimulates conversion of dietary carbohydrates and proteins to
  • - Insulin

stimulates conversion of dietary carbohydrates and proteins to fat. ( Individuals with diabetes mellitus either lack insulin

or are insensitive to it. )

  • - This results in diminished fatty acid synthesis, and the acetyl- CoA arising from catabolism of carbohydrates and proteins is shunted instead to

ketone body production.

* People in severe ketosis smell of acetone, so the condition is sometimes mistaken for drunkenness

Regulation of Triacylglycerol Synthesis by Insulin - Insulin stimulates conversion of dietary carbohydrates and proteins to

Triacylglycerol breakdown and re-synthesis create a futile cycle

Triacylglycerol breakdown and re-synthesis create a futile cycle • 75% of free fatty acids (FFA) released

75% of free fatty acids (FFA) released by lipolysis are reesterified to form TAGs rather than be used for fuel

– Some recycling occurs in adipose tissue

– Some FFA from adipose cells are transported to liver, remade into TAG, and re-deposited in adipose cells

The triacylglycerol cycle.

• The triacylglycerol cycle .

In mammals, triacylglycerol molecules are broken down and resynthesized in a triacylglycerol cycle during starvation.

Some of the fatty acids released by lipolysis of triacylglycerol in adipose tissue pass into the bloodstream, and the remainder are used for resynthesis of triacylglycerol.

Some of the fatty acids released into the blood are used for energy (in muscle, for example), and some are taken up by the liver and used in triacylglycerol synthesis.

The triacylglycerol formed in the liver is transported in the blood back to adipose tissue, where the fatty acid is released by extracellular lipoprotein lipase, taken up by adipocytes, and reesterified into triacylglycerol.

The Triacylglycerol Cycle

The Triacylglycerol Cycle
The Triacylglycerol Cycle

Benefits of this futile cycle?

Benefits of this futile cycle? • Recycling continues even in starvation • Speculation: – energy reserve

• Recycling continues even in starvation • Speculation:

energy reserve for “fight or flight” crises that might occur during fasting

The total # of FFA in flux may change but the % recycled remains

– unless a pharmacological intervention happens

(i.e., thiazolidinedione drugs)

Adipose tissue generates glycerol 3-phosphate by

Adipose tissue generates glycerol 3-phosphate by glyceroneogenesis (p850) • - glyceroneogenesis • is a shortened version

glyceroneogenesis

(p850)

- glyceroneogenesis

is a shortened version of gluconeogenesis, from pyruvate to DHAP, followed by conversion of the DHAP to glycerol 3-phosphate (Fig 21-21)

   

• - Cortisol and Dexamethasone hormone

- stimulate glyceroneogenesis and gluconeogenesis in the liver,

- suppressing glyceroneogenesis

in the adipose tissue (Fig 21-22)

Glyceroneogen

esis

Glyceroneogen esis The pathway is essentially an abbreviated version of gluconeogenesis , from pyruvate to dihydroxyacetone

The pathway is essentially an abbreviated version of gluconeogenesis, from pyruvate to dihydroxyacetone phosphate (DHAP),

followed by conversion of DHAP to glycerol 3-phosphate, which is used for the synthesis of triacylglycerol.

Glyceroneogen esis The pathway is essentially an abbreviated version of gluconeogenesis , from pyruvate to dihydroxyacetone

What is the source of the glycerol 3-phosphate needed for fatty acid re- esterification?

What is the source of the glycerol 3-phosphate needed for fatty acid re- esterification? • During

During lipolysis (stimulated by glucagon or epinephrine),

glycolysis is inhibited

– So DHAP is not readily available to make glycerol 3-phosphate

And adipose cells don’t have glycerol kinase to make glycerol 3-phosphate on-site

So, cells make DHAP via glyceroneogenesis See next slide

Regulation of Glyceroneogenesis via Glucocorticoid Hormones

Glucocorticoid hormones stimulate

glyceroneogenesis and gluconeogenesis in the liver,

• while suppressing glyceroneogenesis in the adipose tissue (by reciprocal regulation of the gene expressing PEP carboxykinase (PEPCK) in the two tissues);

• this increases the flux through the triacylglycerol cycle. The glycerol freed by the breakdown of triacylglycerol in adipose tissue is released to the blood and transported to the liver,

• where it is

rimaril

converted to

lucose

Regulation of Glyceroneogenesis via Glucocorticoid Hormones

Regulation of Glyceroneogenesis via Glucocorticoid Hormones
Regulation of Glyceroneogenesis via Glucocorticoid Hormones

Regulation of Glyceroneogenesis via Thiazolidinediones

Regulation of Glyceroneogenesis via Thiazolidinediones • A class of drugs called thiazolidinediones are now used to

• A class of drugs called thiazolidinediones are now used to treat type 2 diabetes.

• In this disease, high levels of free fatty acids in the blood interfere with glucose utilization in muscle and promote insulin resistance.

Thiazolidinediones activate a nuclear receptor

called

peroxisome proliferator-activated

receptor γ (PPARγ), which induces the activity of PEP

carboxykinase.

• Therapeutically, thiazolidinediones increase the rate of glyceroneogenesis,

thus increasin

the res

nthesis of triac l

l

cerol

Regulation of Glyceroneogenesis via Thiazolidinediones

Regulation of Glyceroneogenesis via Thiazolidinediones
Regulation of Glyceroneogenesis via Thiazolidinediones

Thiazolidinedione drugs target insulin resistance

Thiazolidinedione drugs target insulin resistance by increasing glyceroneogenesis • Elevated FFA levels seem to promote insulin

by increasing

glyceroneogenesis

Elevated FFA levels

seem to promote insulin resistance

Thiazolidinediones (glitazones) upregulate PEPCK in adipose tissue, lead to glyceroneogenesis, resynthesis of TAG in adipose tissue and release of FFA

Thus the drugs promote sensitivity to insulin

Thiazolidinediones

Thiazolidinediones Have this group in common Avandia (Rosiglitazone) – removed from market due to association with
Thiazolidinediones Have this group in common Avandia (Rosiglitazone) – removed from market due to association with

Have this group in common

Thiazolidinediones Have this group in common Avandia (Rosiglitazone) – removed from market due to association with

Avandia (Rosiglitazone) – removed from market due to association with heart at

Pioglitazone (Actos)
Pioglitazone (Actos)

21.3 Biosynthesis of membrane phospholipids

21.3 Biosynthesis of membrane phospholipids • Two major classes of membrane phospholipids : (p852) glycerophospholipids and

Two major classes of membrane phospholipids:

(p852)

glycerophospholipids and sphingolipids

All the biosynthetic pathways follow a few basic patterns.

(1) synthesis of backbone molecule (glycerol or sphingosine)

(2) attachment of fatty acid to the backbone

through as ester or amide linkage (3) addition of a hydrophilic head group to the backbone

through a phosphodiester linkage, in some

case (4) alternation or exchange of the head group to yield the final phospholipid product

Attachment of Head Group to Diacylglycerol

Attachment of Head Group to Diacylglycerol - The phospholipid head group is attached to a diacylglycerol

- The phospholipid head group is attached to a diacylglycerol by a phosphodiester bond, formed when phosphoric acid condenses with two alcohols, eliminating two molecules of H 2 O.

Attachment of Head Group to Diacylglycerol - The phospholipid head group is attached to a diacylglycerol

Forming the Phosphodiester Bond of the Head Group

Forming the Phosphodiester Bond of the Head Group • Either one of the alcohols is activated

• Either one of the alcohols

is activated by attaching to CDP (cytidine diphosphate)

• The free (not bound to CDP) alcohol then does nucleophilic attack on the CDP-activated phosphate

• Releases CMP and a glycerophospholipid –See Fig. 21-24

Forming the Phosphodiester Bond of the Head Group

Forming the Phosphodiester Bond of the Head Group Cells have two strategies for attaching phospholipid head

Cells have two strategies

for attaching phospholipid head groups

- CDP is used as activating group

(Fig 21-24)

Forming the Phosphodiester Bond of the Head Group Cells have two strategies for attaching phospholipid head

Phospholipid synthesis in E Coli employs CDP-diacylglycerol

- Phosphatidylserine, phosphatidylethanolamine, phosphatidylglycerol

(Fig

21-25)

***Eukaryotes synthesize anionic phosphoslipids from CDP-diacylglycerol

- phosphatidylglycerol, cardiolipin, and phosphatidylinocitols

Phospholipid Synthesis in E. coli employs CDP-diacylglycerol (Fig 21-

25)

Phospholipid Synthesis in E. coli employs CDP-diacylglycerol (Fig 21- 25) - Initially, a head group (either
Phospholipid Synthesis in E. coli employs CDP-diacylglycerol (Fig 21- 25) - Initially, a head group (either
  • - Initially, a head group

(either serine or glycerol 3-phosphate)

is attached via a CDP-diacylglycerol intermediate (strategy 1 in Fig. 21–24).

  • - For phospholipids other than

phosphatidylserine, the head group is further modified, as shown here.

In the enzyme names, PG represents phosphatidylglycerol; PS, phosphatidylserine.

Synthesis of Anionic Lipids Cardiolipin and Phosphatidylinositol in Eukaryotes

Synthesis of Anionic Lipids Cardiolipin and Phosphatidylinositol in Eukaryotes Cardiolipin –a phospholipid existing in heart and

Cardiolipin

–a phospholipid existing in heart and mitochondria

Synthesis of Anionic Lipids Cardiolipin and Phosphatidylinositol in Eukaryotes Cardiolipin –a phospholipid existing in heart and

Synthesis of Phosphatidylethanolamine and Phosphatidylcholine in all eukaryotes (Fig 21-27)

Synthesis of Phosphatidylethanolamine and Phosphatidylcholine in all eukaryotes (Fig 21-27) The major path from phosphatidylserine to

The major path from phosphatidylserine to phosphatidylethanolamine and

phosphatidylcholine

in all eukaryotes.

AdoMet is S-adenosylmethionine;

adoHcy, is S-adenosylhomocysteine.

Synthesis of Phosphatidylethanolamine and Phosphatidylcholine in all eukaryotes (Fig 21-27) The major path from phosphatidylserine to

Phosphatidic acid is formed by transacylation of L-glycerol 3-phosphate with fatty acyl groups donated from fatty acyl-CoA.

Summary of the pathway for synthesis of major phospholipids and triacylglyceroles

Summary of the pathway for synthesis of major phospholipids and triacylglyceroles
Summary of the pathway for synthesis of major phospholipids and triacylglyceroles

Pathways for phosphatidylserine synthesis in mammals (a)

Pathways for phosphatidylserine synthesis in mammals (a)
Pathways for phosphatidylserine synthesis in mammals (a) A salvage pathway using serine or ethanolamine PSS1; Phosphatidylserine

A salvage pathway using serine or ethanolamine

PSS1; Phosphatidylserine synthase 1

Pathways for phosphatidylcholine synthesis in mammals (b)

A salvage pathway using choline

Pathways for phosphatidylcholine synthesis in mammals (b) A salvage pathway using choline

Synthesis of ether lipids and plasmalogens

Synthesis of ether lipids and plasmalogens
Synthesis of ether lipids and plasmalogens Plasmogen , ether lipids , involves the displacement of an

Plasmogen, ether lipids, involves the displacement of an esterified fatty acyl group by a long-chain alcohol to form the ether linkage

Sphigolipid and glycerophospholipid synthesis share precursors and some mechanisms

- The biosynthesis of sphingolipid take place in four stages (Fig 21-31)

Palmitoyl CoA +

Serine + NADPH

---

(1) Sphinganine synthesis (2) N-acylsphingasine (3) N-acysphingosine (4) cerebroside or sphigomyelin produce

Polar lipids are targeted to specific cellular membranes

  • - synthesis on the smooth ER

  • - are inserted into specific cellular memnrane

Biosynthesis of sphingolipids Condensation of palmitoyl CoA and serine followed by reduction with NADPH yields sphiganine,

Biosynthesis of sphingolipids

Biosynthesis of sphingolipids Condensation of palmitoyl CoA and serine followed by reduction with NADPH yields sphiganine,

Condensation of palmitoyl CoA and serine followed by reduction with NADPH yields sphiganine,

which is then acylated to a

ceramide

then produce a sphingolipid such as a cerebroside or sphigomyelin

  • 21.4 Cholesterol, Steroids, and Isoprenes

21.4 Cholesterol, Steroids, and Isoprenes • Topics: – Cholesterol synthesis – Cholesterol transport – Cholesterol uptake

• Topics:

– Cholesterol synthesis – Cholesterol transport – Cholesterol uptake – Normal and defective regulation of cholesterol synthesis, uptake and transport – Cholesterol derivatives (bile acids, steroid hormones) – Synthesis of compounds derived from isoprene

21.4 Biosynthesis

of cholesterol, steroids, isoprenoids

21.4 Biosynthesis of cholesterol, steroids, isoprenoids Cholesterol is important for human cardiovascular disease , a component

Cholesterol is important for human cardiovascular disease,

a component of cellular membranes,

Cholesterol is made from acetyl-CoA in four

stages

and as a precursor of steroid hormones and bile

acids

- is made from acetyl CoA (Fig 21-32)

- synthesized by four steps (Fig 21-33)

Acetate -> Mevalonate -> Squalene -> Cholesterol

(Fig 21-34, 35, 36, 37)

Origin of the carbon atoms of cholesterol

Origin of the carbon atoms of cholesterol
Origin of the carbon atoms of cholesterol

Four Steps of Cholesterol Synthesis

Four Steps of Cholesterol Synthesis 1) Three acetate condense to form mevalonate 2) Mevalonate converts to

1) Three acetate

condense to form mevalonate

2) Mevalonate

converts to phosphorylated 5-C

isoprene 3) Six isoprene polymerize to form the 30-C linear squalene 4) Squalene

cyclizes to form the four rings that are modified to produce

cholesterol

Cholesterol

Synthesis

Cholesterol Synthesis The four stages are discussed in the above. Isoprene units in squalene are set

The four stages are discussed in the above.

Isoprene units in squalene are set off by red dashed lines.

Cholesterol Synthesis The four stages are discussed in the above. Isoprene units in squalene are set

Step 1:

Formation of Mevalonate from Acetyl- CoA

Step 1: Formation of Mevalonate from Acetyl- CoA The origin of C-1 and C-2 of mevalonate

The origin of C-1 and C-2 of mevalonate from acetyl-CoA is shaded light red.

Step 1: Formation of Mevalonate from Acetyl- CoA The origin of C-1 and C-2 of mevalonate

Step 2:

Conversion of Mevalonate to Activated Isoprenes

Step 2: Conversion of Mevalonate to Activated Isoprenes Six of these activated units combine to form

Six of these activated units combine to form squalene (see Fig. 21–36).

Step 2: Conversion of Mevalonate to Activated Isoprenes Six of these activated units combine to form

Formatio

n

Formatio n of Squalene

of

Squalene

Formatio n of Squalene

Conversion of Squalene to Cholesterol

Conversion of Squalene to Cholesterol The first step in this sequence is catalyzed by a mixed-function

The first step in this sequence is catalyzed by a mixed-function oxidase (a monooxygenase), for which the cosubstrate is NADPH.

The product is an epoxide, which in the next step is cyclized to the steroid nucleus.

The final product of these reactions in animal cells is cholesterol; in other organisms, slightly different sterols are produced, as shown.

Cholesterol has several fates

Cholesterol has several fates

(p864)

three form:

- synthesized in liver, exported in one of

biliary cholesterol, bile acids, cholesteryl

ester

- used as membrane synthesis, steroid hormone precursor, vitamin D

precursor

Cholesterol and other lipids

(Fig 21-38, explain!!!)

are carried on plasma lipoproteins

Cholesterol has several fates (p864) • three form: - synthesized in liver, exported in one of

- is insoluble in water, - is carried in blood plasma as plasma lipoprotein, apolipoprotein

(Fig 21-39)

- major classes of plasma protein (Table 21-1)

(Table 21-2)

- chylomicron, VLDL, LDL, HDL

Metabolic fates of cholesterol

Metabolic fates of cholesterol • Modification of the cholesterol structure are shown in red . •

• Modification of the cholesterol structure are shown in red.

• Esterification converts cholesterol to an even more hydrophobic form for storage and transport

Metabolic fates of cholesterol

Metabolic fates of cholesterol
Metabolic fates of cholesterol

Lipoprotein

Lipoprotein
Lipoprotein

Electron Microscope Pictures of Lipoproteins

Electron Microscope Pictures of Lipoproteins chylomicrons , 50 to 200 nm in diameter; VLDL , 28

chylomicrons, 50 to 200 nm in diameter;

VLDL, 28 to 70 nm;

HDL, 8 to 11 nm;

and

LDL, 20 to 25 nm.

Four Major Classes of Lipoprotein Particles

Four Major Classes of Lipoprotein Particles • Named based on position of sedimentation (density) in centrifuge

• Named based on position of sedimentation (density) in centrifuge

• Large enough to see in electron microscope (see Fig. 21-39b; next slide).

• Includes:

– Chylomicrons (largest and least dense) – Very low-density lipoproteins (VLDL) – Low-density lipoproteins (LDL)

– High-density lipoproteins (HDL) – smallest, most dense

Apolipoproteins in Lipoproteins

Apolipoproteins in Lipoproteins • “Apo” for “without”… – So “apolipoprotein” refers to the protein part of

• “Apo” for “without”…

– So “apolipoprotein” refers to the protein part of a lipoprotein particle • Provide sites for the particle to bind to

cell surface receptors, activate enzymes,

etc.

• At least ten have been characterized in humans

• See Table 21-2

Lipoproteins and lipid transport

Lipoproteins and lipid transport

Lipoproteins and lipid transport

Lipoproteins and lipid transport

Lipids are transported in the bloodstream as lipoproteins, which exist as several variants that have different functions, different protein and lipid compositions and thus different densities.

LDLs carry cholesterol from liver to muscle and adipose tissue

LDLs carry cholesterol from liver to muscle and adipose tissue • Muscle and adipose tissue have

• Muscle and adipose tissue have LDL receptors, recognize

apoB-100

– See Fig. 21-41

Enable myocytes and adipocytes to take up cholesterol via receptor-mediated endocytosis

Cholesterol esters enter cell by receptor

mediated endocytosis (Fig 21-41) (Fig 21-42)

Cholesterol biosynthesis is regulated at several steps

(Fig 21-43) (Fig 21-44)

  • - is regulated by cholesterol concentration, glucagon, insulin and HMG-CoA reductase ……

Atherosclerosis

- caused by unregulated cholesterol production in blood vessels

  • - high level of LDL-bound cholesterol; there is negative correlation between HDL level

  • - caused heart failure & death

  • - In familial hypercholesterolemia, severe atherosclerosis develops in childhood

Cholesterol Uptake by Receptor-Mediated Endocytosis

Cholesterol Uptake by Receptor-Mediated Endocytosis
Cholesterol Uptake by Receptor-Mediated Endocytosis

Lecithin-Cholesterol Acyl Transferase (LCAT) - Catalyzed Reaction

Lecithin-Cholesterol Acyl Transferase (LCAT) - Catalyzed Reaction This enzyme is present on the surface of HDL

This enzyme is present on the surface of HDL and is stimulated by the HDL component apoA-I.

Lecithin-Cholesterol Acyl Transferase (LCAT) - Catalyzed Reaction This enzyme is present on the surface of HDL

Cholesteryl esters accumulate within nascent HDLs, converting them to mature HDLs.

Five Modes of Regulation of Cholesterol Synthesis and Transport

Five Modes of Regulation of Cholesterol Synthesis and Transport 1) Covalent modification of HMG-CoA reductase 2)

1) Covalent modification of HMG-CoA reductase 2) Transcriptional regulation of HMG-CoA gene 3) Proteolytic degradation of HMG-CoA reductase 4) Activation of ACAT, which increases esterification for storage 5) Transcriptional regulation of the LDL receptor

HMG-CoA reductase is most active when dephosphorylated

HMG-CoA reductase is most active when dephosphorylated 1) AMP-dependent protein kinase - when AMP rises, kinase

1) AMP-dependent protein kinase - when AMP rises,

kinase phosphorylates the enzyme

activity , cholesterol synthesis

2) Glucagon, epinephrine

3)

  • - cascades lead to phosphorylation, activity Insulin

  • - cascades lead to dephosphorylation,activity

Covalent modification provides short-term regul

Regulation of Cholesterol Metabolism

Regulation of Cholesterol Metabolism

Regulation of Cholesterol Metabolism

Longer-term Regulation of HMG-CoA Reductase

through Transcriptional Control

Longer-term Regulation of HMG-CoA Reductase through Transcriptional Control (Fig 21-44) • Sterol regulatory element-binding proteins (SREBPs)

(Fig 21-44)

• Sterol regulatory element-binding proteins (SREBPs)

– When sterol levels are low, SREBP is in ER membrane with other proteins – When sterol levels rise, complex is cleaved, moves to the nucleus activates transcription of HMG-CoA reductase and LDL receptor as well as other genes

Regulation of Cholesterol Synthesis by SREBP

Regulation of Cholesterol Synthesis by SREBP
Regulation of Cholesterol Synthesis by SREBP

Steroid hormones

are formed by side- chain cleavage

and oxidation of cholesterol

- Human derive all their steroid hormones from cholesterol

(Fig 21-45) (Fig 21-46)

Intermediates in cholesterol biosynthesis

have many

alternative fates

Regulation by LXR- mediated Transcription

Regulation by LXR- mediated Transcription • Liver X Receptor (LXR) – a transcription factor activated by

• Liver X Receptor (LXR)

a transcription factor activated by cholesterol • Binds to retinoid X receptor (RXR) LXR-RXR dimer activates transcription of a host

of genes

Action RXR-LXR dimer on expression of genes for lipid and glucose metabolism

Action RXR-LXR dimer on expression of genes for lipid and glucose metabolism

Review of Cholesterol Regulation

Review of Cholesterol Regulation • Insulin cascades stimulate HMG-CoA reductase • Transcription increases when cholesterol levels

• Insulin cascades

stimulate HMG-CoA reductase

• Transcription

increases when cholesterol levels are

low

• Transcription

- SREBPs activated

slows when cholesterol levels are high

- LXRs activated

Cardiovascular disease (CVD) is multi-factorial

Cardiovascular disease (CVD) is multi-factorial • Very high LDL-cholesterol levels tend to correlate with atherosclerosis –

Very high LDL-cholesterol levels tend to correlate with atherosclerosis

– Although many heart attack victims have normal cholesterol, and many people with high cholesterol do not have heart attacks

Low HDL-cholesterol levels

are negatively associated with heart disease

Role of form cells in formation of atherosclerotic plaques

Role of form cells in formation of atherosclerotic plaques
Role of form cells in formation of atherosclerotic plaques

How Plaques Form

How Plaques Form • LDL with partly oxidized fatty acyl groups sticks to the lining of

LDL with partly oxidized fatty acyl groups sticks to the lining of arteries. - attracts macrophage cells of the immune system. • These cells don’t regulate their uptake of sterols, so they accumulate cholesterol and cholesteryl esters • The macrophages become foam cells (named for appearance)

How Plaques Form (cont.)

How Plaques Form (cont.) • Foam cells undergo apoptosis • Remnants accumulate, along with scar tissue,

Foam cells undergo apoptosis Remnants accumulate, along with scar tissue, etc.

can occlude a blood vessel or break off and travel to another artery

Occlusion of blood vessels in the heart cause heart attack; occlusion in the brain causes stroke

Familial

Hypercholesterolemia

Familial Hypercholesterolemia • Due to genetic mutation in LDL receptor • Impairs receptor-mediated uptake of cholesterol

• Due to genetic mutation in LDL receptor • Impairs receptor-mediated uptake of cholesterol from LDL Cholesterol accumulates in the blood

and in foam cells Regulation mechanisms based on cholesterol sensing inside the cell don’t

work • Homozygous individuals can experience severe CVD as youths

Statin drugs inhibit HMG-CoA reductase to lower cholesterol

Statin drugs inhibit HMG-CoA reductase to lower cholesterol • Statins resemble mevalonate (p873)  competitive inhibitors

Statins resemble mevalonate

(p873)

competitive inhibitors of HMG-CoA reductase • First statin, lovastatin, found in fungi lowers serum cholesterol by tens of percent

• Side-effects include muscle weakness, coenzyme Q depletion

• Also reported to improve circulation, stabilize plaques by removing cholesterol from them, reduce vascular inflammation

Reverse Cholesterol Transport

Reverse Cholesterol Transport Apo A-I and HDLs pick up excess cholesterol from peripheral cells, with the

Apo A-I and HDLs pick up excess cholesterol from peripheral cells, with the participation of ABCA1 and ABCG1 transporters, and return it to the liver.

In individuals with genetically

defective ABCA1, the failure of reverse cholesterol transport leads to severe and early cardiovascular diseases:

Tangier disease and familial HDL deficiency disease.

Reverse Cholesterol Transport Apo A-I and HDLs pick up excess cholesterol from peripheral cells, with the

p HDL explains why HDL is

y

p HDL explains why HDL is y cardioprotective • HDL picks up cholesterol from non-liver tissues,

cardioprotective

HDL picks up cholesterol from non-liver tissues, including foam cells at growing plaques

(p873)

• ABC (ATP-Binding Cassette) transporters

bring cholesterol from inside the cell to the plasma membrane • HDL carries cholesterol back to liver

There are several classes of cholesterol-derived steroids

There are several classes of cholesterol-derived steroids • Adrenal gland-synthesized steroids: – Mineralocorticoids • Control electrolyte

• Adrenal gland-synthesized steroids:

– Mineralocorticoids

• Control electrolyte balance, reabsorption of Na + , Cl , HCO 3 from kidney

– Glucocorticoids

• Regulate gluconeogenesis, reduce inflammation

• Gonad-synthesized steroids:

– Progesterone, androgens, estrogens

Steroid Hormones Derived from Cholesterol

Steroid Hormones Derived from

Steroid Hormones Derived from Cholesterol

Cholesterol

How Steroids Are Made from Cholesterol

How Steroids Are Made from Cholesterol • Takes place in mitochondria • The “side chain” on

• Takes place in mitochondria • The “side chain” on C-17 of the D ring is modified or

cleaved • Two adjacent carbons are hydroxylated • Uses mixed-function oxidases, NADPH and

cytochrome P450 • See Fig. 21-49

Side-chain Cleavage in

Side-chain Cleavage in Steroid Synthesis - Cytochrome P-450 acts as electron carrier in this mixed-function oxidase

Steroid

Synthesis

  • - Cytochrome P-450 acts as electron carrier in this mixed-function oxidase system that oxidizes adjacent carbons.

  • - The process also requires the electron- transferring proteins adrenodoxin and adrenodoxin reductase. This system for cleaving side chains is found in mitochondria of the adrenal cortex, where active steroid production occurs.

Pregnenolone is the precursor of all other steroid hormones

(p875)

Side-chain Cleavage in Steroid Synthesis - Cytochrome P-450 acts as electron carrier in this mixed-function oxidase

Intermediates in Cholesterol Synthesis

Intermediates in Cholesterol Synthesis   3 -Isopentenyl pyrophosphate can be made into over 20,000 additional

3 -Isopentenyl pyrophosphate

can be made into over 20,000 additional products

• Includes Vitamins A, E, and D to plant hormones, rubber, fragrant oils, quinone electron carriers, and the phytol chain of chlorophyll, etc.

• Usually involve prenylation • See Fig. 21-50

Synthesis of Isoprenoids; Overview

Synthesis of Isoprenoids; Overview
Synthesis of Isoprenoids; Overview

Finish…!!!

Oxidases, Monooxygenases, and Dioxygenases

Oxidases, Monooxygenases, and Dioxygenases Many enzymes use oxygen as an e − acceptor, but not all

Many enzymes use oxygen as an e acceptor, but not all of them incorporate oxygen into the product.

Oxidases do not incorporate oxygen into the product Oxygen atoms usually end up in H 2 O 2 Oxygenases do incorporate oxygen into the product Monooxygenases incorporate one of the oxygen atoms into the product Dioxygenases incorporate both oxygen atoms into the product

Monooxygenases incorporate one oxygen into the product

Monooxygenases incorporate one oxygen into the product AH + BH + O-O  A-OH + B

AH + BH 2 + O-O A-OH + B + H 2 O

Product is often hydroxylated, so also called hydroxylases or mixed-function oxygenases

– Example: Phenylanine hydroxylase hydroxylates phenylalanine to form tyrosine

– Deficiency causes phenylketonuria

Monooxygenases incorporate one oxygen into the product AH + BH + O-O  A-OH + B

Cytochrome P450s are monooxygenases

Cytochrome P450s are monooxygenases • Important in drug metabolism • Hydroxylate nonpolar molecules – usually inactivating

Important in drug metabolism Hydroxylate nonpolar molecules

usually inactivating them and making them more H 2 O-solubile for excretion

If two drugs (or alcohol and a drug) use the same P450, they will compete, and levels of the drug or alcohol will not be cleared as quickly

Can be deadly

Dioxygenases incorporate two oxygens in the product

Dioxygenases incorporate two oxygens in the product • Usually metalloproteins – Active sites have Fe or

• Usually metalloproteins

– Active sites have Fe or Mn ions

• Rxs often involve opening an aromatic ring • Used to convert catechols to ketones • Example: Catechol 1,2-dioxygenase, Tryptophan 2,3-dioxygenase (Next slide)

Eicosanoids are potent short-range hormones made from arachidonate

Eicosanoids are potent short-range hormones made from arachidonate • Eicosanoid hormones include prostaglandins, leukotrienes, thromboxanes •

• Eicosanoid hormones include prostaglandins, leukotrienes, thromboxanes

• Created from the arachidonate 20:4 that’s incorporated into the phospholipids of membranes

• In response to stimuli (hormone, etc.), phospholipase A 2 is activated and attacks the C-2 fatty acid, releasing arachidonate

Prostaglandins are made by prostaglandin H 2 synthase

(PGH 2 )

Prostaglandins are made by prostaglandin H synthase (PGH ) • PGH functions in the smooth ER.

PGH 2 functions in the smooth ER. Step 1: PGH 2 ’s cyclooxygenase activity (COX), adds 2 O 2 to form PGG 2 Step 2: PGH 2 ’s peroxidase activity converts peroxide to alcohol, creates PGH 2 PGH 2 is precursor to other eicosanoids See Fig. 21-15a

PGH 2 has two isoforms

PGH has two isoforms • COX-1 catalyzes synthesis of prostaglandins that regulate gastric mucin secretion •

• COX-1 catalyzes synthesis of prostaglandins that regulate gastric mucin secretion

• COX-2 catalyzes synthesis of prostaglandins that mediate pain, inflammation, and fever

– NSAIDs (Aspirin, Ibuprofen, Acetaminophen) inhibit COX-2

– See Fig. 21-15b

COX-2-specific inhibitors have a checkered history

COX-2-specific inhibitors have a checkered history • Developed to inhibit prostaglandin formation without harming stomach •

• Developed to inhibit prostaglandin formation without harming stomach • Includes Vioxx, Bextra, and Celebrex

– See Fig. 21-15c

• Vioxx and Bextra removed from market due to increased rates of stroke and heart attack

– May disrupt balance between blood-thinning prostacyclin and blood-clotting thromboxanes

Fat (Triacylglycerol) and Phospholipids in Animals, Plants, and Bacteria

Fat (Triacylglycerol) and Phospholipids in Animals, Plants, and Bacteria • Animals and plants store fat for

Animals and plants store fat for fuel

Plants: in seeds, nuts Typical 70-kg human has ~15 kg fat

• Enough to last 12 wks

• Compare with 12 hrs’ worth glycogen in liver and muscle

Animals and plants and bacteria make phospholipids for cell membranes

The precursor for the backbone of fat and phospholipids is glycerol 3-phosphate

The precursor for the backbone of fat and phospholipids is glycerol 3-phosphate • Most glycerol 3-phosphate

• Most glycerol 3-phosphate comes from siphoning off dihydroxyacetone phosphate (DHAP) from glycolysis

– via glycerol 3-phosphate dehydrogenase

• Some glycerol 3-phosphate made from glycerol

– Via glycerol kinase – Minor pathway in liver and kidney

Acyl transferases attach two fatty acids to glycerol 3- phosphate

Acyl transferases attach two fatty acids to glycerol 3- phosphate • Phosphatidic acid is the precursor

• Phosphatidic acid is the precursor to TAGs and phospholipids

– Made of glycerol 3-phosphate + 2 fatty acids – Fatty acids are attached by acyl transferases – Releases CoA – See Fig. 21-17

• Advantage of making phosphatidic acid:

– It can then be made into triacylglycerol OR phospholipid

To make TAG, phosphatidic acid is dephosphorylated and acylated

To make TAG, phosphatidic acid is dephosphorylated and acylated • Phosphatidic acid phosphatase (lipin) removes the

• Phosphatidic acid phosphatase

(lipin) removes the 3-phosphate from the phosphatidic acid

– Yields 1,2-diacylglycerol

• Third carbon is then acylated with a third fatty acid

– Yields triacylglycerol – See Fig. 21-18

Regulation of Triacylglycerol Synthesis by Insulin

Regulation of Triacylglycerol Synthesis by Insulin • Insulin results in stimulation of triacylglycerol synthesis • Lack

Insulin results in stimulation of triacylglycerol synthesis Lack of insulin results in:

Increased lipolysis Increased fatty acid oxidation

• Sometimes to ketones if citric acid cycle intermediates (oxaloacetate) that react with acetyl CoA are depleted

Failure to synthesize fatty acids

Regulation of Fat Metabolism by Glucagon and Epinephrine

Regulation of Fat Metabolism by Glucagon and Epinephrine • Glucagon and epinephrine result in stimulation of

• Glucagon and epinephrine result in stimulation of triacylglycerol

breakdown (mobilization of fatty acids)

– Also decrease glycolysis – Also increase gluconeogenesis

Regulation of PEPCK expression is tissue- dependent

Regulation of PEPCK expression is tissue- dependent • Cortisol and glucagon both increase PEPCK expression in

Cortisol and glucagon both increase PEPCK expression in liver.

– Results in more TAG synthesis, so more released to the blood

Cortisol and other glucocorticoids decrease PEPCK expression in adipose tissue

glyceroneogenesis in adipose means less recycling; more FFA are released into the blood

– Most glycerol freed from TAG in adipose is sent to liver and converted to glucose

Cortisol and glucagon can elevate blood sugar

Cortisol and glucagon can elevate blood sugar   PEPCK expression in liver   gluconeogenesis

  PEPCK expression in liver gluconeogenesis (so [glucose])

  PEPCK expression in adipose tissue glycerol freed, sent to liver, converted to glucose

3) Plus, the FFA associated with increased flux through TAG cycle interfere with glucose uptake in muscle, keep [glucose] blood high

CDP-diacylglycerol is a precursor to phosphatidylserine and derivatives in E. coli

CDP-diacylglycerol is a precursor to phosphatidylserine and derivatives in E. coli • See Fig. 21-25 •

• See Fig. 21-25

• CDP-diacylglycerol is attacked by the nucleophilic OH of serine phosphatidylserine (and CMP)

– Catalyzed by PS synthase

• Phosphatidylserine is decarboxylated Phosphatidylethanolamine

– Catalyzed by PS decarboxylase

CDP-diacylglycerol is also a precursor to cardiolipin in E. coli

CDP-diacylglycerol is also a precursor to cardiolipin in E. coli • See Fig. 21-25 • CDP-diacylglycerol’s

• See Fig. 21-25

• CDP-diacylglycerol’s activated phosphate is attacked by glycerol 3-phosphate Phosphatidylglycerol 3-phosphate (and CMP)

Catalyzed by PG3-phosphate synthase

• PG 3-phosphate is hydrolyzed PGglycerol

Catalyzed by PG3-phosphate phosphatase

• Phosphatidylglycerol dimerizes to cardiolipin

Catalyzed by cardiolipin synthase

Synthesis of anionic phospholipids in eukaryotes uses similar strategies to that of E. coli

Synthesis of anionic phospholipids in eukaryotes uses similar strategies to that of E. coli • Phosphatidylinositol

Phosphatidylinositol made from CDP-diacylglycerol and inositol

• Specific kinases make phosphorylated derivatives

• Difference from bacteria:

Cardiolipin made from attack of phosphatidylglycerol on CDP-diacylglycerol rather than from two phosphatidylglycerols

Synthesis of Phosphatidylethanolamine and Phosphatidylcholine in Yeast

Synthesis of Phosphatidylethanolamine and Phosphatidylcholine in Yeast • See Fig. 21-27 • Phosphatidylserine is decarboxylated to

• See Fig. 21-27

• Phosphatidylserine is decarboxylated to

phosphatidylethanolamine

– (as in bacteria, but enz is phosphatidylserine decarboxylase)

• Phosphatidylethanolamine acted on by S- adenosylmethione (methyl group donor),

adds three methyl groups to amino group phopshatidylcholine (lecithin)

– Catalyzed by methyltransferase

Phospholipid Synthesis in Mammals

Phospholipid Synthesis in Mammals • Phosphatidylserine isn’t synthesized from CDP-diacylglycerol as it is in yeast and

Phosphatidylserine isn’t synthesized from CDP-diacylglycerol as it is in yeast and bacteria

• See Fig. 21-29a • Made “backwards” from phosphatidylethanolamine or phosphatidylcholine via head group exchange rxs

Catalyzed by specific synthases Pathway “salvages” the choline

Synthesis of Phosphatidylserine and Phosphatidylcholine in Mammals

Synthesis of Phosphatidylserine and Phosphatidylcholine in Mammals
Synthesis of Phosphatidylserine and Phosphatidylcholine in Mammals

Synthesis of Phosphtidylcholine in Mammals by Another Pathway

Synthesis of Phosphtidylcholine in Mammals by Another Pathway
Synthesis of Phosphtidylcholine in Mammals by Another Pathway

Formation of Plasmalogens and Other Ether Lipids

Formation of Plasmalogens and Other Ether Lipids • Ether linkage is made by displacing an esterified

• Ether linkage is made by displacing an esterified fatty acid with a long- chain alcohol

• Attachment of head groups is similar to the pathways for ester- linked phospholipids

– Fig. 21-30 in the text summarizes these pathways

Sphingolipids are made in four steps

Sphingolipids are made in four steps 1) Synthesis of sphinganine from palmitoyl- CoA and serine 2)

1) Synthesis of sphinganine from palmitoyl- CoA and serine

2) Attachment of fatty acid via amide linkage

3) Desaturation of sphinganine

Yields N-acylsphingosine (ceramide)

4) Attachment of head group

Can yield a cerebroside or ganglioside

See Fig. 21-31 in text for detailed pathways

Phospholipids must be transported from the ER to membranes

Phospholipids must be transported from the ER to membranes • Phospholipids are: – synthesized in the

• Phospholipids are:

synthesized in the smooth ER transported to Golgi complex for additional synthesis

• Must be inserted into specific membranes in specific proportions but can’t diffuse because they are nonpolar

• So transported in membrane vesicles that fuse with target membrane

• Details of the process are not well-understood

Step 1: Formation of Mevalonate from Acetyl-CoA

Step 1: Formation of Mevalonate from Acetyl-CoA • 2 Acetyl-CoAs  Acetyoacetyl-CoA – Catalyzed by acetyl-CoA

2 Acetyl-CoAs Acetyoacetyl-CoA

– Catalyzed by acetyl-CoA acyl transferase

Acetyl-CoA + Acetoacetyl-CoA -hydroxyl- -methylglutaryl-CoA (HMG-CoA)

– Catalyzed by HMG-CoA synthase

• NOT the mitochondrial HMG-CoA synthase used in ketone body formation

HMG-CoA + 2 NADPH mevalonate

– Catalyzed by HMG-CoA reductase Rate-limiting step and point of regulation!

Formation of Mevalonate

Formation of Mevalonate • HMG-CoA reductase is a target for some cardiovascular drugs

• HMG-CoA reductase is a target for some

cardiovascular drugs

Step 2: Conversion of Mevalonate to Two Activated Isoprenes

Step 2: Conversion of Mevalonate to Two Activated Isoprenes • 3 PO 3− transferred stepwise from

• 3 PO 4 3− transferred stepwise from ATP to mevalonate

• Decarboxylation and hydrolysis creates a diphosphorylated 5-C product (isoprene) with a double bond • Isomerization to a second isoprene

• The two “activated” isoprene units are 3 -isopentyl pyrophosphate and dimethylallylpyrophosphate

Step 3: Six Activated Isoprene Units Condense to Form Squalene

Step 3: Six Activated Isoprene Units Condense to Form Squalene • The two isoprenes join head-to-tail,

• The two isoprenes join head-to-tail, displacing one set of diphosphates

forms geranyl pyrophopshate

• Geranyl pyrophosphate joins to another isopentenyl pyrophosphate

forms 15-C farnesyl pyrophosphate

• Two farnesyl pyrophosphates join head- to-head to form phosphate-free

squalene

Step 4: Conversion of Squalene to Four-Ring Steroid Nucleus

Step 4: Conversion of Squalene to Four-Ring Steroid Nucleus • Squalene monooxygenase adds one oxygen to

• Squalene monooxygenase adds one oxygen to the end of the squalene chain

forms squalene 2,3-epoxide

• Here pathways diverse in animal cells vs. plant cells

• The cyclization product in animals is lanosterol, which converts to cholesterol

• In plants, the epoxide cyclizes to other sterols such as ergosterol

Bile Acids Assist in Emulsification of Fats

Bile Acids Assist in Emulsification of Fats • Bile is stored in the gall bladder, secreted

• Bile is stored in the gall bladder, secreted into small intestine after fatty meal

• Bile acids such as taurocholic acid (see Fig. 21-23) emulsify fats

– Surround droplets of fat, increase surface area for attack by lipases

Cholsteryl esters are more nonpolar than cholesterol

Cholsteryl esters are more nonpolar than cholesterol • Contain a fatty acid esterified to the oxygen

• Contain a fatty acid esterified to the oxygen (see Fig. 21-38)

– Comes from a fatty acyl-CoA

– Makes the cholesterol more hydrophobic, unable to enter membranes

• Transported in lipoproteins to other tissues or stored in liver

Cholesterol and other lipids are carried on lipoprotein particles

Cholesterol and other lipids are carried on lipoprotein particles • Lipids are carried through the plasma

• Lipids are carried through the plasma on spherical particles

– Surface is made of protein (called apolipoprotein) and a phospholipid monolayer

– Interior contains cholesterol, TAGs, cholesteryl esters

• See Fig. 21-39a (next slide)

Chylomicrons carry fatty acids to tissues

Chylomicrons carry fatty acids to tissues • Have more TAG and less protein  hence, least

• Have more TAG and less protein hence, least dense.

• Have ApoB-48, ApoE, and ApoC-II

• ApoC-II activates lipoprotein lipase to allow FFA release for fuel in adipose tissue, heart, and skeletal muscle

Chylomicron remnants deposit their cholesterol in the liver

Chylomicron remnants deposit their cholesterol in the liver • When chylomicrons are depleted of their TAG,

• When chylomicrons are depleted of their TAG, “remnants” go to liver

• ApoE receptors in liver bind the remnants, take them up by endocytosis

• Remnants release their cholesterol in the liver

VLDLs transport endogenous lipids

VLDLs transport endogenous lipids • Cholesteryl esters and TAGs from excess FA and cholesterol are packed

• Cholesteryl esters and TAGs from excess FA and cholesterol are packed into very low-density lipoproteins (VLDL)

• Excess carbohydrate in the diet can also be made into TAG in the liver and packed into VLDL

• Contain apoB-100, apoC-I, apoC-II, apoC-III, and apoE

VLDLs take TAGs to adipose tissue and muscle

VLDLs take TAGs to adipose tissue and muscle • Again, ApoC-II activates lipoprotein lipase to release

• Again, ApoC-II activates lipoprotein lipase to release free fatty acids • Adipocytes take up the FFA, reconvert them to TAGs, and store them in lipid droplets • Muscle uses the TAG for energy

VLDL remnants become LDL

VLDL remnants become LDL • Removal of TAG from VLDL produces LDL • Because TAG removed,

• Removal of TAG from VLDL produces LDL

• Because TAG removed, LDL is enriched in cholesterol/chloesteryl esters

• ApoB-100 is the major apolipoprotein

Familial hypercholesterolemia is associated with LDL receptor mutations

Familial hypercholesterolemia is associated with LDL receptor mutations • Mutations in LDL receptor prevent normal uptake

• Mutations in LDL receptor prevent normal uptake of LDL by liver and other tissues

• LDL accumulates in blood

• Homozygous individuals have much increased risk of heart attack

• Heterozygous individuals have risk of heart attack greater than normal

HDL carries out reverse cholesterol transport

HDL carries out reverse cholesterol transport • HDLs contain a lot of protein – Including ApoA-I

• HDLs contain a lot of protein

– Including ApoA-I and – lecithin-cholesterol acyl transferase (LCAT)

• Catalyzes the formation of cholesteryl esters from lecithin and cholesterol

• Enzyme converts the cholesterol of chylomicron and VLDL remnants to cholesteryl esters

• HDL picks up cholesterol from cells and returns them to the liver

• See Fig. 21-42

Regulation of HMG-CoA Reductase by Proteolytic Degradation

Regulation of HMG-CoA Reductase by Proteolytic Degradation • Insig ( ins ulin- i nduced g ene

• Insig (insulin-induced gene protein) senses cholesterol levels.

– Triggers ubiquination of HMG-CoA reductase

– Targets the enzyme for degradation by proteasomes

The genes activated by LXR- RXR are largely for cholesterol transport

The genes activated by LXR- RXR are largely for cholesterol transport • Acetyl-CoA Carboxylase – First

• Acetyl-CoA Carboxylase

– First enzyme in fatty acid synthesis

• Apoproteins (C1, C2, D, and E)

– For cholesterol transport

• GLUT4 • ABC transporters

– For reverse cholesterol transport

Review of Cholesterol Regulation

Review of Cholesterol Regulation • Insulin cascades stimulate HMG- CoA reductase • Transcription increases when cholesterol

• Insulin cascades stimulate HMG- CoA reductase

• Transcription increases when cholesterol levels are low

– SREBPs activated

• Transcription slows when cholesterol levels are high

– LXRs activated

How Plaques Form

How Plaques Form • LDL with partly oxidized fatty acyl groups sticks to the lining of

• LDL with partly oxidized fatty acyl groups sticks to the lining of arteries

• Attracts macrophage cells of the immune system

• These cells don’t regulate their uptake of sterols, so they accumulate cholesterol and cholesteryl esters

• The macrophages become foam cells (named for appearance)

How Plaques Form (cont.)

How Plaques Form (cont.) • Foam cells undergo apoptosis • Remnants accumulate, along with scar tissue,

• Foam cells undergo apoptosis • Remnants accumulate, along with scar tissue, etc.

• Can occlude a blood vessel or break off and travel to another artery

• Occlusion of blood vessels in the heart cause heart attack; occlusion in the brain causes stroke

Familial

Hypercholesterolemia

Familial Hypercholesterolemia • Due to genetic mutation in LDL receptor • Impairs receptor-mediated uptake of cholesterol

• Due to genetic mutation in LDL receptor • Impairs receptor-mediated uptake of cholesterol from LDL • Cholesterol accumulates in the blood and in foam cells • Regulation mechanisms based on cholesterol sensing inside the cell don’t work • Homozygous individuals can experience severe CVD as youths

Statin drugs inhibit HMG- CoA reductase to lower cholesterol

Statin drugs inhibit HMG- CoA reductase to lower cholesterol • Statins resemble mevalonate  competitive inhibitors

• Statins resemble mevalonate competitive inhibitors of HMG-CoA reductase • First statin, lovastatin, found in fungi • Lowers serum cholesterol by tens of percent • Side-effects include muscle weakness, coenzyme Q depletion • Also reported to improve circulation, stabilize plaques by removing cholesterol from them, reduce vascular inflammation

Reverse cholesterol transport by HDL explains why HDL is cardioprotective

Reverse cholesterol transport by HDL explains why HDL is cardioprotective • HDL picks up cholesterol from

• HDL picks up cholesterol from non- liver tissues, including foam cells at growing plaques

• ABC (ATP-Binding Cassette) transporters bring cholesterol from inside the cell to the plasma membrane • HDL carries cholesterol back to liver

There are several classes of cholesterol-derived steroids

There are several classes of cholesterol-derived steroids • Adrenal gland-synthesized steroids: – Mineralcorticoids • Control electrolyte

• Adrenal gland-synthesized steroids:

– Mineralcorticoids

• Control electrolyte balance, reabsorption of Na + , Cl , HCO 3 from kidney

– Glucocorticoids

• Regulate gluconeogenesis, reduce inflammation

• Gonad-synthesized steroids:

– Progesterone, androgens, estrogems