Ketone Body Formation; Fatty acid and Cholesterol...

Ketone Body Formation; Fatty acid

and Cholesterol Biosynthesis

Ketone Body Formation; Fatty acid

and Cholesterol Biosynthesis

• Most acetyl CoA from β-oxidation is routed through the TCA cycle

• Excess is used to synthesize ketone bodies; β-hydroxybutyrate, acetoacetate and

acetone

• Ketone bodies serve as fuel molecules

• Liver is the site of synthesis in mammals (occurs in the mitochondrial matrix)

H3C-C-S-CoAO

H3C-C-S-CoAO

Acetyl CoA Acetyl CoA

+

Ketone Body Synthesis:

H3C-C-S-CoAO

H3C-C-S-CoAO

Acetyl CoA Acetyl CoA

+

H3C-C-CH2-C-S-CoA

O O

Acetoacetyl CoA

thiolaseHSCoA

H3C-C-CH2-C-S-CoA

O O

Acetoacetyl CoA

H3C-C-S-CoAO

+

Acetyl CoA

H3C-C-CH2-C-S-CoA

O O

Acetoacetyl CoA

H3C-C-S-CoAO

+

Acetyl CoA

3-Hydroxy-3-Methylglutaryl CoA (HMG CoA)

-OOC-CH2-C-CH2-C-S-CoAOOH

CH3

HMG CoA Synthase

HSCoA

3-Hydroxy-3-Methylglutaryl CoA (HMG CoA)

-OOC-CH2-C-CH2-C-S-CoAOOH

CH3

HMG CoA Lyase

H3C-C-S-CoAO

Acetyl CoA

-OOC-CH2-C-CH3

O

Acetoacetate

-OOC-CH2-C-CH3

O

Acetoacetate

β-Hydroxybutyrate

D’HaseNADH

NAD+

-OOC-CH2-CH-CH3

OH

β-Hydroxybutyrate

CO2

CH3-C-CH3

O

Acetone

Ketone Body Breakdown:

-OOC-CH2-CH-CH3

OH

β-Hydroxybutyrate

β-Hydroxybutyrate

D’Hase

NADH

NAD+

-OOC-CH2-C-CH3

O

Acetoacetate

*Liver enzyme catalyzes a near equilibrium

reaction

*Isozyme found in other cells catalyzes

an irreversible reaction

-OOC-CH2-C-CH3

O

Acetoacetate

+-OOC-CH2-CH2-C-S-CoA

O

Succinyl CoASuccinyl CoA Transferase

H3C-C-CH2-C-S-CoA

O O

Acetoacetyl CoA

+

-OOC-CH2-CH2-COO-

Succinate

H3C-C-CH2-C-S-CoA

O O

Acetoacetyl CoA

Thiolase

H3C-C-S-CoAO

Acetyl CoA

H3C-C-S-CoAO

Acetyl CoA

Fatty acid BiosynthesisFatty acid

Biosynthesis

• Occurs through the condensation of C2 units

• Occurs through the condensation of C2 units

• Occurs largely in adipocytes and liver and to a lesser extent in specialized tissues

• Occurs largely in adipocytes and liver and to a lesser extent in specialized tissues

• Occurs in three stages: 1) transport of acetyl Co A to the cytosol; 2) formation of malonyl CoA; 3) assembly of the fatty acid chain

• Occurs in three stages: 1) transport of acetyl Co A to the cytosol; 2) formation of malonyl CoA; 3) assembly of the fatty acid chain

Citrate Transport System:Citrate Transport System:

MatrixMatrixCytosolCytosol

Citrate:Dicarboxylic acid Carrier

Acetyl CoA + OAA

Citrate

α-Kg or

malate




Acetyl CoA + OAA

Citrate

α-Kg or

malate

Citrate

α-Kg

or malate




Citratecitrate lyase ATP

ADP

OAA + Acetyl CoAmalate d’hase NAD+

NADH

malate

Malic enzyme

NADP+ NADPH

CO2

pyruvatepyruvate translocase pyruvate

carboxylase

• Formation of Malonyl CoA by Acetyl CoA carboxylase

• Loading Step: Transfer of acetyl CoA and malonyl CoA to ACP to form acetyl-ACP

and malonyl ACP

• Successive rounds of condensation; reduction;

dehydration, reduction

Fatty Acid Synthetase Fatty Acid Synthetase }}

H3C-C-S-CoA

O

acetyl-CoA

+ HCO3

bicarbonate

Acetyl CoA Carboxylase

ATP

ADP, Pi

-OOC-CH2-C-S-CoAO

malonyl-CoA

Fatty Acid Synthetase Complex:

Bacteria, PlantsSeven activities in 7 separate proteins

YeastSeven activities in 2 separate proteins

VertebratesSeven activities in 1 large protein

H3C-C-S-CoA

Oacetyl-CoA

-OOC-CH2-C-S-CoAO

malonyl-CoA

Acetyl CoA:ACP transacylase

Malonyl CoA:ACP transacylase

HS-ACP

HS-CoA

HS-ACP

HS-CoA

H3C-C-S-ACP

O-OOC-CH2-C-S-ACP

O

acetyl-ACP malonyl-ACP

Loading Step

H3C-C-S-ACP

O-OOC-CH2-C-S-ACP

O


ketoacyl-ACP synthase

Condensation

H3C-C-S-ACP

O-OOC-CH2-C-S-ACP

O


H3C-C-CH2-C-S-ACP

OO

acetoacetyl-ACP

ketoacyl-ACP synthaseCO2

HS-ACP

Condensation

H3C-C-CH2-C-S-ACP

OO

acetoacetyl-ACPketoacyl-ACP

reductase

NADP+

NADPH

b-hydroxybutyryl-ACP

H3C-C-CH2-C-S-ACP

OOH

H

Reduction

b-hydroxybutyryl-ACP

H3C-C-CH2-C-S-ACP

OOH

H

b-hydroxyacyl-ACP dehydrase

H20

H3C-C=C-C-S-ACPO

H

H

butenoyl-ACP

Dehydration

H3C-C=C-C-S-ACPO

H

H

butenoyl-ACP

enoyl-ACP reductase

NADPHNADP+

H3C-CH2-CH2-C-S-ACPO

butyryl-ACP

Reduction

• Fatty Acid Synthesis is sometimes called palmitate synthesis because palmitate is the

predominant product (in mammals)

• Rounds of synthesis continue until a C16 palmitoyl group is formed

• Thiolase then catalyzes the formation of palmitate and ACP-SH

Palmitoyl-ACPPalmitate +

ACP-SHthiolase

Overall stoichiometry for palmitate synthesis:

Acetyl-CoA + 7 Malonyl CoA + 14 NADPH, H+

palmitate + 7CoA + 7HCO3 14 NADP+

Regulation:Regulation:• Key regulatory step: acetyl-CoA carboxylase

• Citrate allosterically activates.

• Palmitate is a feedback inhibitor

• Fatty acyl-CoAs allosterically inhibit

• Phosphorylation (stimulated by glucagon and epinephrine) inactivates

Regulation:

• Fatty acid synthesis occurs in the chloroplast (stroma) in plants

• The plant acetyl-CoA carboxylase is not regulated by phosphorylation; it is activated by increases in stromal

pH and magnesium

• Synthesis of unsaturated fatty acids requires the activity of a number of desaturases

• Animal desauturases form double bonds as much as 9 carbons removed from the carboxyl

end of a fatty acid

• Only plant desaturases form double bonds positioned farther than 9 carbons from the

carboxyl end; thus fatty acids such as linoleate (18:2∆9,12) must be acquired in the diet of animals

and are considered essential fatty acids

Linoleate

arachidonic acid (C20:4∆5,8,11,14)

leukotrienes

lipoxygenase

trigger allergic responses

eicosanoids

cyclooxygenase

prostaglandins (smooth muscle contraction; pain; inflammation)

Linoleate

arachidonic acid (C20:4∆5,8,11,14)

leukotrienes

lipoxygenase

trigger allergic responses

eicosanoidsprostaglandins (smooth muscle contraction; pain; inflammation)

cyclooxygenase

Aspirin

X

Cholesterol BiosynthesisCholesterol Biosynthesis

• Pathway substantially active only in liver cells

• All carbon atoms arise from acetyl-CoA

• Squalene, C30 linear hydrocarbon, is an intermediate

• Squalene is formed from 5 carbon units (isoprene)

H3C CH

2

H2C

C CH

Stage 1: Acetyl Co-A to Isopentenyl Pyrophosphate


2 Acetyl Co-A Acetoacetyl Co-Athiolase

HMG CoA

HMG CoA synthase

-OOC-CH2-C-CH2-CH2-OH

OH

CH3 Mevalonate

NADPHNADP+

HS-CoA2

2

HMG CoA reductaseHMG CoA reductase



2 Acetyl Co-A Acetoacetyl Co-Athiolase

HMG CoA

HMG CoA synthase


OH

CH3 Mevalonate

HMG CoA reductaseHMG CoA reductase

NADPHNADP+

HS-CoA

Lovostatin (Mevacor);Zocor

X


OH

CH3

melvalonate kinase

ATPADP, PiMevalonate

Phosphomevalonate kinase-OOC-CH2-C-CH2-CH2-OPO3

=

OH

CH3 PhosphomevalonateATP

ADP, Pi

-OOC-CH2-C-CH2-CH2-OP2O63-

OH

CH3Pyrophosphomevalonate

PyrophosphomevalonateDecarboxylase

ATP

ADP, Pi CO2

Isopentenyl pyrophosphateIsopentenyl pyrophosphate

CH2=C-CH2-CH2-OP2O63-CH2=C-CH2-CH2-OP2O63-

CH3CH3

isomerase

CH3-C=CH-CH2-OP2O63-CH3-C=CH-CH2-OP2O63-

CH3CH3

Dimethylallyl pyrophosphateDimethylallyl pyrophosphate

Stage 2: Isopentenyl Pyrophosphate to Squalene

Stage 2: Isopentenyl Pyrophosphate to Squalene

Isopentenyl pyrophosphate + Dimethylallyl pyrophosphate

prenyl transferase

condense head to tail

PPi

OP2O

63-

Geranyl pyrophosphate

(C10)

OP2O

63- OP

2O

63-

+Geranyl pyrophosphate

(C10)

Isopentenyl pyrophosphate

(C5)

OP2O

63-

Farnesyl pyrophosphate

(C15)

prenyl transferase

PPi

OP2O

63-

2 Farnesyl pyrophosphates

condense head to head

2 PPi

NADPHNADP+

Reaction catalyzed by Squalene

synthase

Product is Squalene (C30) !

Stage 3: Squalene to CholesterolStage 3: Squalene to Cholesterol

• Numerous steps involving addition of oxygen (to form squalene 2,3 epoxide) and closure of rings

• Lanosterol is the first intermediate containing all four fused rings; it accumulates in large amounts

in cells actively synthesizing cholesterol

• Lanosterol is converted to cholesterol in 20 more reactions

• Squalene 2,3 epoxide is precursor to plant sterols

Cholesterol metabolism is a source of a large number of other cellular constituents:

Cholesterol metabolism is a source of a large number of other cellular constituents:

• Isopentenyl pyrophosphate is precursor to fat soluble vitamins (A, E, K), ubiquinone in

animal cells, plastoquinone and the phytol side chain of chlorophyll in plant cells, certain oils

such as musk, lemon, eucalayptus

• Cholesterol is precursor for bile salts, vitamin D, steroid hormones, mineralocorticoids, and

is also an important component of membranes.

Regulation of HMG-CoA Reductase:

• Phosphorylation by c-AMP dependent protein kinase inactivates.

• Gene expression: Cholesterol levels control the amout of mRNA. If [cholesterol] is high, mRNA levels are reduced. If [cholesterol] is low, mRNA is increased.

• Degradation of the enzyme: Half-life depends upon the cholesterol level; high cholesterol means a short half-life.

Lipoproteins: Good vs. Bad Cholesterol

Phospholipids, triacylglycerols and cholesterol circulate in blood in the form of lipoproteins.Phospholipids, triacylglycerols and cholesterol circulate in blood in the form of lipoproteins.

Classified according to the relative amounts of lipid and protein in the complex (the more protein and less lipid the denser the complex).

Classified according to the relative amounts of lipid and protein in the complex (the more protein and less lipid the denser the complex).

Thus, we have high-density lipoproteins (HDL); low-density lipoproteins (LDL); intermediate-density lipoproteins (IDL); very-low density lipoproteins (VLDL) and chylomicrons.

Thus, we have high-density lipoproteins (HDL); low-density lipoproteins (LDL); intermediate-density lipoproteins (IDL); very-low density lipoproteins (VLDL) and chylomicrons.

All lipoproteins consist of a core of triacylglycerols or cholesterol esters surrounded by a single phospholipid layer into which is inserted a mixture of cholesterol and proteins. The proteins serve as recognition sites for lipoprotein receptors throughout the body.

HDL and VLDL are synthesized primarily in the ER of the liver; chylomicrons are synthesized in the intestine.

LDL is synthesized from VLDL and is the major circulatory complex for transport of cholesterol and cholesterol esters from liver to other tissues.

Chylomicrons transport triacylglycerols mostly (and some cholesterol esters) from intestines to other tissues.

VLDL released in to the bloodstream is converted to IDL and LDL by the action of lipases which cleave triacylglycerols.

The half-life of LDL is about 24 hours. It is removed from circulation by endocytosis in coated vesicles and degradation by lysosomal acid lipases.

HDL has a half-life of 5-6 days. Newly formed HDL contains no cholesterol esters. These accumulate over time through the action of enzymes that transfer cholesterol esters. HDL functions to return cholesterol and cholesterol esters to the liver and hence remove them from circulation. Hence, HDL is the “good cholesterol”. LDL, because it is the major circulating form of cholesterol is the “bad cholesterol” correlated with increased risk of cardiovascular disease.

Receptor-Mediated Endocytosis

Discovered by Brown and Goldstein (1985 Nobel Prize).Discovered by Brown and Goldstein (1985 Nobel Prize).

ApoB-100, the major protein in LDL particles, is recognized by the LDL receptor.ApoB-100, the major protein in LDL particles, is recognized by the LDL receptor.

Binding of LDL to the LDL receptor initiates endocytosis which brings LDL and its receptors into the cell inside an endosome.

Binding of LDL to the LDL receptor initiates endocytosis which brings LDL and its receptors into the cell inside an endosome.

Endosomes fuse with lysosomes, which contain enzymes that hydrolyze cholesterol esters, releasing cholesterol and fatty acids into the cytosol.

Endosomes fuse with lysosomes, which contain enzymes that hydrolyze cholesterol esters, releasing cholesterol and fatty acids into the cytosol.

ApoB-100 is degraded to amino acids, but the LDL receptor is recycled back to the cell surface.ApoB-100 is degraded to amino acids, but the LDL receptor is recycled back to the cell surface.

Cholesterol entering the cell in this manner is used to synthesize membranes, or is reesterified for storage in cytosolic lipid droplets.

Cholesterol entering the cell in this manner is used to synthesize membranes, or is reesterified for storage in cytosolic lipid droplets.

Defective LDL receptors result in the genetic disease familial hypercholesterolemia. This is characterized by the inability of cells to take up cholesterol; hence blood cholesterol levels are extremely high.

Defective LDL receptors result in the genetic disease familial hypercholesterolemia. This is characterized by the inability of cells to take up cholesterol; hence blood cholesterol levels are extremely high.

The excess blood cholesterol accumulates and contributes to the formation of atherosclerotic plaques. Heart failure from atherosclerosis is the leading cause of death in industrialized societies.

The excess blood cholesterol accumulates and contributes to the formation of atherosclerotic plaques. Heart failure from atherosclerosis is the leading cause of death in industrialized societies.

Clathrin

Coated pit

Receptor with ligandAdaptor

Ketone Body Formation; Fatty acid and Cholesterol...

Documents

Transcript of Ketone Body Formation; Fatty acid and Cholesterol...