LIPOLYSIS, BETA OXIDATION, KETONES, LIPOGENESIS

BIOC 460DR. TISCHLER

LECTURE 31

OBJECTIVES

1. For the lipolytic pathway (lipolysis): describe the pathway, identify where it occurs, name the principal enzyme involved, and explain the role of albumin and fatty acid binding protein in the transport and metabolism of free fatty acids liberated by lipolysis

2. For the degradation of fatty acyl CoAs: describe the roles of acyl CoA synthetase, carnitine-palmitoyl transferases (CPT-I and CPT-II),

and carnitine acylcarnitine translocase (CAT); discuss the relationship of products of the -oxidation pathway to energy production.

3. For ketone body metabolism: identify where and when ketone body formation (ketogenesis) occurs, state the role of ketogenesis, identify

where ketone oxidation occurs and explain why normally individuals do not develop ketoacidosis even when producing ketone bodies.

OBJECTIVES

4. Describe the reactions catalyzed by malic enzyme and acetyl CoA carboxylase

5. For the fatty acid synthase reaction: list the substrates and key products, identify the sources of NADPH for the reaction, and describe its

general mechanism.

6. Describe how fatty acids are stored as a source of fuel during starvation or stress.

PHYSIOLOGICAL PREMISE

Would you believe that diabetics having a ketotic crisis have actually been arrested for DUIs even though they have consumed no alcohol? Indeed a blood analysis would show no alcohol. Why would this occur? During a ketotic crisis a byproduct of the excess ketone production is acetone. Having nowhere else to go, it is expired through the lungs. It is the acetone that arresting officers have smelled on the breath of these individuals and despite their protestations have innocently believed them to be consuming alcohol.

LIPOLYSIS OF STORED TRIACYLGLYCEROL

fatty acids hydrolytically cleaved from triacylglycerol

largely in adipose to release fatty acids as a fuel

may also occur in muscle or liver - smaller amounts of fatty acids are stored

hormone-sensitive (cyclic AMP-regulated) lipase initiates lipolysis – cleaves first fatty acid

this lipase and others remove remaining fatty acids

fatty acids/glycerol released from adipose to the blood

hydrophobic fatty acids bind to albumin, in the blood, for transport

LipolysisTriacylglycerol Glycerol + Fatty acids

MITOCHONDRION

cell membraneFA = fatty acidLPL = lipoprotein lipaseFABP = fatty acid binding protein

ACS

FABP

FABPFA

[3]

FABPacyl-CoA

[4]

CYTOPLASM

CAPILLARY

FAalbuminFA FA

FA

fromfatcell

FA

[1]

acetyl-CoA TCAcycle

-oxidation[6]

[7]

carnitinetransporter

acyl-CoA[5]

Figure 1. Overview of fatty acid degradation

ACS = acyl CoA synthetase

LPL

Lipoproteins(Chylomicrons or VLDL)

[2]

Figure 2 (top). Activation of palmitate to palmitoyl CoA (step 4, Fig. 1) and conversion to palmitoyl carnitine

IntermembraneSpace

OUTERMITOCHONDRIALMEMBRANE

palmitoyl-carnitine

CoApalmitoyl-CoA

carnitine

Cytoplasm

palmitoyl-CoA

AMP + PPiATP + CoA

palmitate

CPT-I [2]

ACS

[1]

CPT-I defects cause severe muscle weakness because fatty acids are an important muscle fuel during exercise.

Figure 2 (bottom). Mitochondrial uptake via of palmitoyl-carnitine via the carnitine-acylcarnitine translocase (CAT) (step 5 in Fig. 1).

Matrix

INNERMITOCHONDRIALMEMBRANE

Intermembrane Space

palmitoyl-carnitinecarnitine

CoApalmitoyl-CoA

CAT [3]

palmitoyl-carnitineCPT-II

carnitine

CoApalmitoyl-CoA

[4]

CPT-I

CAT

IntermembraneSpace

OUTERMITOCHONDRIALMEMBRANE

palmitoyl-carnitine

CoA

carnitine

Cytoplasmpalmitoyl-CoA

AMP + PPiATP + CoA

palmitate

palmitoyl-CoA

Matrix

INNERMITOCHONDRIALMEMBRANE

[3]

palmitoyl-carnitinecarnitine

CoApalmitoyl-CoA

[4]

CPT-I [2]

ACS[1]

CPT-II

Figure 3. Processing and -oxidation of palmitoyl CoA

matrix side

inner mitochondrialmembrane

2 ATP3 ATP

respiratory chain

recycle6 times

Carnitinetranslocase

Palmitoylcarnitine

Palmitoylcarnitine

Palmitoyl-CoA

+ Acetyl CoACH3-(CH)12-C-S-CoA

O

oxidationFAD

FADH2

hydration H2O

thiolase CoA

oxidationNAD+

NADH

Citricacid cycle 2 CO2

Figure 4. Ketone body formation (ketogenesis) in liver mitochondria from excess acetyl CoA derived from the -oxidation of fatty acids

MITOCHONDRION

(excess acetyl CoA)

Hydroxymethylglutaryl CoA

HMG-CoA synthaseacetyl CoA

CoA

Acetoacetate

HMG-CoA-lyase

acetyl CoA

-Hydroxybutyrate

-Hydroxybutyratedehydrogenase

NAD+

NADH

Acetone

(non-enzymatic)

2 Acetyl CoAFatty acid-oxidation

Citric acid cycle

oxidation to

CO2

Acetoacetyl CoACoA

Thiolase

high rates of lipolysis (e.g., long‑term starvation or in uncontrolled diabetes) produce sufficient ketones in the blood to be effective as a fuel

ketones are the preferred fuel if glucose, ketones, fatty acids all available in the blood

primary tissues: using ketones, when available, are brain, muscle, kidney and intestine, but NOT the liver.

-Hydroxybutyrate + NAD+ acetoacetate + NADH -hydroxybutyrate dehydrogenase in mitochondria;

reverse of ketogenesis

KETONE BODY OXIDATION

XAdiposeTissue Free fatty

acidsLiver

Ketone BodiesInsulin

Pancreas

Figure 5. Mechanism for prevention of ketosis due to excess ketone body production that can lead to ketoacidosis

KETOSISExcessive build-up of ketone bodies results in ketosis eventually

leading to a fall in blood pH due to the acidic ketone bodies.

LIPOGENESIS

principally in adipose tissue and liver

lipogenesis – cytoplasm; requires acetyl CoA

adipose: FA stored as triacylglycerols via esterification

liver: produces TAG packaged into VLDL and exported

compounds metabolized to acetyl CoA can serve as a fat precursor

glucose = primary source of carbons for fat synthesis.

LIPOGENESIS

CYTOPLASM MITOCHONDRIAL MATRIX

Pyruvate

Citrate

CSOxaloacetate

PCATP, CO2

ADP, Pi

PPP

Pyruvate

Glucose

Glycolysis

FAS

FattyAcids

Citrate

Acetyl CoA

CLATP, CoA

ADP+Pi

Oxaloacetate ACCADP, Pi

CO2, ATP

Malonyl CoA

Acetyl CoA

NAD, CoA NADH, CO2

PDH

MDH

NADH

NAD+Malate

MENADP+

NADPH

CO2

Figure 6. Export of acetyl CoA as citrate for fatty acid biosynthesis, generation of NADPH and pathway of lipogenesis. (similar to diagram discussed for cholesterol synthesis exxept this involves PDH reaction)

KEY MITOCHONDRIAL REACTIONS

PYRUVATE CARBOXYLASE

pyruvate + CO2 + ATP oxaloacetate + ADP + Pi

PYRUVATE DEHYDROGENASE

pyruvate + NAD+ + coenzyme A (CoA) acetyl CoA + CO2 + NADH

Citrate Lyasecitrate + CoA + ATP acetyl CoA + oxaloacetate + ADP + Pi

Malate dehydrogenaseoxaloacetate + NADH malate + NAD+

Malic Enzymemalate + NADP+ pyruvate + NADPH

KEY CYTOPLASMIC REACTIONS INDIRECTLY NEEDED FOR LIPOGENESIS

KEY CYTOPLASMIC REACTIONS DIRECTLY NEEDED FOR LIPOGENESIS AND FATTY ACID ACTIVATION

Acetyl CoA Carboxylase: acetyl CoA + HCO3

- + ATP malonyl CoA + ADP + Pi

Fatty Acid Synthase: acetyl CoA + 7 malonyl CoA + 14 NADPH + 14 H+ palmitate + 7 CO2 + 8 CoA + 14 NADP+

Acyl CoA Synthetase: (also used for fatty acids other than palmitate) palmitate + ATP + CoA palmitoyl CoA + AMP + PPi

condensation

reductiondehydration

reduction

2 NADPH 2 NADP+

ACP

CEacp

ACP

CEacp

ACP

CEacp

Figure 7. General mechanism for the fatty acid synthase reaction. CE is condensing enzyme. ACP is acyl carrier protein. This row represents the initial steps for priming the reaction with acetyl CoA and the addition of two carbons from malonyl CoA.

COO-

C=O

CH2

C=O

CH2

C=O

CH3

C=O

CH2

CH2

CH3malonyl CoA

CH3

C=O

acetyl CoA

CO2

CO2

CO2

CO2

CH3

C=O

CH3

C=OC=O

CH2

CH3

C=O

C=O

CH2

CH3

C=O

C=O

CH2

CH3

C=O

4-Cunit

Figure 7. General mechanism for the fatty acid synthase reaction. CE is condensing enzyme. ACP is acyl carrier protein. This row depicts a typical cycle of adding two more carbons to the fatty acid chain.

malonyl CoA

condensation

CO2

reductiondehydration

reduction

2 NADPH 2 NADP+

ACP

CEacp

6-Cunit

ACP

CEacp

6-Cunit

ACP

CEacp

4-Cunit

Figure 7. General mechanism for the fatty acid synthase reaction. CE is condensing enzyme. ACP is acyl carrier protein. This row shows the release of the finished product, palmitate, through cleavage by thioesterase.

malonyl CoA

ACP

CEacp

16-Cunit

palmitate

ACP

CEacp

6-Cunit

5 more cycles adding 10 more carbons

5CO2

10NADP+

5malonyl CoA10NADPH

ACP

CEacp

thioesterasecleavage

palmitate

malic enzyme:

Malate + NADP+ Pyruvate + CO2 + NADPH

pentose phosphate pathway:

Glucose-6-P + 2 NADP+ Ribulose-5-P + 2 NADPH + CO2

Sources of NADPH for the Biosynthesis of Fatty Acids.

Figure 8. Formation of phosphatidic acid from glycerol-3-P or DHAP, and its conversion to triacylglycerol

Lysophosphatidic acid

Phosphatidic acid

Triacylglycerol

NADPH

NADP+

Diacylglycerolphosphatase

CoA

Acyldihydroxyacetone phosphate

fatty acyl CoA

Dihydroxyacetone phosphate

fatty acyl CoA

CoA

ADP

ATP glycerolkinase

Glycerol-3-P

Glycerol

CoA

fatty acyl CoA

CoA

fatty acyl CoA

Pi