MTY1107 : Lipid Metabolism

MTY1107 Laboratory Finals Topic 1

Stomach

Where lipid digestion starts with the action of gastric lipase, breaking down triglycerides into diglycerides and free fatty acids.

By itself, it cannot break down fats

Gastric Lipase

Initiates the hydrolysis of triacylglycerols to mono/diacylglycerols, cleaves bond linking glycerol to fatty acid

In the stomach

Small Intestine (Duodenum)

Is the primary site for lipid digestion and absorption, where bile salts and pancreatic lipase further break down lipids into fatty acids and monoglycerides.

Pancreatic Lipase

An enzyme secreted by the pancreas that catalyzes the hydrolysis of dietary fats into free fatty acids and monoglycerides in the small intestine.

Further degrade triacylglycerols

Bile Salts

Compounds derived from cholesterol that emulsify fats in the intestine, aiding in their digestion and absorption.

Secreted by the Gallbladder, emulsifies the mono/diacylglycerols and free fatty acids to form mixed micelles

Micelles

Aggregates of lipid molecules formed in the intestine that facilitate the absorption of fatty acids and monoglycerides by transporting them to the intestinal mucosa.

Small Intestine (Mucosa)

The innermost layer of the small intestine that is involved in nutrient absorption, featuring villi and microvilli to increase surface area.

Apolipoproteins

Proteins that bind to lipids to form lipoproteins, playing key roles in lipid transport and metabolism.

Inactive lipoproteins that reform the degraded free fatty acids and mono/diacylglycerols into triglycerides for storage.

Bind the reformed triacylglycerols with cholesterol to form Chylomicrons and are absorbed

Chylomicrons

Are lipoprotein particles that transport dietary lipids from the intestines to other locations in the body, particularly adipose tissue.

Triacylglycerol + Cholesterol

Lymphatic System (Mucosa)

The absorbed chylomicrons are transported to the __________ and are carried via the blood to various organs.

Lipoprotein Lipase

An enzyme that hydrolyzes triglycerides in lipoproteins into free fatty acids and glycerol, facilitating the uptake of fatty acids by tissues.

Breaks the Chylomicrons and hydrolyzes the triacylglycerols inside them to free fatty acids and Glycerol.

In Muscles or Adipocytes

Uptake of fatty acids for energy production or storage can be

In Muscles

The fatty acids are oxidized for energy

In Adipocytes

The fatty acids are re-esterified to triacylglycerols for storage

Triacylglycerols

Storage form of fats

Stage 1 : Lipolysis and Activation

Stage 2 : Mitochondrial Transport

Stage 3 : Beta Oxidation of Fatty Acids

Stages of Lipolysis and B-Oxidation

Lipolysis and B-Oxidation

How body produces energy using triacylglycerols

Lipolysis

The hydrolysis of Triacylglycerols to Glycerol and Fatty Acids using Lipase

Happens primarily in the Duodenum

Lipase

A hydrolase enzyme needed for the hydrolysis of triacylglycerols to glycerol and fatty acids

Used alongside 3 mol of H2O

Glycerol Metabolism

5% of the available energy in Triacylglycerols is stored in Glycerol

Glycerol is transported to the Liver and is phosphorylated in a two-step reaction for it to enter Glycolysis

1. Glycerol Kinase

2. Glycerol-3-Phosphate

Glycerol Metabolism begins with glycerol being phosphorylated by the enzyme ____ to form ____.

1. Dihydroxyacetone Phosphate (DHAP)

2. Glycerol-3-Phosphate Dehydrogenase

Glycerol-3-phosphate can then be converted into _______ by the enzyme _______.

1. Gluconeogenesis

2. Glycolysis

DHAP can enter ____ to form glucose or enter _____ to generate pyruvate, which can then be used for ATP production or as a precursor for fatty acid synthesis.

Phosphate Isomerase

After glycerol is converted to glycerol-3-phosphate and then to dihydroxyacetone phosphate (DHAP), ______ catalyzes the reversible interconversion between DHAP and glyceraldehyde-3-phosphate (G3P).

Activation

95% of the available energy available in Triacylglycerols is stored in Fatty acids.

Once the fatty acids have been freed through lipolysis, they are transported to target tissues for oxidation (degradation).

The fatty acids enter the cell through Fatty Acid Transporter.

The Fatty acid transporter also converts Fatty Acids to Fatty Acyl-CoA.

The conversion needs ATP and CoA-SH

Fatty Acyl-CoA Synthetase

• Is an enzyme that activates fatty acids in the cytoplasm by converting them into fatty acyl-CoA.

• This activation is a necessary step for beta-oxidation and involves two key reactants:

  • ATP (providing energy)

  • CoA-SH (coenzyme A)

• The reaction forms a high-energy thioester bond between the fatty acid and CoA, creating fatty acyl-CoA.

• The activated fatty acyl-CoA is then ready to be transported into the mitochondria (via the carnitine shuttle) for beta-oxidation, where it will be broken down into acetyl-CoA to enter the citric acid cycle for ATP production.

Carnitine Shuttle

Fatty acyl-CoA cannot directly cross the mitochondrial membrane, so the ______ is required to transport long-chain fatty acids into the mitochondria for beta-oxidation.

While fatty acyls with short chain lengths (<12 Carbon units) can enter the Mitochondria freely, most dietary fatty acyls have >14 carbon units.

For long chain fatty acyls to be able to enter the Mitochondria, they need to undergo the enzymatic reactions of the _________.

Step 1 : Conversion of the Fatty Acyl-CoA to Acylcarnitine

Step 2 : Entry of Acylcarnitine Inside the Mitochondrial Matrix and Re-conversion to Fatty Acyl-CoA

2 Steps of the Carnitine Shuttle

Step 1 : Conversion of Fatty Acyl-CoA to Acylcarnitine

• The enzyme carnitine acyltransferase I (CAT-I), located on the outer mitochondrial membrane, transfers the fatty acyl group from fatty acyl-CoA to carnitine, forming acylcarnitine.

• This allows the fatty acid to be carried across the mitochondrial membrane inside the Intermembrane Space because acylcarnitine is able to pass through, whereas fatty acyl-CoA cannot.

Carnitine Acyltransferase I (CAT-I)

• located on the outer mitochondrial membrane.

• its main role is to transfer the fatty acyl group from fatty acyl-CoA to carnitine, forming acylcarnitine.

• this step is essential for the transport of fatty acids into the mitochondria, as acylcarnitine can pass through the mitochondrial membrane, while fatty acyl-CoA cannot.

Carnitine-Acylcarnitine Translocase I

• is a membrane transporter located in the inner mitochondrial membrane.

• it facilitates the exchange of acylcarnitine (from the cytoplasm) with carnitine (from the mitochondrial matrix), allowing acylcarnitine to enter the matrix and carnitine to exit the mitochondrion.

• this exchange is crucial for maintaining the carnitine pool inside the mitochondria and enabling the transport of fatty acids for beta-oxidation.

Step 2: Entry of Acylcarnitine into the Mitochondrial Matrix and Re-conversion to Fatty Acyl-CoA

• Acylcarnitine is transported into the mitochondrial matrix via the carnitine-acylcarnitine translocase I.

• Once inside the matrix, the enzyme carnitine acyltransferase II (CAT-II) removes the carnitine group and transfers the fatty acyl group back to CoA-SH, regenerating fatty acyl-CoA.

• The fatty acyl-CoA can now undergo beta-oxidation inside the mitochondria to generate ATP.

Carnitine Acyltransferase II (CAT-II)

• is located on the inner mitochondrial membrane, within the mitochondrial matrix.

• catalyzes the conversion of acylcarnitine back to fatty acyl-CoA by transferring the fatty acyl group from carnitine to CoA-SH.

• this is the final step before the fatty acyl-CoA enters beta-oxidation, where it will be broken down to generate energy.

Beta Oxidation or Fatty Acyl Degradation

• Is the process by which fatty acyl-CoA molecules are broken down into acetyl-CoA units in the mitochondria.

• This process generates NADH and FADH₂, which are used in the electron transport chain to produce ATP.

• The acetyl-CoA produced enters the citric acid cycle (Krebs cycle) for further energy production.

• In Eukaryotes, all enzymes used in this process is found in the Mitochondria

STEP1: Oxidation

STEP2: Hydration

STEP3: Oxidation

STEP4: Cleavage

4 Steps of Beta Oxidation

STEP 1: OXIDATION – The Conversion of the Single Bond to a trans-Double Bond

• In the first step of beta-oxidation, the enzyme Acyl-CoA dehydrogenase catalyzes the dehydrogenation of fatty acyl-CoA.

• This results in the formation of a trans-Δ²-enoyl-CoA intermediate, and FADH₂ is produced.

• Acyl-CoA dehydrogenase is specific for different chain lengths of fatty acids, such as short-, medium-, long-, and very long-chain fatty acids.

STEP 2: HYDRATION – Addition of Water Across the trans-Double Bond

• In the second step, the enzyme Enoyl-CoA hydratase adds water (hydration) to the trans-Δ²-enoyl-CoA intermediate.

• This forms L-β-hydroxyacyl-CoA, a hydroxylated fatty acyl intermediate.

• This step is essential for setting up the molecule for the subsequent oxidation step.

STEP 3: OXIDATION – Addition of Water Across the trans-Double Bond

• In the third step, the enzyme B-Hydroxyacyl-CoA dehydrogenase catalyzes the oxidation of L-β-hydroxyacyl-CoA to form β-ketoacyl-CoA.

• Hydroxyl is oxidized to a carbonyl group.

• This step produces NADH2 as a by-product, which will be used in the electron transport chain for ATP generation.

STEP 4: CLEAVAGE – Cleavage of the ɑ–β bond to produce Acetyl-CoA and a Shortened Fatty Acyl

• In the final step, the enzyme acyl-CoA acetyltransferase Thiolase catalyzes the cleavage of the a-B bond of β-ketoacyl-CoA by adding CoA-SH (Coenzyme A).

• This forms a shortened fatty acyl-CoA (which is 2 carbon atoms shorter than the original fatty acid) and acetyl-CoA.

• The acetyl-CoA produced enters the citric acid cycle (Krebs cycle) for further energy production.

How does the beta-oxidation cycle repeat, and what is the final outcome?

• The beta-oxidation cycle repeats until the fatty acyl-CoA chain is fully broken down into acetyl-CoA units.

• Each cycle shortens the fatty acid chain by 2 carbon atoms, and for each cycle, 1 FADH₂, 1 NADH, and 1 acetyl-CoA are produced.

• The process continues until the entire fatty acid is converted into acetyl-CoA, which is then used in the citric acid cycle for further ATP production.

β-Oxidation of Monounsaturated Fatty Acids

• Have a cis double bond at the Δ⁹ position (between the 9th and 10th carbon).

• Step 1: In the first round of β-oxidation, the double bond is at the wrong position for the typical β-oxidation process.

• Additional Step: The enzyme enoyl-CoA isomerase rearranges the cis double bond to a trans configuration at the Δ² position.

• Once the bond is isomerized, the fatty acid proceeds through the usual β-oxidation cycle, involving the same enzymes (Acyl-CoA dehydrogenase, enoyl-CoA hydratase, hydroxyacyl-CoA dehydrogenase, and thiolase).

Δ²,Δ³-enoyl-CoA isomerase

• Catalyzes the isomerization of cis-Δ³ double bonds in monounsaturated fatty acids to trans-Δ² double bonds.

• This isomerization is crucial because β-oxidation requires a trans-Δ² configuration at the double bond to proceed through the standard β-oxidation steps.

• The enzyme ensures that unsaturated fatty acids with cis-Δ³ bonds can continue through β-oxidation, enabling the complete breakdown of these fatty acids.

β-Oxidation of Polyunsaturated Fatty Acids

• Require additional enzymatic steps during β-oxidation due to their multiple cis double bonds.

• Since most unsaturated fatty acids have cis doible bonds, enoyl-CoAhydratse cannot act on it.

• The cis double bond must first be converted to the trans configuration using the enzyme Δ²,Δ³-enoyl-CoA isomerase.

• After conversion, Steps 1-4 is recontinued producing 1 Acetyl-CoA.

• Another cycle is started stopping at Step 1.

2,4-dienoyl-CoA reductase

In β-Oxidation of Polyunsaturated Fatty Acids, the second double bond is reduced using __________ and NADPH

This enzyme reduces the conjugated double bonds to a single double bond at Δ⁴ or Δ⁶.

Enoyl-CoA isomerase

In β-Oxidation of Polyunsaturated Fatty Acids, the enzyme _______ acts again to move the double bond into the correct position for β-oxidation (trans-Δ²).

This moves the double bond to C-2, the product is now free to proceed to 5 more cycles of Steps 1-4.

Tricarboxylic acid (TCA)

Ketogenesis

Fates of Acetyl-CoA

Citric Acid Cycle

Acetyl-CoA enters the _________ where it combines with oxaloacetate to form citrate. This cycle generates NADH, FADH₂, and GTP/ATP, which are used in the electron transport chain to produce ATP.

Ketogenesis

• When glucose is low (e.g., during fasting or prolonged exercise), excess acetyl-CoA is converted into ketone bodies (acetoacetate, β-hydroxybutyrate, and acetone) in the liver.

• These ketone bodies are transported to other tissues (like the brain and muscles) where they can be used as an alternative energy source.

• These ketone bodies are used by the body as alternative sources of fatty acids during starvation, fasting, uncontrolled Diabetes mellitus I, and even sleep.

• Aside from the Liver, the Brain can also produce Ketone bodies as source of energy in extreme cases of starvation.

Acetoacetate, Acetone, D-β-Hydroxybutyrate

3 Ketone Bodies

Condensation of Acetyl-CoA

Formation of HMG-CoA

Cleavage of HMG-CoA

Reduction to β-Hydroxybutyrate

4 Steps of Ketogenesis

Condensation of Acetyl-CoA

Two molecules of acetyl-CoA are condensed by the enzyme thiolase to form acetoacetyl-CoA.

Formation of HMG-CoA

Acetoacetyl-CoA reacts with another molecule of acetyl-CoA in the presence of the enzyme HMG-CoA synthase to form 3-hydroxy-3-methylglutaryl-CoA (HMG-CoA)

Cleavage of HMG-CoA

HMG-CoA is broken down by HMG-CoA lyase, releasing acetoacetate (the primary ketone body) and acetyl-CoA.

Reduction to β-Hydroxybutyrate

Acetoacetate can be spontaneously decarboxylated to form acetone (which is exhaled), or it can be reduced by β-hydroxybutyrate dehydrogenase to form β-hydroxybutyrate, another key ketone body.

Ketosis

Diabetic individuals have a characteristic fruity-smelling breath due to excess ketone bodies, particularly acetoacetate as a product of ______.

Skeletal Muscle and Heart

Peripheral tissues such as ______ can use ketone bodies as alternative sources of energy.

Lipogenesis

• Is the process by which the body synthesizes fatty acids and triglycerides from smaller molecules like acetyl-CoA and glycerol-3-phosphate.

• It occurs mainly in the liver and adipose tissue, especially when there is an excess of carbohydrates.

• The key steps involve converting acetyl-CoA into malonyl-CoA (the rate-limiting step), elongating fatty acid chains, and then esterifying them with glycerol to form triglycerides for storage in fat cells.

Stage 1 : Cytosolic Transport

Stage 2 : Conversion of Acetyl-CoA to Malonyl-CoA

Stage 3 : Fatty Acid Biosynthesis

3 Stages of Lipogenesis

Stage 1 : Cytosolic Transport

• Involves the Citrate Shuttle

• Acetyl-CoA needs to be transported outside the Mitochondria

• To circumvent this, Acetyl-CoA is transported to the Cytosol in the form of Citrate

Cytosol

Where lipogenesis occurs

Citrate Shuffle

Using Citrate Transport Protein (CTP), Citrate is transported outside the Mitochondria

Once in the Cytosol, Citrate is converted to Acetyl-CoA using the enzyme ATP-Citrate lyase

The conversion of Citrate to Acetyl-CoA needs ATP and CoA-SH

Citrate Transport Protein (CTP)

located in the inner mitochondrial membrane.

It facilitates the transport of citrate from the mitochondrial matrix to the cytoplasm.

ATP-citrate lyase

located in the cytoplasm.

It cleaves citrate into acetyl-CoA and oxaloacetate using energy from ATP.

STAGE 2: Conversion of Acety-CoA to Malonyl-CoA

First step in fatty acid biosynthesis

Catalyzed by the biotin-dependent Acetyl-CoA carboxylase

Acetyl-CoA is carboxylated by acetyl-CoA carboxylase (ACC) to form malonyl-CoA.

This is the rate-limiting and committed step in fatty acid synthesis.

STAGE 3: Fatty Acid Biosynthesis

Made up of 4 sequential reactions : Condensation, Reduction, Hydration, and Reduction

Occurs under lipogenic conditions such as after eating and low fat-high carbohydrate diets

The ultimate goal is to create fatty acids from acetyl-CoA

Acyl Carrier Protein (ACP)

After the first step, Malonyl-CoA is activated by being anchored to the ________.

Fatty Acid Synthase

The four subsequent reactions of fatty acid synthesis is catalyzed by enzymes attached to the multi-enzyme complex ________.

Condensation - Combination of Malonyl-ACP and Acetyl-ACP

Two carbons from Malonyl-ACP is combined with Acetyl-ACP, extending the acyl chain

CO2 is released

Forms Acetoacetyl-ACP

Acetoacetyl-ACP

Product of Condensation reaction in Fatty Acid Biosynthesis

Reduction - Reduction of the Carbonyl Group at the β Carbon

The carbonyl group at the β Carbon of Acetoacetyl-ACP is reduced

NADH is oxidized

Forms β-Hydroxybutyryl-ACP

β-Hydroxybutyryl-ACP

Product of first Reduction reaction in Fatty Acid Biosynthesis

DEHYDRATION – Removal of Water and Formation of a trans Double Bond

Water is removed from β-Hydroxybutyryl-ACP

The removal of water allows the formation of a trans double bond

a,β-trans-Butenoyl-ACP is formed

a,β-trans-Butenoyl-ACP

Product of Dehydration reaction of Fatty Acid Biosynthesis

REDUCTION – Reduction of the trans Double Bond

The trans double bond is reduced forming a saturated fatty acyl

NADH is oxidized

The fatty acyl, Butyryl-ACP is formed

Butyryl-ACP

Product of the second Reduction reaction of Fatty Acid Biosynthesis