Lecture 16 biochem

Complete Hydrolysis of Triacylglycerol

• Complete hydrolysis yields:
  • Glycerol molecule
  • Fatty acids (number not specified but implied to be three based on triacylglycerol structure)

Chapter 25: Lipid Metabolism

• Overview of metabolism comparing triacylglycerols, carbohydrates, and proteins:
  • Triacylglycerols are broken down into fatty acids and glycerol.
  • Carbohydrates are broken down into monosaccharides.
  • Proteins are broken down into amino acids.
• These building blocks then feed into:
  • Glycolysis
  • Fatty acid oxidation
  • Amino acid catabolism
• Which all converge on Acetyl CoA, feeding into:
  • Citric acid cycle
  • Reduced coenzymes
  • Electron transport chain and oxidative phosphorylation

Major Difference: Digestion

• Dietary triacylglycerols require a more complex system to reach the bloodstream due to solubility.

Digestion of Triacylglycerols (TAGs)

• 1. Mouth:
  • Saliva has no effect on digestion.
• 2. Stomach:
  • Churning action produces small fat droplets (chyme).
  • Gastric lipases hydrolyze some (10%) TAGs to monoacylglycerols.
• 3. Small Intestine:
  • Bile solubilizes fat droplets.
  • Pancreatic lipases produce monoacylglycerols, which form fatty acid micelles.
• 4. Intestinal Cells:
  • Monoacylglycerols in micelles are repackaged into TAGs, forming chylomicrons.
• 5. Lymphatic System:
  • Chylomicrons are transported to the bloodstream via the lymphatic system.
• 6. Bloodstream:
  • TAGs in chylomicrons are hydrolyzed to free fatty acids, mediated by lipoprotein lipases.

Digestion - Detailed Explanation

• Stomach:
  • Physical Change: TAGs are converted into chyme (semiliquid material).
  • Chemical Change: Gastric lipases hydrolyze approximately 10% of TAGs, breaking ester bonds.
• Small Intestine:
  • Physical Change: Bile (released by the gallbladder) and bile salts emulsify the fat droplets.
  • Chemical Change: Pancreatic enzymes (lipases) continue to hydrolyze ester bonds.

TAG Hydrolysis & Micelle Formation

• Incomplete TAG hydrolysis results in nonpolar tails forming a nonpolar oil droplet.
• Fatty acid micelles are formed, which are small enough to be absorbed by intestinal cells.

Intestinal Cell Processing

• Fatty acid micelles are repackaged back into TAGs within the intestinal cells, along with cholesterol and phospholipids, forming chylomicrons.

Chylomicrons

• Chylomicrons are lipoproteins that transport TAGs from intestinal cells to the bloodstream via the lymphatic system.
• Chylomicrons are too large to enter the bloodstream directly through capillary walls; they enter via the thoracic duct.

Bloodstream Changes

• In the bloodstream, lipoprotein lipases hydrolyze TAGs into glycerol and fatty acids.
• Glycerol can enter glycolysis.
• Fatty acids can be used for energy (beta-oxidation to Acetyl CoA) or stored.

TAG Storage

• TAGs are stored in adipose cells (adipocytes).
• Adipose tissue contains many adipocyte cells.

Regulation of TAG Metabolism

• Hormones regulate TAG (lipids) and carbohydrate metabolism.
• Three major hormones involved:
  • Insulin (low blood glucose)
  • Glucagon (high blood glucose)
  • Epinephrine (quick action)
• These hormones also regulate carbohydrates.

Triacylglycerol Mobilization

• Hydrolysis of TAGs stored in adipose cells/tissue, followed by the release of fatty acids and glycerol into the bloodstream.

Mechanism of Epinephrine Action

• Epinephrine binds to a receptor site on the cell membrane.
• This activates adenyl cyclase, which converts ATP to cyclic AMP (cAMP).
• cAMP activates hormone-sensitive lipase (HSL) by phosphorylation (HSL becomes HSL-P).
• Active HSL-P hydrolyzes triacylglycerols into fatty acids and glycerol.

Comparison of Stored Energy Reserves

• The lecturer will likely ask which energy reserve (TAGs, carbohydrates, or proteins) would last longer.

Metabolism of Glycerol

• Complete hydrolysis of TAG yields glycerol and fatty acids.
• Glycerol has 3 carbons.
• Glycerol can be used in glycolysis.

Glycerol Conversion to Glyceraldehyde-3-Phosphate

• Glycerol is converted into glyceraldehyde-3-phosphate, which is an intermediate in glycolysis.
• Two-step process:
  • Phosphorylation
  • Oxidation

Step #1: Phosphorylation

• Glycerol + ATP \rightarrow Glycerol-3-phosphate + ADP
• Enzyme: Glycerol kinase

Step #2: Oxidation

• Glycerol-3-phosphate + NAD^+ \rightarrow Dihydroxyacetone phosphate + NADH + H^+
• Enzyme: Glycerol 3-phosphate dehydrogenase

Fatty Acid Oxidation

• A 3-part process:
  • Activation (requires energy investment of ATP)
  • Transport (from outer mitochondrial membrane to matrix)
  • Beta-oxidation (repeated oxidation steps)

Activation of Fatty Acids

• Happens in the outer mitochondrial membrane.
• ATP \rightarrow AMP + 2 Pi (pyrophosphate).
• Energy expenditure to activate acyl (not acetyl).

Transport of Acyl CoA

• Large acyl group is transferred to carnitine to transport it into the mitochondrial matrix.

Beta-Oxidation Pathway

• Repeated β-oxidation involves cleavage at the α and β carbons.
• Acyl CoA is converted to Acetyl CoA, shortening the acyl chain by 2 carbons with each cycle.

4 Reaction Sequence of Beta-Oxidation

• Dehydrogenation
• Hydration
• Dehydrogenation (ketone formation)
• Thiolysis (reverse condensation)

Beta-Oxidation - Step 1: Dehydrogenation

• Alkane \rightarrow Alkene.
• Enzyme: Acyl CoA dehydrogenase.
• FAD is the reagent; it is reduced to FADH2.
• R-CH2-CH2-C(=O)-SCoA + FAD \rightarrow R-CH=CH-C(=O)-SCoA + FADH_2
• Only makes trans alkene.

Step #2: Hydration

• Alkene \rightarrow Alcohol.
• Enzyme: Enoyl CoA hydratase.
• R-CH=CH-C(=O)-SCoA + H2O \rightarrow R-CH(OH)-CH2-C(=O)-SCoA

Stereospecificity of Hydration

• Stereospecific reaction that yields the L stereoisomer.

Step #3: Dehydrogenation (Ketone Formation)

• Enzyme: β-hydroxyacyl CoA dehydrogenase.
• Only reacts with the L stereoisomer.
• R-CH(OH)-CH2-C(=O)-SCoA + NAD^+ \rightarrow R-C(=O)-CH2-C(=O)-SCoA + NADH +H^+

Step #4: Thiolysis

• Reaction that cleaves off 2 carbons.
• Enzyme: Thiolase.
• Result: Acetyl CoA + Acyl CoA (shortened by 2 carbons).
• R-C(=O)-CH2-C(=O)-SCoA + CoA-SH \rightarrow R-C(=O)-SCoA + CH3-C(=O)-SCoA

Beta-Oxidation Cycle

• With each cycle, the fatty acid chain is shortened by 2 carbons in the form of Acetyl CoA, and the process repeats.
• For example, an 18-carbon fatty acid undergoes 8 cycles.

Summary of Beta-Oxidation Steps

Illustration with a specific fatty acid chain

Unsaturated Fatty Acids

• Require extra steps.
• Epimerase is needed to convert D-β-Hydroxyacyl CoA to L-β-Hydroxyacyl CoA for Step #3.

Cis-trans Isomerase

• Additional enzyme needed to convert cis double bonds to trans double bonds.
• Walk alkene down a bond and make it trans.

ATP Production from Beta-Oxidation

• 1 Acetyl CoA = 10 ATP (via citric acid cycle, electron transport chain, and oxidative phosphorylation)
• 1 FADH2 = 1.5 ATP
• 1 NADH = 2.5 ATP
• 1 GTP (equivalent to ATP)

Total ATP Production

• Question on total ATP production (gross vs. net).

Example Calculation

• C16 fatty acid generates 8 Acetyl CoA molecules.
  • 8 Acetyl CoA * 10 ATP/Acetyl CoA = 80 ATP
• 7 cycles of beta-oxidation:
  • 7 FADH2 * 1.5 ATP/FADH2 = 10.5 ATP
  • 7 NADH * 2.5 ATP/NADH = 17.5 ATP
• Total ATP: 80 + 10.5 + 17.5 = 108 ATP

Energy per Gram Comparison

• Stearic acid (C18):
  • \frac{1 \text{ g stearic acid}}{284 \text{ g/mol}} \times \frac{120 \text{ mol ATP}}{1 \text{ mol stearic acid}} = 0.423 \text{ mol ATP}
• Glucose:
  • \frac{1 \text{ g glucose}}{180 \text{ g/mol}} \times \frac{30 \text{ mol ATP}}{1 \text{ mol glucose}} = 0.167 \text{ mol ATP}
• Nutritionists:
  • 1 g carbohydrate = 4 kcal
  • 1 g fat = 9 kcal

Fuel Usage Generalizations

• Skeletal Muscle:
  • Glucose/glycogen when active
  • Fatty acids at rest
• Cardiac Muscle: Prefers Fatty acids and ketone bodies.
• Liver: Fatty acids.
• Brain: Glucose and ketone bodies.

Ketone Bodies & Ketogenesis

• Lipid droplets in hepatocytes.
• Acetoacetate, D-β-hydroxybutyrate and acetone are the ketone bodies.
• Acetoacetate and D-β-hydroxybutyrate exported for use by heart, muscle, kidney and brain.
• Glucose is made from pyruvate via gluconeogenesis and is exported as fuel for the brain and other tissues.

Conditions Leading to Ketogenesis

• Low-carbohydrate diet
• Diabetes
• Fasting
• Leads to high acetyl CoA concentration and low oxaloacetate concentration; acetyl CoA is then shunted to ketone body synthesis.

Ketone Bodies Structures

• Acetoacetate (C4)
• β-hydroxybutyrate (C4)
• Acetone (C3)

Interconversion of Ketone Bodies

• Acetoacetate ---> β-hydroxybutyrate (Reduction, NADH required)
• Acetoacetate ---> Acetone (Decarboxylation, releases CO2)

Ketogenesis Location

• Occurs in liver mitochondria.
• First ketone produced is acetoacetate.
• Acetone is synthesized in the blood.

Ketogenesis Process

• Acetyl CoA + Acetyl CoA \rightarrow Acetoacetyl CoA
• Acetoacetyl CoA + Acetyl CoA \rightarrow 3-Hydroxy-3-methylglutaryl CoA
• 3-Hydroxy-3-methylglutaryl CoA \rightarrow Acetoacetate + Acetyl CoA
• Acetoacetate \rightarrow β-Hydroxybutyrate