Lipid Metabolism — Study Notes

1. Beta-Oxidation of Fatty Acids

Beta-oxidation is the catabolic pathway by which fatty acids are broken down in the mitochondrial matrix to generate Acetyl-CoA, which enters the Citric Acid Cycle, and $NADH$/$FADH_2$, which feed into the Electron Transport Chain.

Activation and Carnitine Shuttle

Before entering the mitochondria, fatty acids must be activated in the cytosol: $$\text{Fatty Acid} + \text{CoASH} + \text{ATP} \xrightarrow{\text{Acyl-CoA Synthetase}} \text{Acyl-CoA} + \text{AMP} + \text{PP}_i$$

Because the inner mitochondrial membrane is impermeable to Acyl-CoA, the Carnitine Shuttle is required: 1. CPT-I (Carnitine Palmitoyltransferase-I) in the outer mitochondrial membrane converts Acyl-CoA and carnitine to Acyl-carnitine. Key rate-limiting step, inhibited by Malonyl-CoA. 2. A translocase moves Acyl-carnitine across the inner membrane. 3. CPT-II in the inner membrane converts Acyl-carnitine back to Acyl-CoA and free carnitine.

The Four Reoccurring Steps of Beta-Oxidation

For saturated fatty acids, each cycle cleaves a two-carbon unit (as Acetyl-CoA) through four steps:

1. Oxidation: Acyl-CoA $\rightarrow$ trans-$\Delta^2$-Enoyl-CoA by Acyl-CoA Dehydrogenase. Generates $FADH_2$.
2. Hydration: trans-$\Delta^2$-Enoyl-CoA $\rightarrow$ L-$\beta$-Hydroxyacyl-CoA by Enoyl-CoA hydratase.
3. Oxidation: L-$\beta$-Hydroxyacyl-CoA $\rightarrow$ $\beta$-Ketoacyl-CoA by $\beta$-Hydroxyacyl-CoA dehydrogenase. Generates $NADH$.
4. Thiolysis: $\beta$-Ketoacyl-CoA + CoASH $\rightarrow$ Acyl-CoA (shortened by $2\text{C}$) + Acetyl-CoA by Thiolase.

Energetics of Palmitate ($16\text{C}$): - Requires 7 cycles. - Yields: $8\text{ Acetyl-CoA} + 7\text{ NADH} + 7\text{ FADH}_2$. - Total net ATP generated ≈ $106\text{ ATP}$ (after subtracting the 2 ATP equivalents used in activation).

graph TD
    A[Mitochondrial Acyl-CoA] -->|Acyl-CoA Dehydrogenase / FADH2| B[Enoyl-CoA]
    B -->|Hydratase| C[Hydroxyacyl-CoA]
    C -->|Dehydrogenase / NADH| D[Ketoacyl-CoA]
    D -->|Thiolase / CoASH| E[Acetyl-CoA + Shortened Acyl-CoA]

2. Fatty Acid Synthesis (Lipogenesis)

Fatty acid synthesis occurs in the cytosol, primarily in the liver and lactating mammary glands. It uses Acetyl-CoA and $NADPH$ to construct palmitate ($16\text{C}$).

Mitochondrial Acetyl-CoA Export

Acetyl-CoA cannot cross the mitochondrial membrane. It is converted to Citrate by Citrate Synthase, exported to the cytosol, and cleaved back to Acetyl-CoA and Oxaloacetate by ATP-Citrate Lyase.

The Committed Step

Acetyl-CoA is carboxylated to Malonyl-CoA by Acetyl-CoA Carboxylase (ACC). This is the rate-limiting and highly regulated step of lipogenesis: $$\text{Acetyl-CoA} + \text{HCO}_3^- + \text{ATP} \xrightarrow{\text{ACC}} \text{Malonyl-CoA} + \text{ADP} + \text{P}_i$$

ACC is activated by insulin and citrate, and inhibited by glucagon, epinephrine, and palmitoyl-CoA (feedback).

Fatty Acid Synthase (FAS) Complex

FAS is a multifunctional homodimeric enzyme containing an Acyl Carrier Protein (ACP) domain. Synthesis involves repeated cycles of condensation, reduction (using $NADPH$), dehydration, and reduction, extending the chain by two carbons (derived from Malonyl-CoA) per cycle until palmitate ($16\text{C}$) is released by thioesterase.

3. Ketogenesis

During prolonged fasting, starvation, or uncontrolled diabetes, oxaloacetate is consumed by gluconeogenesis in the liver. This depletes the oxaloacetate needed for the Citric Acid Cycle to oxidize Acetyl-CoA (which is being produced in large amounts by rapid beta-oxidation of adipose-derived fatty acids).

The excess Acetyl-CoA is converted in liver mitochondria into ketone bodies: Acetoacetate, $\beta$-Hydroxybutyrate, and Acetone (non-enzymatic decarboxylation, excreted in breath).

Synthesis Pathway

1. $2\text{ Acetyl-CoA} \rightarrow$ Acetoacetyl-CoA (Thiolase).
2. Acetoacetyl-CoA + Acetyl-CoA $\rightarrow$ HMG-CoA by HMG-CoA Synthase (mitochondrial). Rate-limiting step of ketogenesis.
3. HMG-CoA $\rightarrow$ Acetoacetate + Acetyl-CoA by HMG-CoA Lyase.
4. Acetoacetate is reduced to $\beta$-Hydroxybutyrate (using $NADH$).

Ketone bodies are exported to extrahepatic tissues (brain, skeletal and cardiac muscle), where they are converted back to Acetyl-CoA to enter the TCA cycle for energy. The liver cannot use ketone bodies because it lacks the enzyme thiophorase ($\beta$-ketoacyl-CoA transferase).

4. Cholesterol Metabolism

Cholesterol is essential for membrane structure, steroid hormones, and bile acid synthesis.

Biosynthesis (Cytosolic)

All 27 carbon atoms of cholesterol are derived from Acetyl-CoA. 1. Synthesis of HMG-CoA from Acetyl-CoA (via cytosolic HMG-CoA Synthase). 2. HMG-CoA is reduced to Mevalonate by HMG-CoA Reductase (integral membrane protein of ER, uses $2\text{ NADPH}$). Key rate-limiting and committed step of cholesterol synthesis. 3. Mevalonate is converted to active isoprenes, which condense to form squalene, which is cyclized to lanosterol and ultimately cholesterol.

Regulation of HMG-CoA Reductase

• Feedback inhibition: High cholesterol levels trigger proteasomal degradation of the enzyme and inhibit transcription via the SREBP/SCAP system.
• Covalent modification: Phosphorylation by AMPK inactivates the enzyme (under low energy state).
• Hormonal control: Insulin activates the enzyme, while glucagon inactivates it.
• Pharmacological: Statins are competitive inhibitors of HMG-CoA Reductase used to lower blood cholesterol.

5. Lipoproteins

Lipids are insoluble in water and must be transported in blood as macromolecular complexes called lipoproteins. They consist of a hydrophobic core (triglycerides, cholesterol esters) surrounded by a hydrophilic monolayer of phospholipids, free cholesterol, and apolipoproteins.

• Apo C-II activates Lipoprotein Lipase (LPL) on capillary walls, hydrolyzing triglycerides in chylomicrons and VLDLs to release free fatty acids for tissues.
• Apo E mediates the clearance of chylomicron remnants and VLDL/IDL by liver receptors.
• Apo B-100 acts as the ligand for the LDL Receptor, triggering receptor-mediated endocytosis of LDL.

6. Triglyceride Mobilization

During fasting, epinephrine and glucagon bind to G-protein coupled receptors on adipocytes: 1. PKA phosphorylates and activates Hormone-Sensitive Lipase (HSL) and perilipin (which coats the lipid droplet). 2. Activated Adipose Triglyceride Lipase (ATGL) initiates lipolysis by converting triglycerides to diglycerides. 3. HSL converts diglycerides to monoglycerides. 4. Monoacylglycerol lipase (MGL) completes the process, yielding 3 free fatty acids and 1 glycerol.

Free fatty acids bind to serum albumin in blood and are transported to tissues for beta-oxidation. Glycerol is taken up by the liver for gluconeogenesis or glycolysis.