Metabolism Lecture Notes

Lecture Overview

This lecture covers several key metabolic pathways:

  • Glycolysis, Citric Acid Cycle (Krebs Cycle), and Electron Transport Chain
  • Fat Catabolism and Biosynthesis of Fats and Sugars
  • Amino Acid Metabolism and the Urea Cycle
  • Localization and Integration of Processes
  • Metabolism Problem Solving

Lecture 2 Focus

  • Electron Transport Chain
  • Gluconeogenesis
  • Introduction to Fats
  • Digestion, Mobilization, and Transport of Fats (Brief Overview)
  • Fatty Acid Activation
  • Fatty Acid Oxidation
  • Fatty Acid Synthesis (Note: Not covered in this lecture)

Glycolysis, Citric Acid Cycle, and Electron Transport Chain

Glycolysis (Cytoplasm)

  • Glucose (C6) is converted, consuming 2 ATP in the energy investment stage.
  • Glucose 6-phosphate (C6-P) and fructose 6-phosphate (C6-P) are intermediates.
  • 2 molecules of glyceraldehyde 3-phosphate (P-C3) are produced.
  • In the energy harvesting stage, 4 ATP are generated, resulting in a net gain of 2 ATP.
  • 2 NADH molecules are produced.
  • 2 pyruvate molecules (C3) are the end product.

Citric Acid Cycle (Mitochondria)

  • Pyruvate is converted to Acetyl-CoA, producing CO2 and NADH.
  • Citrate (C6) is formed.
  • CO2 is released, and NADH is produced in the conversion to ketoglutarate (C5).
  • Succinyl-CoA, succinate (C4), fumarate (C4) and oxaloacetate (C4) are intermediates in the cycle. FADH2 and ATP are generated The Krebs Cycle produces 6 NADH, 2 FADH2, and 2 ATP/GTP.

Electron Transport Chain

  • NADH and FADH2 from glycolysis and the citric acid cycle are used to reduce oxygen (O2O_2) and generate ATP.
  • Electrons are transferred through a series of protein complexes (Complex I, Complex II, Complex III, and Complex IV) along the inner mitochondrial membrane.
  • Protons (H+H^+) are pumped from the mitochondrial matrix to the intermembrane space, creating a proton gradient.
  • ATP synthase uses the proton gradient to synthesize ATP from ADP and inorganic phosphate (PiP_i).
  • Oxygen is the final electron acceptor, forming water (H2OH_2O).

Electron Transport Chain – Overview

  • The electron transport chain is a series of 4 proteins that receive the high energy electrons from NADH and FADH2 molecules and move those electrons along the inner mitochondrial membrane and on to the final electron acceptor, oxygen. In the process a proton gradient is established which is used to create ATP molecules with the help of a special enzyme called ATP synthase.

  • High energy electrons are transferred onto the electron transport chain

  • The movement of electrons stimulates the movement of H+H^+ ions out of the matrix and into the intermembrane space of the mitochondria, establishing an electrical potential difference.

  • The electrons are ultimately captured by oxygen to form water.

  • The unequal distribution of H+H^+ ions causes the spontaneous movement of the H+H^+ down their electrochemical gradient and back into the matrix. This flow of ions helps generate ATP via the enzyme ATP synthase.

Gluconeogenesis

  • Gluconeogenesis is the synthesis of glucose from non-carbohydrate precursors.
  • Key precursors include pyruvate, lactate, amino acids, and oxaloacetate.
  • Several enzymes are involved, including:
    • Pyruvate carboxylase: Converts pyruvate to oxaloacetate.
    • Phosphoenolpyruvate carboxykinase: Converts oxaloacetate to phosphoenolpyruvate.
    • Fructose 1,6-bisphosphatase: Converts fructose 1,6-bisphosphate to fructose 6-phosphate.
    • Glucose 6-phosphatase: Converts glucose 6-phosphate to glucose.
  • The process bypasses the irreversible steps of glycolysis, utilizing different enzymes to overcome the large negative free-energy changes.
  • Magnesium ions (Mg++Mg^{++}) act as cofactors for many of these enzymes.
  • ATP and GTP are consumed in the process.

Introduction to Fats: Energy Storage

  • Glycogen (Liver/Muscle):
    • Approximately 480g stored.
    • Energy extracted: 4 Kcal/g.
    • Total energy: ~2,000 Kcal; lasts about 1 day
  • Protein (Muscles):
    • Approximately 6000g.
    • Energy extracted: 4 Kcal/g.
    • Total energy: ~24,000 Kcal; lasts about 12 days
  • Fat (Adipose):
    • Approximately 12,000g.
    • Energy extracted: 9 Kcal/g.
    • Total energy: ~120,000 Kcal; lasts 60 days

Digestion, Mobilization, and Transport of Fats

  1. Emulsification: Bile salts, released by the gallbladder, emulsify dietary fats in the small intestine, forming mixed micelles.
  2. Degradation: Intestinal lipases degrade triacylglycerols into fatty acids and glycerol.
  3. Uptake and Conversion: Fatty acids and other breakdown products are taken up by the intestinal mucosa and converted back into triacylglycerols.
  4. Chylomicron Formation: Triacylglycerols are incorporated, along with cholesterol and apoproteins (like ApoC-II), into chylomicrons.
  5. Transport: Chylomicrons move through the lymphatic system and bloodstream to tissues.
  6. Fatty Acid Release: Lipoprotein lipase, activated by ApoC-II in the capillaries, releases fatty acids and glycerol from chylomicrons.
  7. Cellular Uptake: Fatty acids enter cells and are either oxidized for fuel or re-esterified for storage.

Fatty Acid Activation

  • Occurs in the cytoplasm.
  • Fatty acids are activated by converting them to Fatty Acyl-CoA using Acyl-CoA Synthetase.
  • This process occurs in two steps and requires ATP.
  • In first step, Acyl-CoA synthetase catalyzes the transfer of adenosine monophosphate from ATP on to the fatty acid. This forms Acyl adenylate and releases a pyrophosphate
  • Pyrophosphatase hydrolyses the pyrophosphate into 2 orthophosphate molecules which can be used in many other reactions
  • A coenzyme A molecule attacks acyl adenylate forming acyl- coenzyme A and releasing AMP in the process
  • The reaction yields AMP and pyrophosphate (PPiPP_i).
Carnitine Shuttle
  • Transports fatty acyl-CoA from the cytoplasm into the mitochondrial matrix for beta-oxidation.
  • Carnitine acyltransferase 1 (CAT1) located on the outer mitochondrial membrane, transfers the acyl group from CoA to carnitine, forming fatty acyl-carnitine.
  • The outer mitochondrial membrane contains porins, allowing fatty acyl-carnitine to diffuse into the intermembrane space.
  • Acyl-carnitine translocase, an inner membrane transporter, moves fatty acyl-carnitine into the mitochondrial matrix.
  • Carnitine acyltransferase 2 (CAT2), located on the inner mitochondrial membrane, converts fatty acyl-carnitine back to fatty acyl-CoA and releases carnitine.
  • Carnitine is then shuttled back into the cytoplasm via acyl carnitine translocase and the porins of the outer mitochondrial membrane.

Fatty Acid Oxidation (Beta-Oxidation)

  • Occurs in the mitochondrial matrix.
  • A repetitive four-step process that shortens the fatty acyl-CoA by two carbon atoms with each cycle.
  • Step 1: Oxidation by FAD
    • Acyl-CoA dehydrogenase catalyzes the formation of a double bond between the alpha and beta carbons of the fatty acyl-CoA, producing trans-Δ2-enoyl-CoA and FADH2.
  • Step 2: Hydration
    • Enoyl-CoA hydratase adds water across the double bond of trans-Δ2-enoyl-CoA, forming L-β-hydroxyacyl-CoA.
  • Step 3: Oxidation by NAD+
    • L-β-hydroxyacyl-CoA dehydrogenase oxidizes L-β-hydroxyacyl-CoA to β-ketoacyl-CoA, producing NADH + H+.
  • Step 4: Thiolytic Cleavage by CoA
    • Thiolase cleaves β-ketoacyl-CoA, releasing acetyl-CoA and a fatty acyl-CoA molecule shortened by two carbons.
  • Acetyl-CoA enters the citric acid cycle for further oxidation.
  • FADH2 and NADH enter the electron transport chain, generating ATP.

Fatty Acid Synthesis

  • Occurs in the cytosol.
  • Acetyl-CoA is transported from the mitochondrial matrix to the cytosol via the citrate shuttle.
  • Acetyl-CoA is converted to malonyl-CoA by acetyl-CoA carboxylase, a biotin-dependent enzyme. CO2 is also incorporated
  • Fatty acid synthase, a multienzyme complex, catalyzes the synthesis of palmitate (a 16-carbon saturated fatty acid) from acetyl-CoA and malonyl-CoA.
  • The process involves a series of condensation, reduction, dehydration, and reduction reactions, adding two carbons to the growing fatty acid chain with each cycle.
  • NADPH is the reducing agent.
  • Palmitate can be further elongated and desaturated in the endoplasmic reticulum.

Steps in Fatty Acid Synthesis:

  1. Citrate Shuttle: Acetyl-CoA is transported from the mitochondrial matrix to the cytosol as citrate.
  2. Malonyl-CoA Formation: Acetyl-CoA carboxylase converts acetyl-CoA to malonyl-CoA.
  3. Palmitate Synthesis: Fatty acid synthase synthesizes palmitate from acetyl-CoA and malonyl-CoA through repeated condensation, reduction, dehydration, and reduction reactions.
  4. Modification: Palmitate can undergo elongation and desaturation to form other fatty acids.

Fatty Acid Synthase Reactions:

  • Acetyl-CoA (from the citrate shuttle) and Malonyl-CoA (formed from Acetyl CoA) bind to the enzyme. This releases CoA.
  • Condensation: B-Ketoacyl-ACP is formed and carbon dioxide is released
  • Reduction: NADPH is converted to NADP
  • Dehydration: Water is released
  • Reduction: NADPH is converted to NADP
  • Acyl(C+2)-ACP) is formed. Note that each increment produces to additional carbons.