Glycolysis Notes

Carbohydrate Metabolism - Glycolysis

Learning Objectives

  • Glycolysis is an energy conversion pathway involving 10 reactions.
  • Regeneration of NAD+ occurs from the metabolism of pyruvate.
  • Fructose and galactose are converted into glycolytic intermediates.
  • Glycolysis is regulated in both muscle and liver tissues.
  • Understanding lactose intolerance.
  • Understanding Galactosaemia.
  • Understanding Hypoxia and the Warburg effect.

Glycolysis: Turning Glucose into Energy

  • Glucose is the primary carbohydrate in living systems.
  • It is the sole fuel source for the brain under non-starvation conditions.
  • Red blood cells can only use glucose as fuel.
  • Glucose tends to exist in a ring formation.
  • This reduces the likelihood of glucose non-specifically modifying proteins and affecting their activity.

How Glucose Enters the Cell

  • Insulin facilitates glucose entry into the cell.
  • Insulin binds to insulin receptors.
  • This triggers a signal transduction cascade.
  • GLUT4 transporters become active.
  • Glucose is transported from outside to inside the cell.
  • Inside the cell, glucose is utilized for glycogen, lipid, and protein synthesis.
  • Gene expression and growth regulation are also influenced.

What is Glycolysis?

  • Glycolysis is a process common to almost all prokaryotic and eukaryotic cells.
  • The 10 reactions of glycolysis occur in the cytoplasm.
  • Glucose is converted into 2 molecules of pyruvate and 2 molecules of ATP.
  • NAD+ is reduced to NADH, storing energy as reducing power.
  • It is the initial step in both aerobic and anaerobic respiration.
  • Glycolysis is divided into two consecutive stages.

Stages of Glycolysis

  • Glycolysis can be divided into two main stages:
    • Stage 1: Energy investment phase (Steps 1-5).
      • Glucose is converted into 2 molecules of glyceraldehyde 3-phosphate (G3P).
      • Two ATP molecules are invested in this stage.
    • Stage 2: Energy payoff phase (Steps 6-10).
      • Two molecules of G3P are converted into 2 molecules of pyruvate + 4 ATP molecules.
      • Results in a net production of two ATP molecules and two NADH molecules.

Irreversible Steps in Glycolysis

  • Irreversible steps in glycolysis along with their Gibbs free energy changes ΔG\Delta G (kcal/mol):
    • Hexokinase: −33.5-33.5
    • Phosphoglucose isomerase: −2.5-2.5
    • Phosphofructokinase: −22.2-22.2
    • Aldolase: −1.3-1.3
    • Triose phosphate isomerase: −1.7-1.7
    • Glyceraldehyde 3-phosphate dehydrogenase: +1.3+1.3
    • Phosphoglycerate kinase: +0.8+0.8
    • Phosphoglycerate mutase: −3.3-3.3
    • Pyruvate kinase: −16.7-16.7

Regulation of Glycolysis

  • The irreversible steps, which have a significant negative ΔG\Delta G, are the points where the pathway is regulated.

Trapping and Generation Phase

  • The initial strategy is to trap glucose inside the cell and form a compound that can be readily cleaved into phosphorylated three-carbon units.
    • Step 1: Hexokinase phosphorylates glucose using ATP to form glucose 6-phosphate.
      • The phosphoryl group prevents glucose 6-phosphate from passing through the cell membrane, effectively locking it inside the cell.
      • Hexokinase is a kinase, a class of enzymes catalyzing the transfer of a phosphoryl group from ATP to an acceptor.
    • Step 2: Phosphoglucose isomerase catalyzes the isomerization of glucose 6-phosphate to fructose 6-phosphate.
      • This is a conversion of an aldose into a ketose.
      • Only fructose 6-phosphate can be readily cleaved into two three-carbon fragments.
    • Step 3: Phosphofructokinase (PFK) phosphorylates fructose 6-phosphate (F-6P) using ATP to form fructose 1,6-bisphosphate (F-1,6-BP).
      • This reaction is irreversible under cellular conditions and is the committed step of glycolysis.
      • PFK is the key regulatory enzyme of glycolysis.
      • Bisphosphate indicates two separate monophosphoryl groups, while diphosphate means two phosphoryl groups are connected by an anhydride linkage (e.g., ADP).
    • Step 4: F-1,6-BP is cleaved by aldolase into dihydroxyacetone phosphate (DHAP) and glyceraldehyde 3-phosphate (GAP).
      • Unlike GAP, DHAP is not directly in the glycolytic pathway.
    • Step 5: DHAP is interconverted into GAP by triose phosphate isomerase (TPI).
      • Isomerization ensures that the unused three-carbon fragment is utilized for glycolysis.
      • TPI deficiency is a rare and lethal glycolytic enzymopathy, characterized by severe hemolytic anemia and neurodegeneration.

Three-Carbon Units Oxidation and Energy Generation

  • Energy is produced in the form of ATP.
    • Step 6: Glyceraldehyde 3-phosphate is oxidized and phosphorylated to form 1,3-bisphosphoglycerate.
      • The oxidation-reduction reaction is catalyzed by glyceraldehyde 3-phosphate dehydrogenase.
      • Dehydrogenases catalyze the transfer of a hydride ion from a donor molecule to NAD+.
      • 1,3-Bisphosphoglycerate has a high phosphoryl-transfer potential, which is used to generate ATP in the next step.
    • Step 7: Phosphoglycerate kinase catalyzes the transfer of the phosphoryl group from the acyl phosphate of 1,3-BPG to ADP, producing ATP and 3-phosphoglycerate.
      • 1,3-Bisphosphoglycerate is an energy-rich molecule with a greater phosphoryl-transfer potential than ATP.
      • Therefore, it can be used to synthesize ATP from ADP and orthophosphate.
    • Step 8: 3-Phosphoglycerate is converted into 2-phosphoglycerate by phosphoglycerate mutase, which shifts the position of the phosphoryl group.
      • Mutases are enzymes that catalyze the intramolecular shift of chemical groups.
    • Step 9: Dehydration of 2-phosphoglycerate, catalyzed by enolase, introduces a double bond, creating phosphoenolpyruvate.
      • Enol phosphates are an unstable class of molecules with an elevated phosphoryl group transfer potential.
    • Step 10: The phosphoryl transfer from phosphoenolpyruvate to ADP is catalyzed by pyruvate kinase, resulting in the production of ATP and pyruvate.
      • The phosphoryl group of phosphoenolpyruvate traps the 3-carbon compound in its unstable enol form.
      • When the phosphoryl group is donated to ATP, the enol undergoes conversion into the stable ketone, pyruvate.

Overall (Net) Reaction for Glycolysis

  • Glucose + 2 Pi + 2 ADP + 2 NAD+ → 2 pyruvate + 2 ATP + 2 NADH + 2 H+ + 2 H2O

Regulation of Glycolysis Pathway

  • Flux through the glycolysis pathway is regulated by controlling three enzymes that catalyze spontaneous reactions: hexokinase, phosphofructokinase, and pyruvate kinase.

Hexokinase Regulation

  • Hexokinase is inhibited by its own product, glucose 6-phosphate (negative feedback).
  • If PFK is inhibited by ATP, glucose 6-phosphate will accumulate.
  • This accumulation switches off hexokinase.
  • Glucose 6-phosphate is also the first step on the pathway to make glycogen.
  • When energy charge is high, glycolysis is inhibited, and glycogen is synthesized instead.

Phosphofructokinase (PFK) Regulation

  • PFK is the main site of regulation.
  • PFK is an allosteric protein.
  • Inhibitors:
    • ATP allosterically inhibits PFK by binding to a different site from the active site.
    • Citrate.
  • Activators:
    • ADP, AMP.
    • Fructose 2,6-bisphosphate.

Pyruvate Kinase Regulation

  • Pyruvate kinase is an allosteric tetramer.
  • Inhibited by ATP, Acetyl CoA, and fatty acids.
  • Decreases glycolysis rate when the energy charge is high.
  • Activated by fructose 1,6-bisphosphate.
  • Increases the rate of pyruvate kinase when PFK activity is high; hence, all the enzymes work fast at the same time.