Glycolysis and Gluconeogenesis

• Glycolysis and gluconeogenesis are central metabolic pathways.
• Glycolysis breaks down glucose to pyruvate, while gluconeogenesis synthesizes glucose from pyruvate and other precursors.
• These pathways are reciprocally regulated to maintain glucose homeostasis.

Glycolysis Steps

• Glycolysis occurs in the cytoplasm and consists of ten enzymatic steps.
• Step 1: Hexokinase/Glucokinase: Glucose is phosphorylated to glucose-6-phosphate (G6P).
  • Hexokinase is found in most tissues
  • Glucokinase is found in the liver and pancreas.
  • This traps glucose inside the cell and commits it to glycolysis
  • ATP is consumed, and the reaction is irreversible.
• Step 2: Phosphoglucose Isomerase: G6P is isomerized to fructose-6-phosphate (F6P).
  • This reaction is reversible
  • It prepares the molecule for the next phosphorylation step.
• Step 3: Phosphofructokinase-1 (PFK-1): F6P is phosphorylated to fructose-1,6-bisphosphate (F1,6BP).
  • PFK-1 is the major regulatory point of glycolysis
  • ATP is consumed, and the reaction is irreversible.
• Step 4: Aldolase: F1,6BP is cleaved into two 3-carbon molecules: glyceraldehyde-3-phosphate (GAP) and dihydroxyacetone phosphate (DHAP).
  • The reaction is reversible.
• Step 5: Triose Phosphate Isomerase: DHAP is converted to GAP.
  • This ensures that both products of the aldolase reaction can proceed through the rest of glycolysis.
  • The reaction is reversible.
• Step 6: Glyceraldehyde-3-Phosphate Dehydrogenase (GAPDH): GAP is oxidized and phosphorylated to 1,3-bisphosphoglycerate (1,3BPG).
  • NAD+ is reduced to NADH.
  • This is the first energy-yielding step in glycolysis
  • The reaction is reversible.
• Step 7: Phosphoglycerate Kinase: 1,3BPG transfers a phosphate group to ADP, forming ATP and 3-phosphoglycerate (3PG).
  • This is substrate-level phosphorylation.
  • The reaction is reversible.
• Step 8: Phosphoglycerate Mutase: 3PG is converted to 2-phosphoglycerate (2PG).
  • The reaction is reversible.
• Step 9: Enolase: 2PG is dehydrated to form phosphoenolpyruvate (PEP).
  • The reaction is reversible.
• Step 10: Pyruvate Kinase: PEP transfers its phosphate group to ADP, forming ATP and pyruvate.
  • This is another substrate-level phosphorylation.
  • The reaction is irreversible and is another major regulatory point.

Gluconeogenesis Steps

• Gluconeogenesis occurs mainly in the liver and, to a lesser extent, in the kidneys.
• It involves several enzymes and reactions that bypass the irreversible steps of glycolysis.
• Pyruvate to Phosphoenolpyruvate (PEP): Bypasses pyruvate kinase and involves two enzymes.
  • Pyruvate Carboxylase: Pyruvate is converted to oxaloacetate (OAA) in the mitochondria.
    • Requires ATP and biotin as a cofactor.
  • PEP Carboxykinase (PEPCK): OAA is converted to PEP.
    • This can occur in the mitochondria or cytoplasm, depending on the species and tissue.
    • GTP is consumed.
• Fructose-1,6-bisphosphate to Fructose-6-phosphate: This bypasses phosphofructokinase-1 (PFK-1).
  • Fructose-1,6-bisphosphatase (FBPase-1): F1,6BP is dephosphorylated to F6P.
    • This reaction is irreversible and is a major regulatory point.
• Glucose-6-phosphate to Glucose: This bypasses hexokinase/glucokinase.
  • Glucose-6-phosphatase: G6P is dephosphorylated to glucose in the endoplasmic reticulum.
    • This enzyme is present in the liver and kidney but absent in muscle and brain
    • This explains why these tissues cannot release free glucose into the bloodstream.

Regulation of Glycolysis

• Glycolysis is regulated at three key irreversible steps: hexokinase/glucokinase, PFK-1, and pyruvate kinase.
• Hexokinase/Glucokinase Regulation:
  • Hexokinase is inhibited by its product, G6P.
  • Glucokinase is not inhibited by G6P directly but is regulated by glucose levels and is induced by insulin.
• PFK-1 Regulation:
  • Activated by AMP and fructose-2,6-bisphosphate (F2,6BP).
  • Inhibited by ATP and citrate.
  • F2,6BP is a potent allosteric activator
  • Its levels are controlled by the enzyme PFK-2/FBPase-2.
• Pyruvate Kinase Regulation:
  • Activated by fructose-1,6-bisphosphate (feedforward activation).
  • Inhibited by ATP and alanine.
  • Liver pyruvate kinase is also regulated by phosphorylation; it is inactivated when phosphorylated by protein kinase A in response to glucagon.

Regulation of Gluconeogenesis

• Gluconeogenesis is regulated at the irreversible steps catalyzed by pyruvate carboxylase, PEPCK, and fructose-1,6-bisphosphatase.
• Pyruvate Carboxylase Regulation:
  • Activated by acetyl-CoA.
  • When energy is high, acetyl-CoA accumulates, signaling the need to store glucose.
• PEPCK Regulation:
  • Transcription of the PEPCK gene is increased by glucagon and cortisol.
• Fructose-1,6-bisphosphatase Regulation:
  • Inhibited by AMP and fructose-2,6-bisphosphate (F2,6BP).
  • Citrate activates FBPase-1.

Reciprocal Regulation

• Glycolysis and gluconeogenesis are reciprocally regulated to prevent futile cycling.
• Fructose-2,6-bisphosphate (F2,6BP):
  • This is a key regulator.
  • It activates PFK-1 (glycolysis) and inhibits FBPase-1 (gluconeogenesis).
  • F2,6BP levels are controlled by a bifunctional enzyme, PFK-2/FBPase-2.
  • PFK-2 phosphorylates F6P to form F2,6BP
  • FBPase-2 hydrolyzes F2,6BP back to F6P.
  • The activity of PFK-2/FBPase-2 is regulated by hormones:
    • Insulin activates PFK-2, increasing F2,6BP levels and stimulating glycolysis.
    • Glucagon activates FBPase-2, decreasing F2,6BP levels and stimulating gluconeogenesis.
• Hormonal Control:
  • Insulin promotes glucose utilization by stimulating glycolysis and inhibiting gluconeogenesis.
  • Glucagon promotes glucose production by stimulating gluconeogenesis and inhibiting glycolysis.
• Energy Charge:
  • High ATP levels inhibit glycolysis and stimulate gluconeogenesis.
  • High AMP levels stimulate glycolysis and inhibit gluconeogenesis.

Energetics

• Glycolysis:
  • Net ATP production: 2 ATP per glucose molecule (2 ATP consumed, 4 ATP produced).
  • 2 NADH produced per glucose molecule.
• Gluconeogenesis:
  • Requires energy: 6 ATP equivalents per glucose molecule (2 ATP for pyruvate carboxylase, 2 GTP for PEPCK, 2 ATP for phosphoglycerate kinase).
  • Consumes 2 NADH.

Importance

• Glycolysis is essential for energy production, especially in tissues with limited mitochondria (e.g., red blood cells) or during intense exercise.
• Gluconeogenesis is crucial for maintaining blood glucose levels during fasting, starvation, and prolonged exercise.
• It provides glucose to the brain and other tissues that rely on it as a primary energy source.