Glycolysis and Gluconeogenesis

Learning Objectives

• Understand how energy is harnessed from glucose via glycolysis.

• Describe metabolism under anaerobic conditions.

• Explain the synthesis of glucose from simpler compounds (gluconeogenesis).

Central Importance of Glucose

Glucose as a Metabolic Fuel and Precursor

• Glucose is a primary energy source for most organisms, yielding significant energy upon oxidation.

• It can be efficiently stored as polymers such as starch (plants) or glycogen (animals).

• Many tissues and organisms rely exclusively on glucose for energy needs.

• Glucose serves as a versatile biochemical precursor for the synthesis of:

  • All amino acids

  • Membrane lipids

  • Nucleotides (DNA and RNA)

  • Cofactors required for metabolism

• Life evolved around glucose due to its central metabolic role.

Four Major Destinies of Glucose Utilization

1. Storage as starch or glycogen when energy is abundant.

2. Oxidation via glycolysis and the TCA cycle for short-term energy needs.

3. Entry into the pentose phosphate pathway to generate NADPH for reductive biosynthesis.

4. Formation of structural polysaccharides (e.g., cell walls, glycoproteins, proteoglycans).

Glycolysis

Overview and Significance

• Glycolysis is a nearly universal pathway for glucose metabolism, occurring in the cytosol.

• It consists of ten enzyme-catalyzed reactions that convert glucose to two molecules of pyruvate.

• During glycolysis, ATP and NADH are produced.

• The pathway is highly conserved, but regulation and pyruvate fate can differ among species.

• All glycolytic enzymes are homodimers or homotetramers.

Historical Importance

• Research on glycolysis led to the discovery of coenzymes, the role of ATP, and methods for enzyme purification.

Glycolysis: Phases and Steps

Preparatory Phase (Steps 1–5)

• Converts D-glucose into two molecules of glyceraldehyde 3-phosphate (G3P).

• Requires the investment of two ATP molecules.

• Includes an essential isomerization from glucose to fructose.

• No energy is extracted yet; intermediates are phosphorylated.

Payoff Phase (Steps 6–10)

• Each G3P is converted to pyruvate, yielding 4 ATP and 2 NADH (net gain: 2 ATP, 2 NADH per glucose).

• Much energy remains in pyruvate.

Detailed Steps of Glycolysis

Step 1: Hexokinase Reaction

• Phosphorylation of glucose to glucose 6-phosphate by hexokinase.

• ATP is the phosphate donor; Mg2+ is required as a cofactor.

• ΔG'° = –16.7 kJ/mol; ΔG = –33.4 kJ/mol (highly favorable).

• Equation:

Step 2: Phosphohexose Isomerase

• Isomerization of glucose 6-phosphate (aldose) to fructose 6-phosphate (ketose).

• Catalyzed by phosphohexose isomerase with Mg2+ as a cofactor.

• ΔG'° = 1.7 kJ/mol; ΔG = 0 to 2.5 kJ/mol (near equilibrium).

• Mechanism involves an enediol intermediate and active-site glutamate.

• Equation:

Step 3: Phosphofructokinase-1 (PFK-1)

• Phosphorylation of fructose 6-phosphate to fructose 1,6-bisphosphate.

• ATP is the phosphate donor; Mg2+ is required.

• This is the first committed step of glycolysis and is irreversible.

• PFK-1 is negatively regulated by ATP (feedback inhibition).

• ΔG'° = –22.2 kJ/mol.

• Equation:

Step 4: Aldolase

• Cleavage of fructose 1,6-bisphosphate into dihydroxyacetone phosphate (DHAP) and glyceraldehyde 3-phosphate (G3P).

• Energetically unfavorable; coupled to subsequent reactions to drive forward.

• ΔG'° = –6 to 0 kJ/mol.

• Equation:

Step 5: Triose Phosphate Isomerase

• Interconversion of DHAP (ketotriose) and G3P (aldotriose).

• Only G3P continues in glycolysis; DHAP is converted as needed.

• ΔG'° = 7.5 kJ/mol; ΔG = 0 to 4 kJ/mol.

• Equation:

Step 6: Glyceraldehyde 3-Phosphate Dehydrogenase

• First energy-yielding step: oxidation of G3P to 1,3-bisphosphoglycerate (1,3-BPG).

• NAD+ is reduced to NADH.

• ΔG'° = 6.3 kJ/mol; ΔG = –2 to 2 kJ/mol.

• Equation:

Step 7: Phosphoglycerate Kinase (First Substrate-Level Phosphorylation)

• Transfer of a phosphate from 1,3-BPG to ADP, forming ATP and 3-phosphoglycerate.

• This is an example of substrate-level phosphorylation (SLP).

• Equation:

Step 8: Phosphoglycerate Mutase

• Isomerization of 3-phosphoglycerate to 2-phosphoglycerate.

• Enzyme contains a phosphohistidine intermediate.

• Equation:

Step 9: Enolase

• Dehydration of 2-phosphoglycerate to phosphoenolpyruvate (PEP).

• PEP is a high-energy compound.

• Equation:

Step 10: Pyruvate Kinase (Second Substrate-Level Phosphorylation)

• Transfer of phosphate from PEP to ADP, forming ATP and pyruvate.

• Reaction is driven by the tautomerization of enolpyruvate to keto form.

• Equation:

Summary of Glycolysis

• Overall equation:

• ΔG'° ≈ –85 kJ/mol; energy recovery efficiency is just over 60% under cellular conditions.

• Enzymes often form multienzyme complexes (metabolons) for efficient substrate channeling.

Phosphorylated Intermediates in Glycolysis

• All glycolytic intermediates are phosphorylated for three main reasons:

  1. They are ionized at physiological pH, preventing diffusion out of the cell.

  2. Phosphoryl groups are essential for enzymatic conservation of metabolic energy (ATP formation).

  3. Binding energy from phosphate groups lowers activation energy for enzyme-catalyzed reactions.

Fates of Pyruvate

• Pyruvate, the end product of glycolysis, has several metabolic fates:

  1. Aerobic conditions: Oxidized to acetyl-CoA (with loss of CO2), enters the citric acid cycle, and is fully oxidized to CO2.

  2. Anaerobic conditions: Reduced to lactate (in animals) or ethanol + CO2 (in yeast), regenerating NAD+ for glycolysis.

  3. Can be used for anabolic processes, such as the biosynthesis of alanine.

ATP Formation in Glycolysis

• Some energy from glucose catabolism is conserved in ATP; much remains in pyruvate.

• Net ATP yield per glucose: 2 ATP (4 produced, 2 consumed).

• Net NADH yield per glucose: 2 NADH.

Table: Key Enzymes and Steps of Glycolysis

Additional info:

• Glycolysis is tightly regulated at the hexokinase, phosphofructokinase-1, and pyruvate kinase steps.

• Gluconeogenesis is the reverse pathway, with unique enzymes bypassing the irreversible steps of glycolysis.