CHE414 Lecture 29 (Glycolysis Part II; F24)

Lecture Overview

  • Topic: Glucose Metabolism: Part II

  • Important announcements:

    • Last week of lab

    • Upcoming practical exam and take-home quiz

    • Fun project on Friday with prizes

    • Exam 4 next week and planning for extra credit and winter break

Step 2: Phosphoglucose Isomerase Reaction

  • Isomerization reaction: Conversion of an aldose (glucose) to ketose (fructose)

  • Involves ring opening and closing, reversible due to near-equilibrium state

  • Coupled to the next reaction with highly negative free energy change, facilitating forward progression

Acid-Base Catalysis

  • Involvement of amino acids like Lys, His (proton donation) and Glu

  • Understand the reaction conceptually rather than step-for-step

  • Proton transfer creates a double bond affecting the isomerization

Importance of Isomerization

  • Makes C1 of fructose product available for phosphorylation

  • Direct entry of fructose into glycolysis

Step 3: Phosphofructokinase Reaction

  • Enzyme: Phosphofructokinase (PFK)

  • Phosphorylates fructose-6-phosphate (F6P) to fructose-1,6-bisphosphate (FBP)

  • Requires ATP; reaction is irreversible and represents the first committed step of glycolysis with large negative free energy change (-14.2 kJ/mol)

  • Mg2+ is an essential cofactor

Committed Step of Glycolysis

  • The resulting FBP cannot proceed to other pathways, confirming it as the committed step

  • A second phosphate group is necessary for the following cleavage into two 3C intermediates

Regulation of PFK

  • PFK has four active and allosteric sites (tetrameric enzyme)

  • Inhibition: High ATP concentrations inhibit PFK, increasing KM and lowering substrate affinity

  • Activation: AMP and ADP binding promotes F6P binding; indicates energy demand

  • Inhibition by citrate signals slow down of glycolysis to maintain balance with citric acid cycle

Step 4: Aldolase Reaction

  • Conversion of fructose-1,6-bisphosphate (FBP) to two three-carbon molecules, each containing a phosphate group

Step 5: Triose Phosphate Isomerase Reaction

  • Converts dihydroxyacetone phosphate (DHAP) to glyceraldehyde-3-phosphate (GAP)

  • Only GAP can proceed in glycolysis; conversion keeps reaction moving despite a positive ΔG

Mechanism of TPI

  • Induced fit mechanism where substrate binds triggers closing of the active site

Summary of Glycolysis Progression

  • Each glucose molecule generates two three-carbon intermediates that continue through glycolysis

  • Key goal: Generate ATP and high-energy intermediates for ATP synthesis

GAP Dehydrogenase Reaction (Step 6)

  • Phosphate comes from inorganic phosphate, forming an acyl-phosphate with high transfer potential

  • NAD+ is reduced to NADH; identifies need for subsequent NAD+ regeneration

  • Reaction consists of dual phosphorylation and oxidation-reduction

NAD+ Role in Glycolysis

  • Definitions: Oxidation (loss of electrons), Reduction (gain of electrons)

  • NAD+ acts as a cofactor to accept electrons during glycolytic oxidation

Step 7: Phosphoglycerate Kinase Reaction

  • 1,3-bisphosphoglycerate donates phosphate to ADP, generating ATP via substrate-level phosphorylation

  • Reactions recur twice, leading to recouping 2 ATPs

Step 8: Phosphoglycerate Mutase Reaction

  • Transfers phosphate to C2 via a phosphorylated active site His residue

  • Isomerization is reversible, influencing steady-state phosphate levels

Step 9: Enolase Reaction

  • Catalyzes a dehydration reaction producing water

  • Requires magnesium to coordinate with the hydroxyl group, enhancing the reaction

Phosphoenolpyruvate (PEP) Characteristics

  • PEP is high in energy due to restricted tautomerization; drives the formation of pyruvate in glycolysis

Step 10: Pyruvate Kinase Reaction

  • Final step in glycolysis yielding ATP from substrate-level phosphorylation

  • Regulated by phosphorylation with catalytic efficiency and significant energy drop

Free Energy Changes in Glycolysis

  • Steps 1, 3, and 10 are irreversible due to high energy shifts

  • Glycolysis operates through multiple pathways and conditions, ensuring flexibility in metabolic responses

Overall Yield from Glycolysis

  • One glucose produces two pyruvate, consumes two ATP, and nets four ATPs (net gain of 2 ATPs)

  • Additionally results in two molecules of NADH

Fate of Pyruvate

  • Can enter the citric acid cycle for further energy production based on cellular conditions and oxygen availability

  • During anaerobic respiration, conversion to lactate allows for temporary ATP production

  • Some microorganisms, like yeast, generate ethanol anaerobically for NAD+ regeneration

Gluconeogenesis Overview

  • Conversion of pyruvate back to glucose in the liver when glycogen is depleted

  • Utilizes and modifies many glycolytic enzymes, introducing unique enzymes like pyruvate carboxylase and phosphoenolpyruvate carboxykinase

Specific Enzymes in Gluconeogenesis

  • Four unique enzymes:

    • Pyruvate carboxylase

    • Phosphoenolpyruvate carboxykinase

    • Fructose bisphosphatase

    • Glucose-6-phosphatase