MF

Lecture 6 (II) ATP + Intro to metabolism F24

Page 1: Quiz Announcement

  • Quiz 3 closes tonight!

Page 2: Stability of RNA vs. DNA

  • RNA is rapidly hydrolyzed under alkaline conditions; DNA is more stable.

  • The 2′ hydroxyl of RNA:

    • Acts as a nucleophile in an intramolecular displacement.

    • Breaks the phosphodiester linkage.

Page 3: ATP Hydrolysis Free-Energy Change

  • Chemical basis for ATP hydrolysis:

    [ \Delta G^\prime_0 = \text{Free energy of products} - \text{Free energy of reactants} ]

  • Reasons for large free-energy change:

    1. Hydrolysis releases electrostatic repulsion among negative charges.

    2. Inorganic phosphate ( ( Pi )) has greater resonance stabilization than ATP.

    • Reaction: ( ATP \rightarrow ADP + Pi )

Page 4: Hydrolysis of ATP

  • ATP contains two high-energy bonds (phosphoanhydride linkages).

    • Reaction: ( ATP \rightarrow ADP + Pi )

    • Standard free energy change: ( \Delta G’_0 = -30.5 \text{ kJ/mol} )

    • Hydrolysis of the gamma-beta phosphoanhydride bond through nucleophilic attack:

      • Yields ADP and Pi, releasing ~30 kJ of energy.

Page 5: Additional Hydrolysis Mechanisms

  • Resulting Products:

    1. Yields AMP and pyrophosphate (PPi).

    2. Hydrolysis of alpha-beta linkage (nucleophilic attack of the alpha phosphate):

      • PPi is immediately hydrolyzed by pyrophosphatase: ( PPi \rightarrow 2 Pi )

    • Hydrolysis of ATP to AMP + PPi has a free energy change: ( \Delta G^\prime_0 = -45.6 \text{ kJ/mol} )

    • Overall reaction and net free energy change:

      • ( ATP \rightarrow AMP + 2 Pi )

      • ( \Delta G^\prime_0 = -64.8 \text{ kJ/mol} )

Page 6: ATP Hydrolysis Driving Unfavorable Reactions

  • Analogy: How gasoline drives a car forward.

  • Similar mechanism: ATP hydrolysis drives reactions just like the chemical energy in gasoline.

Page 7: Mechanism of ATP Hydrolysis in Reactions

  • Example: Glutamine synthetase.

    • Reaction coupled to ATP hydrolysis:

      • ( ATP \rightarrow ADP + Pi + NH3 \rightarrow \text{Glutamine} )

Page 8: Steps in Glutamine Synthetase Reaction

  • Step 1: ATP reacts with glutamate to form a covalent intermediate (mixed anhydride of phosphate and glutamate).

  • Step 2: NH3 acts as a nucleophile:

    • Reacts with electrophilic carbonyl carbon of glutamate.

    • Displaces ( Pi ), forming glutamine.

Page 9: Energy Transfer Mechanism of ATP

  • ATP provides energy through group transfer rather than simple hydrolysis.

  • Phosphate group can participate in different chemical reactions with organic groups.

  • ATP can transfer other moieties:

    • Pyrophosphoryl (PPi) or adenylate (AMP) group.

Page 10: Other High-Energy Compounds

  • Comparison of compounds with large free energies of hydrolysis:

    Compound

    ΔG°' hydr (kJ/mol)

    ATP

    -30

    Glucose 6-phosphate

    -50

    Inorganic phosphate (Pi)

    -10

    Phosphoenolpyruvate

    -61

    Creatine Phosphate

    -43

    1,3-Bisphosphoglycerate

    -50

    Acetyl-CoA (thioester)

    -31

Page 11: Short Answer Question on GTP vs. ATP

  • ATP and GTP share similar ΔG0′ of hydrolysis (approx. -30 kJ/mol).

  • Both serve as energy sources; their similarities suggest a shared functional role in metabolic processes.

    • Reactions: ( GTP \rightarrow GDP + Pi ) and ( ATP \rightarrow ADP + Pi )

Page 12: Introduction to Metabolism

  • Metabolism consists of two processes:

    • Catabolism: Breakdown of large molecules into simpler products.

    • Anabolism: Synthesis of larger, complex molecules from simpler precursors.

      • Involves: ADP and oxidized cofactors; ATP and reduced cofactors.

Page 13: Common Themes in Metabolism

  • Catabolic pathways converge:

    • Sugars, fats, and amino acids convert to acetyl coenzyme A (acetyl CoA).

    • Central role of Acetyl CoA in cellular metabolism.

Page 14: Divergent Anabolic Pathways

  • Acetyl CoA can synthesize:

    • Fatty acids

    • Steroids

    • Components of proteins and nucleic acids.

Page 15: Structure of Metabolic Pathways

  • Metabolic pathways involve enzyme-catalyzed reactions to convert a precursor (A) into a product (E) through intermediates:

    • Each step contributes a small chemical change.

    • Typically contains:

      • At least one thermodynamically favorable irreversible reaction.

      • Regulatory mechanisms (transcription control, enzyme inhibition).

Page 16: Elucidating Metabolic Pathways

  • Metabolic inhibitors:

    • Inhibiting enzymes results in accumulation of intermediates, aiding identification.

  • Genetic diseases can illustrate pathways:

    • Example: Alkaptonuria - urine darkens due to homogentisic acid accumulation.

Page 17: Use of Auxotrophic Mutants

  • Inactivation of specific genes to create auxotrophic mutants:

    • Requires presence of pathway end-products for growth.

    • Accumulation of unmetabolized intermediates aids in identifying the affected pathway.

Page 18: Using Radioactively Labeled Substrates

  • Radioactive labels (e.g., 14C) can trace incorporation into metabolic products.

  • Detection of radioactive compounds assists in tracing pathways through complex mixtures.