ATP Synthesis

• The chemiosmotic model describes the coupling of ATP synthesis to an electrochemical proton gradient (the proton motive force).
• ATP synthase drives the synthesis of ATP as protons flow passively back into the matrix through its proton pore.
• To emphasize the role of the proton motive force, the equation for ATP synthesis is:

  • ADP + Pi + nH+P --> ATP + H2O + nH+N
• The chemiosmotic model:

  • supported by the observation that, when isolated mitochondria have an artificial electrochemical gradient imposed on them, electrons flow through the respiratory chain
  • Requires an intact outer membrane
  • Requires that mitochondrial ATP synthesis and electron flow through the respiratory chain be obligately coupled
  • Is supported by the observation the isolated mitochondria actively respiring cause the pH of the solution they are in to increase. 


  Requires that mitochondrial ATP synthesis and electron flow through the respiratory chain be obligately coupled
• Inhibitors of electron transfer block ATP synthesis
• Inhibitors of electron transfer block ATP synthesis, unless oxidation and phosphorylation are uncoupled.
• The addition of oligomycin, an inhibitor of ATP synthase, to mitochondria suspended in a buffered medium blocks both ATP synthesis and respiration. What would happen if 2,4-dinitrophenol (DNP) were also added to the suspension?

  • Respiration and ATP synthesis would both resume
  • Respiration would resume without ATP synthesis
  • ATP synthesis would resume without respiration
  • Neither respiration nor ATP synthesis would resume

  Respiration would resume without ATP synthesis.
• Dinitrophenol (DNP) uncouples respiration from ATP synthesis by entering the matrix in the protonated form and releasing a proton, thus dissipating the proton gradient. This allows respiration to continue without ATP synthesis.
• Physical shear or detergent uncouples electron transfer from ATP synthesis.
• DNP and FCCP are weak acids that carry proteins across the inner mitochondrial membrane. This dissipates the proton gradient
• In the absence of an oxidizable substrate, the proton motive force alone drives ATP synthesis.
• ATP synthase has two functional domains, F0 and F1
• Mitochondrial ATP synthase is an F type ATPase. They are similar in structure and mechanism to the ATP synthases of bacteria and chloroplasts
• Mitochondrial ATP synthase contains two distinct components:

  • F0 is an integral membrane protein with a proton pore
  • F1 (originally F1 ATPase) is a peripheral membrane protein that catalyzes hydrolysis of ATP when isolated.
• On the surface of F1, the reaction ADP + Pi --> ATP + H2O is readily reversible.
• ATP synthase stabilizes ATP relative to ADP + Pi by binding ATP more tightly, releasing enough energy to counterbalance the cost of making ATP.
• What is the major energy barrier for ATP synthase?


  Release of ATP from the enzyme. In a typical enzyme catalyzed reaction, reaching the transition state between substrate and product is the major energy barrier to overcome. In the reaction catalyzed by ATP synthase, release of ATP from the enzyme, not the formation of ATP, is the major energy barrier.
• The P/O ratio is

  • about 2.5 when electrons enter the respiratory chain at complex I
  • about 1.5 when electrons enter at ubiquinone
• The P/O ratio varies among species, depending on the number of c subunits in the F0 complex
• Succinate dehydrogenase is dysfunctional in a species of garden slug. While its metabolism is compromised on a number of levels, it can still undergo oxidative phosphorylation. What is the maximal P/O ratio for these organisms if NADH is used as an electron source?

  • 1
  • 1.5
  • 2
  • 2.5
  • 4

  2.5. If 10 protons are pumped out per NADH and 4 must flow in to produce 1 ATP, the proton based P/O ratio is 2.5 for NADH as an electron donor.
• The proton motive force energies active transport.
• Adenine nucleotide translocase is the antiporter that moves ADP into the matric and ATP out.
• Phosphate translocase promotes symport of one H2PO4- and one H+ into the matrix.
• ATP synthasome is a complex of ATP synthase, phosphate translocase and adenine nucleotide translocase.
• The shuttle system indirectly convey cytosolic NADH into mitochondria for oxidation.
• The inner mitochondrial membrane is impermeable to NADH and NAD.
• NADH equivalents are moved from the cytosol to the matrix by the malate asparate shuttle and the glycerol 3-phosphate shuttle.