11 -

• Oxidative Phosphorylation


  The process in which ATP is formed as a result of the transfer of electrons from NADH or FADH2 to O2 by a series of electron carriers
• Reduced cofactors NADH/FADH2: 10 NADH 2 FADH2
• Mitochondria


  • Inner mitochondrial membrane (inner face)
  • 4208 particles/µm2
• The respiratory complexes


  1. Electrons are transferred from NADH/FADH2 to O2

  2. The free energy released during electron flow through each complex is sufficient to drive ATP synthesis

  3. The complexes do NOT make ATP

  4. They sequester the necessary energy by pumping protons into the intermembrane space
• NADH➞ cpx I ➞ cpx III ➞ cpx IV ➞ O2

  FADH2 ➞ cpx II ➞ cpx III ➞ cpx IV ➞ O2
• ΔG°'


  • 220 kJ/mol
• ΔE°'


  + 1.14 V
• Complex I: NADH-Q oxidoreductase


  • NADH delivers electrons
  • Received by flavin
  • Chain of iron sulphur clusters
  • Delivered to ubiquinone
  • 4 protons pumped across membrane
• Iron Sulphur Clusters


  • Red: iron

  Yellow, Sulphur in Cys

  Green: Sulphur
• Complex III: cyt bc1 or Q cyt c oxidoreductase


  • Ubiquinone delivers electrons
  • Like Cpx I mini electron transport chain inside Cpx III
  • Carriers iron sulphur complexes and cytochromes
  • Delivered to cytochrome C
  • Relay of 2 electron to 1 electron carrier called Q cycle
  • 4 protons pumped across membrane
• Hemes


  • Found in cytochromes
  • Accepts one electron only
  • Accepts and delivers 2 electrons, one at a time
• The Q cycle: how to get 2 e- to Cyt C


  QH2 ➞ Q

  Cpx III ➞ Q ➞ FeS ➞ 1e- Cyt C ox ➞ Cyt C red ➞ 1e- Cyt bL ➞ Cyt bH ➞ Q ➞ SQ ➞ 2H+ ➞ QH2
• Complex IV: Cytochrome C oxidase


  • Cyt C delivers electrons

  Again mini electron transport chain inside Cpx III

  Carriers iron cytochromes and cupper

  Delivered to oxygen: the final acceptor, reduced to water

  2 protons pumped across membrane
• Complex II: Succinate – Q reductase


  • Directly linked to TCA cycle: Succinate is oxidised to fumerate delivering e- to FAD

  FADH2 starts mini electron transport chain

  Chain of iron sulphur clusters

  Delivered to ubiquinone

  NO protons pumped across membrane
• The respiratory super-complexes complexes


  • Complexes exist individually

  Complexes also form super-complexes, respirasomes

  Adapter proteins are needed

  Advantage more efficient transfer

  But too effective might result in reactive oxygen species

  We don't know much about how this is regulated
• ATP synthesis


  Electrons are transported and protons are pumped into the intermembrane space
• Chemiosmotic theory


  The primary energy-conserving event induced by e- transport is the generation of a proton-motive force across the inner mitochondrial membrane
• Chemiosmotic coupling


  All you can do with a proton gradient across an H+ - impermeable membrane:
• Proton-motive force


  • Pumping of protons in the intermembrane space generates:

  A concentration gradient: ΔpH ~ -1.4

  A transmembrane potential: Em ~ 0.14 V

  Both contribute to the proton-motive force, Δp
• Complex V: F0-F1 ATP synthase


  • F1 is the soluble portion

  Without F1, F0 pumps protons but does not make ATP

  Isolated F1 hydrolyses ATP (reverse reaction)

  ATP synthesis requires F0 (proton pumping) and F1 (ATPase) to be coupled

  ATP is formed in the catalytic site of F1 but it is not released unless H+ flow through F0
• High [H+] Low [H+] F0-F1 ATP synthase


  F0 rotates in membrane due to H+ flow

  H+ move from entry to exit channel

  F0 rotates shaft, γ

  F1 is multimer, α and β associate to form a hexamer, the β subunits contain the catalytic sites

  Rotation of γ causes changes in the conformation of the β catalytic sites
• Only γ rotates, β is static but changes conformation when γ rotates
• Rotation of γ


  1. ADP and Pi bound
  2. ATP formed and held
  3. bind and release
• Control of respiration


  The need for ATP is the key control point for oxidative phosphorylation

  Electrons do not flow from NADH to O2 unless ATP is being synthesised

  The level of ADP determines the rate
• Uncoupling of the electron transport chain and ATP synthesis, resulting in no ATP synthesis, has been a pharmacological target for decades to combat obesity, but isn't currently used because it doesn't lead to the expected weight loss, electron transport shuts down, temperature of the individual can increase dangerously, it doesn't encourage lifestyle changes
• Natural uncoupling to generate heat


  • Thermogenin in animals, found in mitochondria of brown fat cells, natural uncoupler, maintains body temperature e.g., in babies and during wake up from hibernation

  Alternative oxidase in plants, oxidises QH2 and generates heat, used by some species to increase temperature of specific organs i.e., flowers or to germinate early
• NADH Shuttles


  The inner mitochondrial membrane is impermeable to NADH

  Two shuttles transport e- from NADH: glycerol phosphate shuttle, malate shuttle
• The glycerol phosphate shuttle


  • carried by FADH2 onto Q

  So, complex I is bypassed, therefore only 6H+ pumped instead of 10

  And less ATP is made from cytoplasmic NADH
• The Malate Shuttle when energy must be conserved


  • Used in heart and liver

  Complex redox shuttle

  Net reaction is NADH moved to mitochondria

  Enter via complex I
• Nucleotide Carriers


  • Two antiporters:

  Pi-OH- antiporter

  ATP-ADP antiporter

  1 H+ is spent to import ADP+Pi and export ATP from the matrix
• ~10 protons are pumped per NADH oxidised

  ~6 protons are pumped per FADH2 oxidised
• Stoichiometry


  • Cpx III: 4 H+

  Cpx IV: 4 H+

  Cpx I: 2 H+

  Cpx II: 0 H+

  ATP synthase: ~3 H+ per ATP

  Nucleotide transport: ~1 H+ per ATP

  ~4 H+ / ATP
• 2.5 ATP made for each NADH

  1.5 ATP made for each FADH2
• ~30 ATP are generated per glucose molecule
• Efficiency of respiration


  Assuming ΔG for ATP hydrolysis in vivo ~-50 kJ/mol

  ΔG for the cellular oxidation of glucose of about -2900 kJ/mol

  ΔG for 30 ATP hydrolysis in vivo -50 kJ/mol * 30 = -1500 kJ/mol

  Efficiency 1500/2900 * 100 = 52%