MCB 2000 Exam 5

• Pyruvate dehydrogenase complex


  Catalyzes the conversion of pyruvate to acetyl CoA
• Enzymes and coenzymes involved in the conversion of pyruvate to acetyl CoA


  • Enzymes: Pyruvate dehydrogenase, Dihydrolipoyl transacetylase, Dihydrolipoyl dehydrogenase
  • Coenzymes: Thiamine pyrophosphate (TPP), Lipoic acid, FAD, NAD+, Coenzyme A
• Conversion of pyruvate to acetyl CoA

  1. Decarboxylation

  2. Oxidation

  3. Acyl transfer to CoA
• Pyruvate dehydrogenase


  Catalyzes the decarboxylation of pyruvate
• Thiamine pyrophosphate (TPP)


  Coenzyme derived from the vitamin thiamine, involved in the decarboxylation step
• Lipoamide


  Derivative of lipoic acid, involved in the oxidation step
• Dihydrolipoyl transacetylase (E2)


  Catalyzes the transfer of the acetyl group from acetyl-lipoamide to coenzyme A to form acetyl CoA
• Dihydrolipoyl dehydrogenase (E3)


  Catalyzes the reoxidation of dihydrolipoamide
• Pyruvate dehydrogenase complex is regulated by

  • Allosteric regulation (inhibition by acetyl CoA and NADH)
  • Covalent modification (phosphorylation of E1 by kinase, dephosphorylation by phosphatase)
• Defects in the regulation of pyruvate dehydrogenase

  Can lead to lactic acidosis (phosphatase deficiency) and beriberi (thiamine deficiency)
• Advantages of organizing the enzymes of the pyruvate dehydrogenase complex into a single large complex


  • Flexible linkages allow rapid movement of substrates and products between active sites
  • Lipoamide arm allows rapid transfer of acetyl group between active sites
• Purposes of the two stages of the Citric Acid Cycle


  • Stage 1: Harvest high-energy electrons from carbon fuels
  • Stage 2: Replenish cycle components and regulate the cycle
• Redox potential (E0')


  Measure of a molecule's tendency to donate or accept electrons
• ΔG0' = -nFΔE0'
• Electron transport chain


  Composed of 4 large protein complexes that capture the energy of high-energy electrons to synthesize ATP
• Key components of the electron transport chain

  • NADH, FADH2 (electron donors)
  • O2 (electron acceptor)
  • FMN, Fe-S proteins, cytochromes (electron carriers)
  • Coenzyme Q (mobile electron carrier)
• Electron flow through the electron transport chain


  1. Complex I (NADH-Q oxidoreductase)

  2. Complex II (Succinate-Q reductase)

  3. Complex III (Q-cytochrome c oxidoreductase)

  4. Complex IV (Cytochrome c oxidase)
• Q cycle


  Mechanism for coupling electron transfer from QH2 to cytochrome c while pumping protons
• Proton gradient is established by proton pumping complexes (I, III, IV) in the electron transport chain
• Complex II (succinate-Q reductase) is not a proton pump
• Anaplerotic reactions

  Replenish the Citric Acid Cycle components when the energy status of the cell changes
• Pyruvate carboxylase

  Catalyzes an anaplerotic reaction that synthesizes oxaloacetate from pyruvate
• Cytochrome c catalyzes the reduction of O2 to 2 molecules of H2O

  1. 8 protons are removed from the matrix

  2. 4 protons used to reduce oxygen

  3. 4 protons are pumped into the intermembrane space
• Mobile electron carriers of the ETC

  Coenzyme Q (Q, ubiquinone, CoQ10)
• Coenzyme Q

  • Binds protons (QH2, ubiquinol) as well as electrons and can exist in several oxidation states
  • Q and QH2 are present in the inner mitochondrial membrane → Q pool
• Proton gradient establishment in the electron-transport chain

  1. Complexes pump protons out of the mitochondria, which generate a proton gradient

  2. Proton pumping complexes: Complex I, Complex III, Complex IV

  3. Complex II is not a proton pump (succinate-Q reductase)

  4. Delivers electrons from FADH2 to Complex III via ubiquinone
• Q cycle


  • Mechanism for coupling electron transfer from QH2 to cytochrome c
  • 4 protons are pumped out of the mitochondria and 2 more are removed from the matrix
  • 2 molecules of QH2 create 2 reduced cytochrome c molecules and 1 new molecule of QH2
• Q pool


  • Q and QH2 are present in the inner mitochondrial membrane
  • Oxidized → Q
  • Reduced Q → QH2
• Cytochrome c oxidase facilitates the reduction of O2 to two molecules of H2O

  1. Accepts 4 electrons from 4 molecules of cytochrome c

  2. 4 protons used to reduce oxygen

  3. 4 additional protons pumped into the intermembrane space
• Reactive oxygen species (ROS)


  • Highly reactive oxygen derivatives created by the partial reduction of O2
  • Superoxide ion, peroxide ion, hydroxyl radical
  • Implicated in many pathological conditions
• Enzymes that mitigate potential damage caused by ROS

  • Superoxide dismutase and catalase
• 1. 4% of oxygen molecules consumed by mitochondria are converted into superoxide ions
• Proton-motive force


  • Proton gradient generated by the oxidation of NADH and FADH2
  • More than just a chemical gradient
  • Proton motive force = chemical gradient + charge gradient
  • Δp =ΔpH + ΔΨ
• ATP synthase structure


  • F0 - embedded in the inner mitochondrial membrane, contains the proton channel and c-ring (rotor)
  • F1 - protrudes into the matrix, contains the active sites
  • 3 catalytic β subunits exist in 3 forms: O (open), L (loose), T (tight)
  • Rotation of γ subunit interconverts the β subunits
  • ATP synthases bind to one another to form dimers, contributing to the formation of cristae (folds in mitochondria)
  • Dimers oligomerize for additional stability (bind with additional peptides)
• Proton flow through ATP synthase leading to ATP synthesis


  1. Proton flow around the c ring powers ATP synthesis

  2. Subunit a has 2 half-channels, one facing intermembrane space and one facing matrix

  3. Protons enter the half-channel facing the intermembrane space, bind to a glutamate residue on a c subunit, and leave the c subunit once they rotate to face the matrix half channel

  4. The force of the proton gradient powers the rotation of the c ring

  5. Rotation of the c ring powers the movement of the γ subunit, which in turn powers the movement of the β subunits

  6. The number of c subunits in the ring determines the number of protons required to synthesize a molecule of ATP

  7. Actual ATP synthesis occurs due to the transition from L to T binding site
• Transfer of electrons from cytoplasmic NADH into the mitochondria

  1. Glycerol phosphate shuttle (muscle)

  2. Malate-aspartate shuttle (heart and liver)
• Acceptor control


  • The regulation of oxidative phosphorylation by ADP
  • Also called respiratory control
  • Example of control of metabolism by energy charge
• Uncoupling leading to heat generation

  1. If electron transport is uncoupled from ATP synthesis, heat is generated

  2. Uncoupling is facilitated by UCP-1 (uncoupling protein 1), also called thermogenin
• Inhibition of oxidative phosphorylation


  1. Inhibition of ETC prevents oxidative phosphorylation by inhibiting the formation of the proton-motive force

  2. Inhibition of ATP synthase by inhibiting proton flow prevents electron transport

  3. Uncouplers carry protons across the inner mitochondrial membrane, so ETC functions but proton gradient never forms (leakiness)

  4. Inhibition of ATP-ADP translocase prevents oxidative phosphorylation