Aerobic Respiration and the Mitochondrion

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CHAPTER 5

• Aerobic Respiration and the Mitochondrion

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Introduction

• The early Earth was populated by anarobes, which
  captured and utilized energy by
  oxygen-independent metabolism.
• Oxygen accumulated in the primitive atmosphere
  after cyanobacteria appeared.
• Aerobes evolved to use oxygen to extract more
  energy from organic molecules.
• In eukaryotes, aerobic respiration takes place in
  the mitochondrion.

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5.1 Mitochondrial Structure and Function (1)

• Mitochondria have characteristic morphologies
  despite variable appearance.
• Typical mitochondria are bean-shaped organelles
  but may be round or threadlike.
• The size and number of mitochondria reflect the
  energy requirements of the cell.

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Mitochondria
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Mitochondrial Structure and Function (2)

• Mitochondria can fuse with one another, or split
  in two.
• The balance between fusion and fission is likely
  a major determinant of mitochondrial number,
  length, and degree of interconnection.

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Mitochondrial fusion and fission
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Mitochondrial Structure and Function (3)

• Inner and outer mitochondrial membranes enclose
  two spaces the matrix and intermembrane space.
• The outer mitochondrial membrane serves as its
  outer boundary.
• The inner mitochondrial membrane is subdivided
  into two interconnected domains
• Inner boundary membrane
• Cristae where the machinery for ATP is located

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The structure of a mitochondrion
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Mitochondrial Structure and Function (4)

• Mitochondrial Membranes
• The outer membrane is about 50 the inner
  membrane is more than 75 protein.
• The inner membrane contains cardiolipin but not
  cholesterol, both are true of bacterial
  membranes.
• The outer membrane contains a large pore-forming
  protein called porin.
• The inner membrane is impermeable to even small
  molecules the outer membrane is permeable to
  even some proteins.

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Porins
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Mitochondrial Structure and Function (5)

• The mitochondrial matrix
• Contains a circular DNA molecule, ribosomes, and
  enzymes.
• RNA and proteins can be synthesized in the matrix.

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Overview of carbohydrate metabolism in eukaryotic
cells
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5.2 Oxidative Metabolism in the Mitochondrion (1)

• The first steps in oxidative metabolism are
  carried out in glycolysis.
• Glycolysis produces pyruvate, NADH, and two
  molecules of ATP.
• Aerobic organisms use O2 to extract more than 30
  additional ATPs from pyruvate and NADH.
• Pyruvate is transported across the inner membrane
  and decarboxylated to form acetyl CoA, which
  enters the next stage.

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An overview of glycolysis
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Oxidative Metabolism in the Mitochondrion (2)

• The tricarboxylic acid (TCA) cycle
• It is a stepwise cycle where substrate is
  oxidized and its energy conserved.
• The two-carbon acetyl group from acetyl CoA is
  condensed with the four-carbon oxaloacetate to
  form a six-carbon citrate.
• During the cycle, two carbons are oxidized to
  CO2, regenerating the four-carbon oxaloacetate
  needed to continue the cycle.

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The TCA cycle
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Oxidative Metabolism in the Mitochondrion (3)

• The TCA cycle (continued)
• Four reactions in the cycle transfer a pair of
  electrons to NAD to form NADH, or to FAD to
  form FADH2.
• Reaction intermediates in the TCA cycle are
  common compounds generated in other catabolic
  reactions making the TCA cycle the central
  metabolic pathway of the cell.

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Catabolic pathways generate compounds that are
fed into the TCA cycle
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Oxidative Metabolism in the Mitochondrion (4)

• The Importance of Reduced Coenzymes in the
  Formation of ATP
• The reduced coenzymes FADH2 and NADH are the
  primary products of the TCA cycle.
• NADH formed during glycolysis enters the
  mitochondria via malate-aspartate or glycerol
  phosphate shuttles.

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The glycerol phosphate shuttle
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Oxidative Metabolism in the Mitochondrion (5)

• The Importance of Reduced Coenzymes
• As electrons move through the electron-transport
  chain, H are pumped out across the inner
  membrane.
• ATP is formed by the controlled movement of H
  back across the membrane through the
  ATP-synthesizing enzyme.

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Oxidative Metabolism in the Mitochondrion (6)

• Reduced coenzymes (continued)
• The coupling of H translocation to ATP synthesis
  is called chemiosmosis.
• Three molecules of ATP are formed from each pair
  of electrons donated by NADH two molecules of
  ATP are formed from each pair of electrons
  donated by FADH2.

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Summary of oxidative phosphorylation
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The Human Perspective The Role of Anaerobic and
Aerobic Metabolism in Exercise (1)

• ATP hydrolysis increases 100-fold during
  exercise, quickly exhausting ATP available.
• Muscles used stored creatine phosphate (CrP) to
  rapidly generate but must rely on aerobic or
  anaerobic synthesis of new ATP for sustained
  activity.
• CrP ADP ? Cr ATP

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The Human Perspective The Role of Anaerobic and
Aerobic Metabolism in Exercise (2)

• Fast-twitch muscle fibers contract rapidly, have
  few mitochondria ad produce ATP anaerobically.
• Anaerobic metabolism produces fewer ATPs per
  glucose but produces them very fast.
• Anaerobic metabolism rapidly depletes available
  glucose and builds up lactic acid which reduces
  cellular pH.

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Skeletal muscles
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The Human Perspective The Role of Anaerobic and
Aerobic Metabolism in Exercise (3)

• Slow-twitch fibers contract slowly, have many
  mitochondria and produce most of their ATP by
  aerobic metabolism.
• Aerobic metabolism initially uses glucose as a
  substrate.
• Free fatty acids are oxidized during prolonged
  exercise.
• Ratio of fast- to slow-twitch fibers is variable
  and depends on the normal function of the muscle.

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5.3 The Role of Mitochondria in the Formation of
ATP (1)

• ATP can be formed by substrate-level
  phosphorylation or oxidative phosphorylation.
• Accounts for gt 160 kg of ATP in our bodies per day

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The Role of Mitochondria in the Formation of ATP
(2)

• Oxidation-Reduction (Redox) Potentials
• Strong oxidizing agents have a high affinity for
  electrons strong reducing agents have a weak
  affinity for electrons
• Redox reactions are accompanied by a decrease in
  free energy.
• The transfer of electrons causes charge
  separation that can be measured as a redox
  potential.

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Redox potential of some reaction couples
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The Role of Mitochondria in the Formation of ATP
(3)

• Electron Transport
• Electrons move through the inner membrane via a
  series of carriers of decreasing redox potential.
• Electrons associated with either NADH or FADH2
  are transferred through specific electron
  carriers that make up the electron transport
  chain.

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The Role of Mitochondria in the Formation of ATP
(4)

• Types of Electron Carriers
• Flavoproteins are polypeptides bound to either
  FAD or FMN.
• Cytochromes contain heme groups bearing Fe or Cu
  metal ions.
• Three cooper atoms are located within a single
  protein complex and alternate between Cu2/Cu3
• Ubiquinone (coenzyme Q) is a lipid-soluble
  molecule made of five-carbon isoprenoid units.

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Structures of three electron carriers
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The Role of Mitochondria in the Formation of ATP
(5)

• Types of Electron Carriers (continued)
• Iron-sulfur proteins contain Fe in association
  with inorganic sulfur.
• These carriers are arranged in order of
  increasingly positive redox potential.
• Sequence of carriers determined by use if
  inhibitors.

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Sequence of electron carriers
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The Role of Mitochondria in the Formation of ATP
(6)

• Electron-Transport Complexes
• Complex I (NADH dehydrogenase) catalyzes transfer
  of electrons from NADH to ubiquinone and
  transports four H per pair.
• Complex II (succinate dehydrogenase) catalyzes
  transfer of electrons from succinate to FAD to
  ubiquinone without transport of H.
• Complex III (cytochrome bc1) catalyzes the
  transfer of electrons from ubiquinone to
  cytochrome c and transports four H per pair.

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The electron-transport chain of the inner
mitochondrial membrane
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The electron-transport chain of the inner
mitochondrial membrane
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The Role of Mitochondria in the Formation of ATP
(7)

• Electron-Transport Complexes (continued)
• Complex IV (cytochrome c oxidase) catalyzes
  transfer of electrons to O2 and transports H
  across the inner membrane.
• Cytochrome oxidase is a large complex that adds
  four electrons to O2 to form two molecules of
  H2O.
• The metabolic poisons CO, N3, and CN bind
  catalytic sites in Complex IV.

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Cytochrome oxidase
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The Role of Mitochondria in the Formation of ATP
(8)

• Cytochrome oxidase
• Electrons are transferred one at a time.
• Energy released by O2 reduction is presumably
  used to drive conformational changes.
• These changes would promote the movement of H
  ions and through the protein.

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5.4 Translocation of Protons and the
Establishment of a Proton-Motive Force (1)

• There are two components of the proton gradient
• The concentration gradient between the matrix and
  intermembrane space creates a pH gradient (?pH).
• The separation of charge across the membrane
  creates an electric potential (?).
• The energy present in both components of the
  gradients is proton-motive force (?p).

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Visualizing the proton-motive force
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Translocation of Protons and the Establishment of
a Proton-Motive Force (2)

• Dinitrophenol (DNP) uncouples glucose oxidation
  and ATP formation by increasing the permeability
  of the inner membrane to H, thus eliminating the
  proton gradient.
• Differences in uncoupling proteins (UCPs) account
  for differences in metabolic rate.

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5.5 The Machinery for ATP Formation (1)

• Isolation of coupling factor 1, or F1, showed
  that it hydrolyzed ATP.
• Under experimental conditions, it behaves as an
  ATP synthase.
• Led to conclusion that an ionic gradient
  establishes a proton-motive force to
  phosphorylate ADP.

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An experiment to drive ATP formation in membrane
vesicles reconstituted with the Na/K-ATPase
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The Machinery for ATP Formation (2)

• The structure of the ATP synthase
• The F1 particle is the catalytic subunit, and
  contains three catalytic sites for ATP synthesis.
• The F0 particle attaches to the F1 and is
  embedded in the inner membrane.
• The F0 base contains a channel through which
  protons are conducted from the intermembrane
  space to the matrixdemonstrated in experiments
  with submitochondrial particles.

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The structure of the ATP synthase
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ATP formation in experiments with
submitochondrial particles
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The Machinery for ATP Formation (3)

• The Basis of ATP Formation According to the
  Binding Change Mechanism
• The binding change mechanism states the
  following
• Movement of protons through ATP synthase alters
  the binding affinity of the active site.
• Each active site goes through distinct
  conformations that have different affinities for
  substrates and product.

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The structural basis of catalytic site
conformation
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The binding change mechanism for ATP synthesis
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The Machinery for ATP Formation (4)

• Binding change mechanism
• Binding sites on the catalytic subunit can be
  tight, loose, or open.
• ATP is synthesized through rotational catalysis
  where the stalk of ATP synthase rotates relative
  to the head.
• There is structural and experimental evidence to
  support this mechanism

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Direct observation of rotational catalysis
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The Machinery for ATP Formation (5)

• Using the Proton Gradient to Drive the Catalytic
  Machinery The Role of the F0 Portion of ATP
  Synthase
• The c subunits of the F0 base form a ring.
• The c ring is bound to ? subunit of the stalk.
• Protons moving through membrane rotate the ring.
• Rotation of the ring provides twisting force that
  drives ATP synthesis.

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A model of the proton diffusion coupled to
rotation of c ring in the F0 complex
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The Machinery for ATP Formation (6)

• Other Roles for the Proton-Motive Force in
  Addition to ATP Synthesis
• The H gradient drives transport of ADP into and
  ATP out of the mitochondrion.
• ADP is the most important factor controlling the
  respiration rate.
• Many factors influence the rate of respiration,
  but the pathways are poorly understood.

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Summary of the major activities during aerobic
respiration in the mitochondrion
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5.6 Peroxisomes (1)

• Peroxisomes are membrane-bound vesicles that
  contain oxidative enzymes.
• They oxidize very-long-chain fatty acids, and
  synthesize plasmalogens (a class of
  phospholipids).
• They form by splitting from preexisting
  organelles, import preformed proteins, and engage
  in oxidative metabolism.

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The structure and function of peroxisomes
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Peroxisomes (2)

• Hydrogen peroxide (H2O2), a reactive and toxic
  compound, is formed in peroxisomes and is broken
  down by the enzyme catalase.
• Plants contain a special peroxisome called
  glyoxysome, which can convert fatty acids to
  glucose by germinating seedlings.

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Glyoxysome localization within plant seedlings
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The Human Perspective Diseases that Result from
Abnormal Mitochondrial or Peroxisomal Function (1)

• Mitochondria
• A variety of disorders are known that result from
  abnormalities in mitochondria structure/function.
• Majority of mutations linked to mitochondrial
  diseases are traced to mutations in mtDNA.
• Mitochondrial disorders are inherited maternally.

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Mitochondrial abnormalities in skeletal muscle
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The Human Perspective Diseases that Result from
Abnormal Mitochondrial or Peroxisomal Function (2)

• It is speculated that accumulations of mutations
  in mtDNA is a major cause of aging.
• In mice encoding a mutation in their mtDNA, signs
  of premature aging develop.
• Additional findings suggest that mutations in
  mtDNA may cause premature aging but are not
  sufficient for the normal aging process.

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A premature aging phenotype caused by mutations
in mtDNA
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The Human Perspective Diseases that Result from
Abnormal Mitochondrial or Peroxisomal Function (3)

• Peroxisomes
• Patients with Zellweger syndrome lack peroxisomal
  enzymes due to defects in translocation of
  proteins from the cytoplasm into the peroxisome.
• Adrenoleukodydstrophy is caused by lack of a
  peroxisomal enzyme, leading to fatty acid
  accumulation in the brain and destruction of the
  myelin sheath of nerve cells.