Chapter 19: Oxidative Phosphorylation sections 19'119'3

1
Chapter 19 Oxidative Phosphorylation(sections
19.1-19.3)

• Dr. Clower
• Chem 4203

2
Oxidative Phosphorylation

• Final step of cellular respiration
• Convergence of pathways
• Reduced cofactors oxidized
• Respiratory chain
• Oxidation coupled to phosphorylation
• Occurs in mitochondria
• Chemiosmotic theory

3
Mitochondrial anatomy

• Elliptical
• Two membranes
• Matrix

4
Oxidation-Reduction Reactions (review)

• Oxidation
• Reduction
• Oxidizing agent
• Reducing agent
• Example reaction 8 of citric acid cycle (malate
  to oxaloacetate)

5
Electrochemistry

• Thermodynamics of electron transport
• Half-reactions
• Oxidation (loss of e-)
• Reduction (gain of e-)
• Standard reduction potential, E
• E at pH 7
• Affinity of electron acceptor for electrons
• Relative to H standard (0.00 V)
• Written as reduction half-reactions
• Electrons flow from species with lower E to
  higher E
• Reduction half-reaction has higher E
• DE sum of E values for each half-reaction
• DG -nFDE

6
Standard Reduction Potentials
7
Universal Electron Acceptors

• Act as electron carriers
• In the form of H atoms (H e-) or hydride ions
  (H 2e-)
• Water-soluble
• Undergo reversible oxidation and reduction
• Function catalytically (regenerated)
• Reduced substrate NAD ? oxidized substrate
  NADH H
• NADP ? NADPH H
• FAD ? FADH2
• FMN ? FMNH2
• Q ? QH2
• Iron-containing proteins

8
Nicotinamide Nucleotides

• NAD, NADP
• Pyridine nucleotides
• Derived from niacin
• Water soluble
• indicates charge on N
• Move from enzyme to enzyme
• High concentration of NAD (favored in oxidation
  reactions) and NADPH (reduction reactions)

PDB ID 3LDH
9
(No Transcript)
10
Flavin Nucleotides

• FMN, FAD
• Flavin nucleotides
• Derived from riboflavin
• Tightly bound to enzymes (flavoproteins)
• Sometimes covalently
• More diverse set of reactions because of ability
  to accept one or two electrons (form FADH or
  FADH2)
• Different values of E depending on specific
  flavoprotein

11
Ubiquinone

• Coenzyme Q
• Lipid soluble
• Benzoquinone with isoprenoid side chain

12
Cytochromes
PDB ID 1CCR
13
(No Transcript)
14
Iron-sulfur Proteins
PDB ID 1FRD
15
Sequence of Electron Carriers
16
Sequence of Electron Carriers
17
Complexes of the Respiratory Chain
Digitonin
18
Electron transfer to Q
19
Complex I NADH to Ubiquinone

• aka NADHubiquinone oxidoreductase or NADH
  dehydrogenase
• Catalyzes two processes
• Transfer of hydride from NADH and H from matrix
  to Q (exergonic) NADH H Q ? NAD QH2
• Transfer of 4 protons from matrix to
  intermembrane space (endergonic) 4H(matrix) ?
  4H(intermembrane)
• We can rewrite equations
• NADH HN Q ? NAD QH2
• 4HN ? 4HP
• And overall reaction is
• NADH 5HN Q ? NAD QH2 4HP

20
Complex II Succinate to Ubiquinone

• aka succinate dehydrogenase
• Subunits
• C and D
• A and B
• Path of electrons

PDB ID 1NEK
21
6. Oxidation of succinate to fumarate

• Dehydrogenation (loss of H2 oxidation)
• Stereospecific to form trans double bond only
• Catalyzed by succinate dehydrogenase complex
• aka succinate dehydrogenase
• aka Complex II
• Embedded in inner mitochondrial membrane, rather
  than in mitochondrial matrix
• Oxidation of alkane requires stronger oxidizing
  agent than NAD (hence FAD)
• FADH2 produced is re-oxidized by coenzyme
  ubiquinone (Q) to reform FAD and ubiquinol (QH2)
• Competitive inhibitor malonate
• -O2C-CH2-CO2-
• Binds to active site through carboxylate groups
• Cannot undergo dehydrogenation
• Inhibition reactions used by Krebs to determine
  citric acid cycle reaction sequence
• Symmetrical molecule evenly distributes carbons
  in remainder of products throughout the cycle

22
Complex III Ubiquinol to Cytochrome c

• aka cytochrome bc1 complex or ubiquinonecytochro
  me c oxidoreductase

QN inhibitor antimycin A QP inhibitor
myxathiazol
PDB ID 1BGY
23
Q cycle
24
Complex IV Cytochrome c to O2

• aka cytochrome oxidase

PDB ID 1OCC
25
Electron Flow (Complex IV)

• Overall reaction
• 4 cyt c (red) 8 HN O2 ? 4 cyt c (ox) 4 HP
  2 H2O
• Or, for each pair of electons
• 2 cyt c (red) 4HN ½ O2 ? 2 cyt c (ox) 2
  HP H2O

26
Summary of Reactions

• Complex I
• NADH 5HN Q ? NAD QH2 4HP
• Complex III
• QH2 2 cyt c1 (ox) 2 HN ? Q 2 cyt c1 (red)
  4 HP
• Complex IV
• 2 cyt c (red) 4HN ½ O2 ? 2 cyt c (ox) 2
  HP H2O

27
Summary, cont.

• Protons transferred from matrix per electron pair
• Overall, for each pair of electrons
• NADH 11HN ½ O2 ? NAD 10HP H2O
• Or, NADH H ½ O2 ? NAD H2O

28
Thermodynamics
(When both are written as reduction half
reactions)

• DE Eelectron acceptor - Eelectron donor
• NAD H 2e- NADH E -0.320 V
• ½ O2 2H 2e- H2O E 0.816 V
• ½ O2 NADH H H2O NAD
• DE (0.816 V) (-0.320 V) 1.136V
• DG -nFDE
• -(2 mol e-/mol reactant)(96.5 kJ/V mol
  e-)(1.136 V)
• -219 kJ/mol reactant
• So, net reaction is highly exergonic

29
Thermodynamics, cont.

• Energy conserved in proton gradient
• For transport of charged species across
  membranes,
• where C2 and C1 are high and low concentrations
  of ions Z is absolute value of charge F is the
  Farraday constant ?? is potential across membrane

30
Thermodynamics, cont.

• For H transport at 25 C
• So,
• In respiring mitochondria, ?? ? 0.15 to 0.20 V
  and ?pH ? 0.75
• Under these conditions, ?G ? 20 kJ/mol H
• Multiply by 10 mol H (pumped when 2 mol e-
  transfer) 200 kJ conserved in proton gradient

31
Next

• Synthesis of ATP

32
ATP Synthesis

• Energy from electron transfer reactions conserved
  in proton gradient
• Chemical (concentration)
• Electrical (charge)
• This energy is used to drive ATP synthesis
• Chemiosmotic model

33
Chemiosmotic Model
ADP Pi nHP ? ATP H2O nHN
34
Coupling of Electron Transfer and ATP Synthesis
Table 19-4
35
(No Transcript)
36
Experimental Support
37
ATP Synthase Complex
PDB ID 1BMF and 1QO1
38
ATP Synthase Complex
39
ATP Synthase
40
Rotational Catalysis
41
Adenine Nucleotide and Phophate Translocases
Table 19-4
42
How do electrons get into the matrix?

• Inner mitochondrial membrane is impermeable to
  NADH
• Two possible methods to transfer reducing
  equivalents through membrane
• Liver, kidney, heart
• Electrons are instead transferred to malate
• Malate-aspartate shuttle
• Skeletal muscle, brain
• Glycerol 3-phosphate shuttle

43
Malate-aspartate Shuttle
44
Glycerol 3-Phosphate Shuttle
45
Stoichiometry of Coupling

• How many ATP are synthesized when electrons pass
  through the respiratory chain?
• P/O ratio (aka P/2e- ratio)
• Experimental values of P/O
• Difficult to measure since ATP and O2 are
  involved in many reactions in mitochondria
• Values between 2 and 3 when NADH is electron
  donor
• Values between 1 and 2 when succinate is electron
  donor
• Textbooks/literature often use values of 3 and 2
• We will use values of 2.5 and 1.5
• Ratio of protons pumped outward by proton
  transfer to protons that flow in through FoF1
  complex to synthesize one ATP
• NADH Succinate

46
ATP Yield from Glucose Oxidation

• How many ATP are produced from the complete
  oxidation of 1 glucose molecule to CO2, assuming
  NADH enters the mitochondrion via the
  malate-aspartate shuttle?
• How many ATP are produced from the complete
  oxidation of 1 glucose molecule to CO2, assuming
  NADH enters the mitochondrion via the glycerol
  3-phosphate shuttle?

47
ATP Yield from Glucose Oxidation
48
Regulation of Oxidative Phosphorylation

• Acceptor Control
• O2
• Regulation of ATP-producing pathways

PDB ID 1OHH