BIO462 - BIOCHEMISTRY

Electron Transport
And
Oxidative Phosphorylation

Lesson Learning Outcomes

Upon completion of this lecture, students should be
able to:

• understand the electron transport chain

• understand the oxidative phosphorylation
Lecture Outline
1. The Role of Electron Transport in Metabolism

2. Reduction Potentials in the Electron Transport Chain

3. Organization of Electron Transport Complexes

4. The Connection between Electron Transport and Phosphorylation

5. The Mechanism of Coupling in Oxidative Phosphorylation

6. Shuttle Mechanisms

7. The ATP Yield from Complete Oxidation of Glucose

The Role of Electron Transport in Metabolism –
Electron Transport Chain (ETC)
• Electron transport is carried out by four closely
related multisubunit membrane-bound complexes
and two electron carriers, coenzyme Q and
cytochrome c
• In a series of oxidation-reduction reactions, electrons
from FADH2 and NADH are transferred from one
complex to the next until they reach O2
• O2 is reduced to H2O
O2 + 4 H+ + 4 e - 2 H2 O

• As a result of electron transport, protons are pumped

across the inner membrane to the intermembrane
space, creating a pH gradient.
ATP Production in the Mitochondrion

• The production of ATP in the mitochondria is the

result of oxidative phosphorylation

• The proton gradient establishes a voltage gradient

• The proton and voltage gradients together provide

the mechanism to couple electron transport with
phosphorylation of ADP
Establishment of the Proton Gradient
Summary - ETC
• Electron transport from one
carrier to another creates a
proton gradient across the
inner mitochondrial membrane

• The proton gradient is coupled

to the production of ATP in
aerobic metabolism
• Schematic representation of the electron transport chain,
showing sites of proton pumping coupled to oxidative
phosphorylation.

• FMN is the flavin coenzyme flavin mononucleotide, which

differs from FAD in not having an adenine nucleotide.

• CoQ is coenzyme Q

• Cyt b, cyt c1, cyt c, and cyt aa3 are the heme-containing
proteins cytochrome b, cytochrome c1, cytochrome c, and
cytochrome aa3, respectively.
Reduction Potentials in the Electron
Transport Chain
• A useful way to look at electron transport is to
consider the change in free energy associated with
the movement of electrons from one carrier to
another
• If we have two electron carriers, for example NADH
and coenzyme Q, are electrons more likely to be
transferred from NADH to coenzyme Q, or vice versa?
• What we need to know is the reduction potential for
each carrier
• A carrier of high reduction potential will tend to be
reduced if it is paired with a carrier of lower reduction
potential
Reduction Potentials (Cont’d)
Reduction Potentials (Cont’d)
Summary
• Standard reduction potentials provide a basis for
comparison among redox reactions

• The sequence of reactions in the electron transport

chain can be predicted by using reduction potentials.
Organization of Electron Transport
Complexes
• Complex I: NADH-CoQ
oxidoreductase
• Electrons are passed from
NADH to FMN

NADH + H+ E-FMN ----> NAD+

+ E-FMNH2
Oxidized and Reduced Forms of CoQ: The
Ubiquinone
Organization of Electron Transport
Complexes (Cont’d)
• Electrons are then passed to the iron-sulfur clusters
• The last step of Complex I involves electrons being
passed to coenzyme Q (also called ubiquinone)
Energetics of Electron Transport
• The transfer of electrons is strongly exergonic and is
sufficient to drive the phosphorylation of ADP
Electron Transport Complexes (Cont’d)
• Complex II: Succinate-coenzyme Q oxidoreductase

Succinate + E-FAD ---> Fumarate + E-FADH2

• The overall reaction is exergonic (-13.5 kj•mol-1), but

not enough to drive ATP production

• No H+ is pumped out of the matrix during this step

Electron Transport Complexes (Cont’d)
• Complex III: CoQH2-cytochrome c oxidoreductase

CoQH2 + 2Cyt c[Fe(III)]--->CoQ + 2Cyt c[Fe(II)] + 2H+

• This reactions of this complex results in a decrease in free

energy that is sufficient to drive the phosphorylation of ADP to
ATP
• The flow of electrons from reduced CoQ, a quinone that can
exist in 3 forms, is known as the Q cycle
Compositions and Locations of Respiratory
Complexes in Inner Mitochondrial Membrane
Oxidized and Reduced Forms of CoQ
Electron Transport Complexes (Cont’d)
• Complex IV: Cytochrome c oxidase
• Catalyzes the final step in electron transport

2Cyt c[Fe(II)] + 2H+ + 1/2O2 ---> 2 Cyt c[Fe(III)] + H2O

• Complex IV contains cytochrome a, cytochrome a3,

and Cu(II), which are also involved in the electron
transport
• Complex IV is the link to molecular oxygen
The Energetics of Electron Transport Reactions
The Heme Group of Cytochromes
• All cytochromes contain a heme group
• There are differences in the side chain depending on
the heme
Fe-S Bonding in Nonheme Iron Proteins
The Connection between Electron
Transport and Phosphorylation
• The energy-releasing oxidations give rise to proton
pumping and a pH gradient across the inner
mitochondrial membrane
• Differences in the concentration of ions across the
membrane generates a voltage gradient
• A coupling process converts the electrochemical
potential to the chemical energy of ATP
• The coupling factor is ATP synthase, a complex
protein oligomer, separate from the electron transport
complexes
• Uncouplers inhibit the phosphorylation of ADP without
affecting electron transport; examples are 2,4-
dinitrophenol, valinomycin, and gramicidin A
F1 and F0 Components of ATP Synthase
Uncouplers
P/O Ratio
• P/O ratio: the number of moles of Pi consumed in
phosphorylation to the number of moles of oxygen
atoms consumed in oxidation
• Phosphorylation: ADP + Pi ----> ATP + H2O
• Oxidation: 1/2O2 + 2H+ + 2e- ---> H2O

• P/O = 2.5 when NADH is oxidized

• P/O = 1.5 when FADH2 is oxidized
Summary
• The coupling of electron transport to oxidative
phosphorylation requires a multisubunit membrane-
bound enzyme, ATP synthase. This enzyme has a
channel for protons to flow from the intermembrane
space into the mitochondrial matrix.

• The proton flow is coupled to ATP production in a

process that appears to involve a conformational
change of the enzyme.
The Mechanism of Coupling in Oxidative
Phosphorylation
• Chemiosmotic coupling
• based on a proton concentration gradient between the
intermembrane space and the matrix
• a proton gradient exists because the various proteins
that serve as electron carriers are not symmetrically
oriented with respect to the two sides of the inner
mitochondrial membrane
• these proteins take up protons from the matrix when
they are reduced and release them to the
intermembrane space when they are reoxidized
• the reactions of NADH, CoQ, and O2 all require
protons
The Mechanism of Coupling in Oxidative
Phosphorylation(Cont’d)
The Mechanism of Coupling in Oxidative
Phosphorylation (Cont’d)
• Evidence for chemiosmotic coupling suggested by Mitchell
(1961):
• A system with definite inside and outside compartments (closed
vesicles) is essential.

• Submitochondrial vesicles can be prepared, which carry out

oxidative phosphorylation and have an asymmetric orientation of
respiratory complexes (Figure 20.16).

• A model system for oxidative phosphorylation can be

constructed with proton pumping in the absence of electron
transport; the model system consists of reconstituted membrane
vesicles, mitochondrial ATP synthase, and a proton pump
(Figure 20.17).

• The existence of the pH gradient has been demonstrated and

confirmed experimentally
The Mechanism of Coupling in Oxidative
Phosphorylation (Cont’d)
• The mechanism by which the proton gradient leads to
the production of ATP depends on ion channels
through the inner mitochondrial membrane
• Protons flow back into the matrix through channels in
the F0 unit of ATP synthase
• The flow of protons is accompanied by formation of
ATP in the F1 unit of ATP synthase
• The details of how phosphorylation takes place as a
result of the linkage to the proton gradient are not
explicitly specified by this mechanism
The Mechanism of Coupling in Oxidative
Phosphorylation (Cont’d)
Conformational Coupling
• The proton gradient leads to changes in conformation
in a number of proteins, including ATP synthase
• There are 3 sites for substrate on ATP synthase, and 3
possible conformations:
- Open (O); a low affinity for substrate
- Loose-binding (L); not catalytically active, binds ADP
and Pi
- Tight-binding (T); catalytically active, binds ATP
• These sites interconvert as a result of proton flux
through ATP synthase
• Proton flux converts L to T, which produces ATP
• Proton flux converts T to O, releasing ATP
Release of ATP from ATP Synthase
Summary
• In chemiosmotic coupling, the proton gradient is the
crux of the matter. The flow of protons through pore
in the synthase drives ATP production.

• In conformational coupling, a change in the shape of

the synthase releases bound ATP that has already
been formed.
Shuttle Mechanisms
• Shuttle mechanisms: transport metabolites between
mitochondria and cytosol
• Glycerol phosphate shuttle:
• We know glycolysis in the cytosol produces NADH
• NADH does not cross the mitochondrial membrane,
but glycerol phosphate and dihydroxyacetone
phosphate do
• Through the glycerol phosphate shuttle, 1.5 ATP are
produced in the mitochondria for each cytosolic NADH
The Glycerol-Phosphate Shuttle
The Malate-Aspartate Shuttle
• The Malate-Aspartate Shuttle:
• Has been found in mammalian kidney, liver, and heart
• Malate crosses the mitochondrial membrane, while
oxaloacetate cannot
• The transfer of electrons from NADH in the cytosol
produces NADH in the mitochondria
• In the malate-aspartate shuttle, 2.5 mitochondrial ATP
are produced for each cytosolic NADH
The Malate-Aspartate Shuttle (Cont’d)
Summary
• Shuttle mechanisms transfer electrons, but not
NADH, from the cytosol across the mitochondrial
membrane

• In the malate-aspartate shuttle, 2.5 molecules of ATP

are produced for each molecule of cytosolic NADH,
rather than 1.5 ATP in the glycerol-phosphate shuttle,
a point that affects the overall yield of ATP in these
tissues
The ATP Yield from Complete Oxidation of
Glucose
• In the complete oxidation of glucose, a total of 30 or
32 molecules of ATP are produced for each
molecule of glucose, depending on the shuttle
mechanism
The ATP Yield from Complete Oxidation of
Glucose (Cont’d)