Chapter 20 Electron Transport and Oxidative Phosphorylation

Major concepts
1. Oxidative phosphorylation is the enzymatic synthesis of ATP coupled to the
transfer of electrons to molecular oxygen.
2. The mitochondrial respiratory chain is an ordered array of electron carriers
arranged in complexes.
3. Complex I is a transmembrane protein complex of the inner membrane.
4. Complex II, or succinate dehydrogenase, transfers electrons from succinate to the
complexs covalently bound FAD and then to its Fe-S centers.
5. Complex III transfers electrons from QH2 to cytochrome c.
6. Cytochrome c is a mobile protein that shuttles electrons, in this case carrying
them from Complex III to Complex IV.
7. The chemiosmotic model explains in molecular terms how the proton gradient
generated by the flow of electrons through the respiratory chain drives the
synthesis of ATP.
8. The proton-motive force also drives other cell activities.

Chapter Outline
20.1 The Role of Electron Transport in Metabolism
What is the importance of mitochondrial structure in ATP production?
20.2 Reduction Potentials in the Electron Transport Chain
How can reduction potentials be used to predict the direction of electron
transport?
20.3 Organization of Electron Transport Complexes
What reactions take place in the respiratory complexes?
What is the nature of the iron-containing proteins of electron transport?
20.4 The Connection between Electron Transport and Phosphorylation
What is the coupling factor in oxidative phosphorylation?
20.5 The Mechanism of Coupling in Oxidative Phosphorylation
What is chemiosmotic coupling?
What is conformational coupling?
20.6 Respiratory Inhibitors Can Be Used to Study Electron Transport
Do respiratory inhibitors have a connection with respiratory complexes?
20.7 Shuttle Mechanisms
How do shuttle mechanisms differ from one another?
20.8 The ATP Yield from Complete Oxidation of Glucose
 SUMMARY

What is the importance of mitochondrial structure in ATP production?

In the final stages of aerobic metabolism, electrons are transferred from NADH
to oxygen (the ultimate electron acceptor) in a series of oxidationreduction reactions
known as the electron transport chain. In the process, protons are pumped across the
inner mitochondrial membrane. This series of events depends on the presence of
oxygen in the final step. This pathway allows for the reoxidation of the reduced
electron carriers produced in glycolysis, the citric acid cycle, and several other
catabolic pathways, and is the true source of the ATPs produced by catabolism.
Phosphorylation depends on the compartmented structure of mitochondria.

How can reduction potentials be used to predict the direction of electron

transport?
The overall reaction of the electron transport chain shows a very large, negative

G ' due to the large differences in reduction potentials between the reactions

involving NADH and those involving oxygen. If NADH were to reduce oxygen

directly, the E ' would be more than 1 V. In reality, many redox reactions are in

between, and the correct order of events in the electron transport chain was predicted
by comparing the reduction potentials of the individual reactions long before the order
was established experimentally.

What reactions take place in the respiratory complexes?

Four separate respiratory complexes can be isolated from the inner mitochondrial
membrane. Each can carry out the reactions of a portion of the electron transport
chain. In addition to the respiratory complexes, two electron carriers, coenzyme Q and
cytochrome c, are not bound to the complexes but are free to move within and along
the membrane, respectively. Complex I accomplishes the reoxidation of NADH and
sends electrons to coenzyme Q. Complex II reoxidizes FADH2 and also sends
electrons to CoQ. Complex III involves the Q cycle and shuttles electrons to
cytochrome c. Complex IV takes the electrons from cytochrome c and passes them to
oxygen in the final step of electron transport.

What is the nature of the iron-containing proteins of electron transport?

 A number of iron-containing proteins are part of the electron transport chain. In
the cytochrome proteins, the iron is bound to a heme group. In other proteins, the iron
is bound to the protein along with sulfur.
What is the coupling factor in oxidative phosphorylation?
A complex protein oligomer is the coupling factor that links oxidation and
phosphorylation. The complete protein spans the inner mitochondrial membrane and
projects into the matrix as well. The portion of the protein that spans the membrane is
called Fo ; it consists of three different kinds of polypeptide chains (a, b, and c).
The portion that projects into the matrix is called F1 ; it consists of five different
kinds of polypeptide chains (, and , in the ratio 33). The F1 sphere is
the site of ATP synthesis. The whole protein complex is called ATP synthase. It is also
known as mitochondrial ATPase. During the process of electron transport, several
reactions occur in which reduced carriers that have both electrons and protons to
donate are linked to carriers that can only accept electrons. At these points, hydrogen
ions are released to the other side of the inner mitochondrial membrane, causing the
formation of a pH gradient. The energy inherent in the charge and chemical separation
of the hydrogen ions is used to phosphorylate ADP to ATP when the hydrogen ions
pass back into the mitochondria through ATP synthase.

What is chemiosmotic coupling?

Two mechanisms, the chemiosmotic mechanism and the conformational
coupling mechanism, have been proposed to explain the coupling of electron transport
and ATP production.
Chemiosmotic coupling is the mechanism most widely used to explain the
manner in which electron transport and oxidative phosphorylation are coupled to one
another. In this mechanism, the proton gradient is directly linked to the
phosphorylation process. The way in which the proton gradient leads to the
production of ATP depends on ion channels through the inner mitochondrial
membrane; these channels are a feature of the structure of ATP synthase. Protons flow
back into the matrix through proton channels in the Fo part of the ATP synthase. The
flow of protons is accompanied by formation of ATP, which occurs in the F1 unit.

What is conformational coupling?

In the conformational coupling mechanism, the proton gradient is indirectly
related to ATP production. Recent evidence appears to show that the effect of the
proton gradient is not the formation of ATP but the release of tightly bound ATP from
the synthase as a result of the conformational change.
Do respiratory inhibitors have a connection with respiratory complexes?
Many of the workings of the electron transport chain have been elucidated by
experiments using respiratory inhibitors. These inhibitors specifically block the
transfer of electrons at specific points in the respiratory complexes. Examples are CO
and CN-, both of which block the final step of the electron transport chain, and
rotenone, which blocks the transfer of electrons from NADH reductase to coenzyme
Q. When such a blockage occurs, it causes electrons to pile up behind the block,
giving a reduced carrier that cannot be oxidized. By noting which carriers become
trapped in a reduced state and which ones are trapped in an oxidized state, we can
establish the link between carriers.

How do shuttle mechanisms differ from one another?

Two shuttle mechanismsthe glycerolphosphate shuttle and the malate
aspartate shuttletransfer the electrons, but not the NADH, produced in cytosolic
reactions into the mitochondrion. In the first of the two shuttles, which is found in
muscle and brain, the electrons are transferred to FAD; in the second, which is found
in kidney, liver, and heart, the electrons are transferred to NAD+. With the malate
aspartate shuttle, 2.5 molecules of ATP are produced for each molecule of cytosolic
NADH, rather than 1.5 ATP in the glycerolphosphate shuttle, a point that affects the
overall yield of
ATP in these tissues. Approximately 2.5 molecules of ATP are generated for each
molecule of NADH that enters the electron transport chain and approximately 1.5
molecules of ATP for each molecule of FADH2. When glucose is metabolized
anaerobically, the only net ATPs that are produced are those from the substrate-level
phosphorylation steps. This leads to a total of only two ATPs per glucose entering
glycolysis. When the pyruvate generated from glycolysis can enter the citric acid
cycle, and the resulting NADH and FADH2 molecules are reoxidized through the
electron transport chain, a total of 30 or 32 ATPs are produced, with the difference
being due to the two possible shuttles.
 Class Note

# ETC OP

ATP

20-01~20-02

The Electron Transport Chain ()

Membrane embedded protein complex serve as series of e- carrier that transfer e-

derived form reduced coenzyme to O2.

Electron Transport Chain (ETC) ATP :

1. NADH 2.5 ATP

2. FADH2 1.5 ATP

ETC ATP synthase mitochondria inner membrane

ETC

Oxidative phosphorylation ()

A process for generating ATP, creation of pH gradient (enzyme complexes

actively pump H+ outward) within mitochondria as a result of electron transfer,
creating energy gradient that drives synthesis of ATP couple to electron transfer,
produced by cytosolic dehydrogenase.

20-03~20-11
5 electron carriers involved in respiratory chain:
Universal e- acceptor:
1. Nicotinamide Nucleotide (NADH, NADPH)
2. Flavin Nucleotide (FADH2, FMNH2)
Mem.-bound carrier:
3. Coenzyme Q (ubiquinone)

CoQ 6-10 isoprene units ()

 Overall rx.: Quinone Semiquinone Hydroquinone

4. Cytochrome protein (b, c1, c, a, a3)

Hemoprotein (heme ), different cytochromes have

different side-substituent of heme to absorb different wavelength of

light.
Mitochondria: Cyt b, c, c1, a, a3; E.coli: Cyt b1; Yeast: Cyt b2;

: Cyt f (chloroplast, f for fron, foliage)

5. Iron-sulfur protein (Fe-S) (Nonheme iron protein, NHI)

Fe ion is bound to S of Cyt. or sulfide (S2-)
Membrane bound multiprotein complexes in the ETC
I, II, III, IV: 4 enzyme complexes, each contains part of the entire respiratory
chain (RC), catalyze transfer between e- donor (NADH, succinate),
intermediate carrier (Q, cyt. c), and terminal acceptor (O2)
V: ATP synthase (ATPase, Adenosine triphosphatase, ATP hydrolase), isolated
ATP synthase has only ATP-hydrolyzing activity.

(): NADH > FADH2 > CoQ > Cyt. b > Cyt. c1 > Cyt. c

> Cyt. a-a3 > O2

20-11~13, 20-15~20-17
Respiratory Chain:
1. Complex I (NADH-CoQ oxidoreductase)

NADH + H+ + CoQ NAD+ + CoQH2

2. Complex II (succinate-CoQ oxidoreductase)

Succinate + CoQ Fumarate + CoQH2 (Fe3+ + e- Fe2+)

3. Complex III (CoQ-cytochrome c oxidoreductase)

CoQH2 + 2 Cyt c (Fe3+) CoQ + 2 Cyt c (Fe2+) + 2 H+

4. Complex IV (cytochrome oxidase): Final step of ETC

2 Cyt c (Fe2+) + 2 H+ + 1/2 O2 2 Cyt c (Fe3+) + H2O

Sites of ATP formation in ETC: NADH CoQ (Complex I); Cyt b c1

 (Complex III); Cyt a-a3 O2 (Complex IV)

20-13~20-14, 20-18~20-21
ATP synthase (Complex V)

Chemiosmotic coupling hypothesis ()

Complex I, III, IV matrix proton pump

intermembrane space, inner membrane asymmetric
, intermembrane space , matrix (I-P, M-N)
Intermembrane space matrix chemical gradient electrical
gradient, proton-motive force (PMF)PMF ATP
PMF intermembrane space proton
ATPase Fo (H+-specific channel) matrix F1 ATP
ATP Synthase functional domain : F1 FOF1 domain
3 & 3 subunits, unit (central shaft) ATP
Fo H+-specific channel
F1 domain ATP high affinity binding energy ATP
domain ATP F1 domain
proton motive force
Rotational catalysisH+ Fo domain pore F1 subunit
(ATP binding, ADP + Pi, Empty)
subunit subunit subunit ATP
ATP synthasome: ATP synthase + Adenine nucleotide translocase + phosphate
translocase, tightly intergrated.
20-21
What do they stand for ?
1) Fo o Oligomycin sensitive
2) F1 1 The first factor isolated from mitochondria
3) F-type ATPase F Energy coupling Factor
4) V-type ATPase V Vacuole ()
5) P-type ATPase P Plasma membrane
6) AoA1 A Archaebacteria
7) cFocF1 c chloroplast

20-22
Brief summarize

20-23~20-24
Two tightly coupled phenomena in OP
1. Oxidation: NADH & QH2 are oxidized by the ETC
2. Phosphorylation: The concentration gradient indirectly drive the reaction
ADP + Pi ATP + H+ (matrix site) + OH- (Intermembrane space-site)

20-25~20-32
Uncouplers () of OP
DNP (2,4-dinitrophenol): uncoupler
Valinomycin: ionophore ()
Thermogenin: natural uncoupler (uncoupling protein, UCP),
intermembrane space proton
(non-shivering thermogenesis),
UCP is found in mitochondria of brown adipose tissue (BAT), plays important
role in thermos-regulation and in overall energy balance. BAT
Sudden infant death
syndrome (SIDS, ) (owing to hypothermia)
Inhibitors of OPblock ETC (oxidation) or ATP synthase (phosphorylation)
For complex I: rotenone, amytal
For complex III: DCMU, antimycin
For complex IV: azide (N3-), cyanide (CN-), carbon monoxide (CO)
For ATPase: oligomycin (block Fo subunit)

20-33~20-40
Glucose glycolysis, pyruvate dehydrogenase complex, TCA (
)
 Net rx: Glc + 10 NAD+ + 2 FAD + 4 H2O + 4 ADP + 4 Pi 6 CO2 + 10
NADH + 4 H+ + 2 FADH2 + 4 ATP
38 ATP () Shuttle ATP
Glycerol 3-phosphate shuttle:
Mechanism: NADH (cytosol) to FADH2 (mit.)
NADH matrix shuttle
matrix CoQ ETC
Two types of Glycerol-3-P Dehydrogenase: Cytosol: NAD+-linked; Outer
face of mitochondria: FAD-linked
shuttle muscle brain BAT
BAT
Malate-Aspartate shuttle ()

38 ATP glycolysis 2 NADH

shuttle 2 ATP 36 ATP

Leakage of proton slippage of ATP synthase OP

NADH = 2.5 ATP, FADH2 = 1.5 ATP ATP yield 30

(29.85, due to newly revised ratios) ATP