ELECTRON TRANSPORT,

CHAPTER
OXIDATIVE
PHOSPHORYLATION
AND MITOCHONDRIAL
MEMBRANE TRANSPORTERS
14

Flow of electrons along the electron transport chain (ETC) is the final event in the cellular respiration which releases energy
for generation of ATP. The electron transport chains are embedded in the inner mitochondrial membrane. Each chain
consists of a series of electron carrying molecules. Electrons move from one carrier protein to another in a stepwise fashion; each
carrier being capable of accepting electrons (either with or without accompanying protons) from the preceding carrier
and donating them to the next one. At the end of ETC, the electrons reach oxygen, the final acceptor of electrons.
The electrons for the ETC are obtained during catabolism of foodstuffs, e.g. carbohydrates (particularly glucose), fatty
acids and amino acids. As these substances undergo oxidation, they lose electrons to the reduction–oxidation cofactors
NAD and FAD (less commonly to others) to generate NADH and FADH2, respectively. These reduced cofactors can give
rise to ATP by transferring their electrons to the electron transport chain. Initially these electrons are energy rich, but as
they flow down the ETC, much of their energy is lost. The lost energy is released in small packets, most of which are used
for the generation of ATP, while the rest is dissipated (i.e. entropy). Flow of electrons along the ETC (i.e. oxidation) and
generation of ATP (i.e. phosphorylation) are coupled processes, together referred to as oxidative phosphorylation. The
phosphorylation follows the oxidation; if the latter is inhibited, the former is also similarly affected. Oxidative phos-
phorylation is described in this chapter followed by a discussion about other carrier proteins that are located in the inner
mitochondrial membrane.
At the end of this chapter, the student should be able to understand:
 Electron transport chain: its localization, carrier proteins, redox couples, free energy changes, reactions and inhibitors.
 Oxidative phosphorylation, hypotheses for ATP generation during ETC and uncouplers.
 Various membrane transport systems located in the inner mitochondrial membrane.

In TCA cycle, an electron pair is removed from each of

I. Electron Transport Chain (ETC) the following substrates: isocitrate, -ketoglutarate, suc-
cinate and malate by specific dehydrogenases (Table 14.1).
A. Sources of Electrons for ETC These dehydrogenases remove a pair of hydrogen atoms
initially. Since each hydrogen atom contains an electron,
The electrons for the ETC are released during catabolic removal of two hydrogen atoms implies removal of an
pathways of biomolecules such as carbohydrates, fats, electron pair.
and amino acids (Fig. 14.1) by action of the enzymes known These electrons travel down the ETC, and combine
as dehydrogenases. These electrons are then funneled into with the last acceptor, i.e. oxygen, and two protons are
the ETC. For example, during glycolytic sequence a pair also taken up from the surrounding medium. This results
of electrons is removed from glyceraldehyde 3-phosphate. in the formation of water (Fig. 14.2).
302 Textbook of Medical Biochemistry

 Coenzymes linked with dehydrogenases: The electron

Respiratory chain or electron transport chain located in inner pairs removed from the substrate molecules do not enter
mitochondrial membrane consists of a series of electron ETC directly. They are first transferred to specialized
carriers. Electrons from the reduced coenzymes, NADH coenzymes, nicotinamide adenine dinucleotide (NAD) or
and FADH2, pass through these before they reduce oxygen. flavin adenine dinucleotide (FAD).
 Certain dehydrogenases, termed NAD-linked dehydroge-
nases, transfer the electrons to NAD-to form NADH.
Fatty acids
Amino acids
NAD+ NADH, H+
Glucose

A B C Substrate Product
(i)
Urea
 Other dehydrogenases, called FAD-linked dehydroge-
Pyruvate Carbon skeleton
nases, transfer electrons to FAD to form FADH2.
FAD FADH2
Acetyl CoA
Substrate Product
(ii)
Subsequently the reduced coenzymes, NADH and
TCA FADH2, transfer electrons to the ETC at different levels.
cycle Some NAD-linked and FAD-linked dehydrogenases pres-
ent in catabolic pathways are given in Table 14.1.

NADH/FADH2

During all major catabolic pathways, electron pair is
removed from the substrate and transferred to NAD or
FAD. This forms reduced coenzymes—NADH and FADH2,
respectively. Indeed, formation of reduced coenzymes
Electron
is common feature of all catabolic pathways.
Transport ATP
Chain DH
Substrate(s) Product(s)

½O2
2H 2H+
1O 2H+
H2O 2e−
2 2 ETC 2e−

Fig. 14.1. Electron transport and oxidative phosphorylation H2O

re-oxidize NADH and FADH2 and trap the energy released as
ATP. The NADH and FADH2 arise from all major catabolic Fig. 14.2. The electron pair, removed from the substrate travels
pathways: glycolysis, (A); -oxidation of fatty acids, (B); down the electron transport chain (ETC) to the final acceptor,
amino acid catabolism, (C); and TCA cycle. oxygen (DH  dehydrogenase enzyme).

Table 14.1. Action of FAD-linked and NAD-linked dehydrogenases

Enzyme Reaction Location
NAD-linked
Pyruvate dehydrogenase Pyruvate  Acetyl CoA M
Isocitrate dehydrogenase Isocitrate -Ketoglutarate M
Malate dehydrogenase Malate  Oxaloacetate M
-Ketoglutarate dehydrogenase -Ketoglutarate  Succinyl CoA M
Glutamate dehydrogenase Glutamate  -Ketoglutarate  Ammonia M
Glyceraldehyde-3-P dehydrogenase Glyceraldehyde 3-P1, 3-Bisphosphoglycerate C
Lactate dehydrogenase Lactate  Pyruvate C
FAD-linked
Succinate dehydrogenase Succinate  Fumarate M
Fatty acyl CoA dehydrogenase Fatty acyl CoA  Enoyl CoA M
M  mitochondrion, C  cytoplasm.
 Electron Transport, Oxidative Phosphorylation and Mitochondrial Membrane Transporters 303

B. Localization of ETC the total membrane weight. Rest of the membrane structure
is formed by lipids. This contrasts with the other cell mem-
Electron transport chains are located in the inner mito- branes which contain less amount of proteins (Chapter 7).
chondrial membrane (IMM). The catabolic pathways Mitochondrial matrix is the space enclosed by the
that yield electrons for ETC occur in the mitochondrial inner mitochondrial membrane. It contains a gel-like
matrix. Thus, there is close proximity of catabolic path- solution in which several catabolic pathways, e.g. citric acid
ways with ETC which ensures that the electrons obtained cycle, b-oxidation of fatty acids, and oxidation of amino acids,
during these pathways are rapidly transferred to ETC occur. In addition to enzymes of these pathways, the
(Fig. 14.3). Moreover, the ATP synthesizing system is also matrix contains coenzymes such as NAD, NADP and
located in IMM. This permits a rapid utilization of the FAD, and components of the phosphorylation reaction,
energy, which was released earlier during the electron e.g. ADP, ATP and phosphate ions.
flow, for the synthesis of ATP. Since all these processes

concerned with oxidative phosphorylation take place in
Proximity of catabolic pathways (occur in matrix) with the
mitochondria, it is appropriate to examine the anatomy electron transport chains and ATP synthesizing systems
of mitochondria in some detail. (located in IMM) ensures that (i) the electrons obtained
during catabolism are promptly transferred to ETC and (ii)
Biochemical Anatomy of Mitochondrion there is rapid utilization of energy for ATP synthesis.
Mitochondrion has two surrounding membranes: the
outer mitochondrial membrane and the inner mitochon-
drial membrane. The outer mitochondrial membrane C. Electron Transport: An Overview
has a relatively simple structure. It contains special pores
which make it permeable to most small molecules and ions. ETC consists of several electron carriers that are arranged
A few enzymes are also located in the membrane. The inner in a sequence shown in Figure 14.4.
mitochondrial membrane (IMM) has a more complex and
specialized structure. It contains a number of protein com- Catabolism
ponents such as enzymes, transport proteins, several sets of
electron transport chains, and the ATP synthesizing system.
NADH
A large surface area is required to accommodate all these
components. For this purpose, the membrane structure is 2e−
highly convoluted, being thrown into numerous folds,
NADH
called cristae, which serve to increase surface area of the Dehydrogenase
IMM several-fold (Fig. 14.3). Thus, the IMM of a single
mitochondrion in liver may contain over 10,000 sets of
electron transport chains and ATP synthase molecules.
Ubiquinone
In contrast to the outer mitochondrial membrane which
is freely permeable, the inner mitochondrial membrane is 2e−
selectively permeable, meaning that it is impermeable to
2 × Cytochrome b
most ions (H, K, Na) and small molecules (ADP, ATP),
and permeable only to a few: only water, carbon dioxide 2e−
and oxygen can freely move across the IMM. Movement
2 × Cytochrome c1
of other substances across the membrane can take place
only through mediation of specific transport proteins. 2e−
Presence of these protein components makes the IMM
unusually rich in proteins which constitute 75% or more of 2 × Cytochrome c

2e−
Mitochondrial Inner mitochondrial
matrix membrane 2 × Cytochrome aa3
Intermembrane
Cristae 2e−
space
Outer mitochondrial
membrane 1
2H++ 2 O2 H2O
ATP synthase molecules

Fig. 14.3. Biochemical anatomy of a mitochondrion. Fig. 14.4. An overview of electron transport chain.
304 Textbook of Medical Biochemistry

The electrons are transported from one carrier molecule to The tendency of a redox couple to donate or accept
the next either as hydride ion (:H–) which bears two electrons electrons is determined by standard redox potential (E0).
(discussed later), or as hydrogen atoms or as free electrons. Every redox couple has a characteristic value of E0.
The starting compound, that loses its electrons to the Electrons always flow from the lower to the higher redox
ETC is NADH and the final acceptor of the electrons is oxygen. potential, so that redox systems with negative E0 tend to
NADH is formed by transfer of energy-rich electron pair donate electrons to redox systems with positive E0. For
removed from a substrate during catabolism, as mentioned example:
earlier (equation i). NADH loses its electrons to the first car-
 Standard redox potential of NADH/NAD is –0.32 V.
rier of ETC (NADH dehydrogenase). Thereafter, the elec-
 That of CoQH2/CoQ is 0.10 V.
trons pass down from one carrier to another. During the
 Therefore, direction of flow of electrons is from
electron transfer, the components of ETC accept or donate
NADH/NAD to ubiquinol/ubiquinone and never
electrons, either with or without accompanying protons. A
the opposite way.
simple transport chain is depicted as below.
For the same reason, electrons flow from FMNH2/
NADH + H+ A BH2 C H2O FMN (E0 0.30 V) to CoQH2/CoQ (E0 0.05 V). In this
way, all components of the respiratory chain are arranged
to range from the most negative E0 (0.32 V of NADH/
1O NAD) to the most positive E0 (0.82 V of H2O/O2)
NADH+
AH2 B CH2
2 2
pair.
The sum of all these reactions is: Standard redox potential of components of ETC are
1 shown in Table 14.2.
NADH  H  O2  NAD  H2O (iii)
2
The electrons may travel, not only as free electron (or 
hydride) but also as hydrogen atoms. This is logical as a Redox reactions are electron-transfer reactions requiring a
hydrogen atom consists of one electron and one proton. reductant (e donor) and an oxidant (e acceptor). Together
they constitute a redox couple. The standard redox
 potential is the most useful property of redox couples.
Oxidations are broken down into several sequential reactions
with smaller free energy changes. In each step the starting car-
rier is oxidized and the next carrier is reduced. This step-wise
arrangement of electron transfer enables the body to capture
E. Free Energy Changes During
the energy for work rather than simply dissipate it as heat. Electron Flow
Just as all spontaneous transformations proceed in such
D. Redox Couples and Redox Potential a direction that there is a loss of free energy, the electron

Why do electrons flow unidirectionally from NADH (or

FADH2) to oxygen, and what is the thermodynamic origin Table 14.2. Redox potential of electron carriers
of the high energy bonds being created? The answer may be
Redox couple E0 (volts)
obtained by looking at the redox potentials of various car-

riers. As noted, a given carrier oscillates between the reduced NADH/NAD 0.32
and the oxidized states. For example, ubiquinone (CoQ) FADH2/FAD (Protein bound) 0.12
interchanges between a reduced form, i.e. ubiquinol or
FMNH2/FMN (Protein bound) 0.30
CoQH2 and an oxidized form, i.e. ubiquinone or CoQ.
The reduced and the oxidized forms of the same carrier are Ubiquinol/ubiquinone 0.10
together referred to as redox couple. Thus, the CoQ and the
0.08
2 3
Cyt b (Fe )/Cyt. b (Fe )
CoQH2 constitute redox couple, as do the oxidized cyto-
chrome b and the reduced cytochrome b. 2 3
Cyt c1 (Fe )/Cyt. c1 (Fe ) 0.23
Flow of electrons between these two redox couples is
Cyt c (Fe2)/Cyt. c (Fe3) 0.22
shown in an expanded form below:
Cyt a (Fe2)/Cyt. a (Fe3) 0.29
CoQH2 CoQ  2H  2e–
2  Cyt b (oxd)  2e– 2  Cyt b (red) 2 3
Cyt a3 (Fe )/Cyt. a3 (Fe ) 0.55
Sum: CoQH2  2  Cyt b (oxd) CoQ  H2O/O2 0.82
2  Cyt b (red)  2H
 Electron Transport, Oxidative Phosphorylation and Mitochondrial Membrane Transporters 305

transfer between the redox pairs is also accompanied by F. Components of Electron Transport
release of free energy. Amount of free energy released is
directly proportional to a difference in the standard
Chain
redox potentials of the redox pairs. A definite relation-
Five distinct types of components are present in ETC:
ship exists between the two as below:
(a) Nicotinamide nucleotides, (b) flavoproteins (con-
G0’  –nF E0 taining flavin mononucleotide or FMN; and flavin ade-
nine dinucleotide or FAD), (c) iron–sulphur centres,
G0’  standard free energy change in kcal/mole.
(d) ubiquinone, and (e) cytochromes. Their arrangement
n  number of reducing equivalents (i.e. electrons) trans-
in the respiratory chain has been depicted in Figure 14.5.
ferred (its value is 2 in the present case, as the elec-
trons are transferred in pairs).
F  Faraday’s constant (23.062 kcal V–1 mol–1). Nicotinamide Nucleotides
E0  difference between the standard redox potentials of Nicotinamide adenine dinucleotide (NAD) and nico-
the electron-donor and the electron-acceptor redox tinamide adenine dinucleotide phosphate (NADP) are
systems. the coenzymes derived from the vitamin niacin. The for-
The given relationship holds good under standard mer is more actively involved in the ETC. It is reduced to
conditions, i.e. concentration of 1.0 M of both the elec- NADH by transfer of a pair of electrons (in form of
tron donor and the electron acceptor, temperature 25ºC hydride ion) from the substrate by action of various dehy-
and pH 7.0. It can be used to calculate the standard free drogenases (see NAD-linked dehydrogenases; Table 14.1).
energy change as a pair of electrons passes along the elec- NAD  H  2e  NADH (Hydride is a proton, H,
tron transport chain, from the first redox couple (i.e. bearing two electrons)
NADH/NAD; Eº’  –0.32 V) to the last one (i.e. H2O)/
O2; Eº’  0.82 V). NADH subsequently loses its electrons to the initial
component of ETC: NADH dehydrogenase, a large protein
G0’ 2 (23.062) [0.82 – (–0.32)]  –52.6 kcal/mole.
complex embedded in IMM.
This relationship can also be used to calculate the NADP is similarly produced by action of NADP-
energy changes for individual segments of the electron dependent dehydrogenases, but it is mostly involved in
transport chain. reductive biosynthesis of biomolecules.

 Flavoproteins
Electrons flow from the redox couples with more nega-
Flavoproteins are important hydrogen carriers. Their pros-
tive redox potential to those with more positive redox
thetic groups are flavins (FMN or FAD), which are deriva-
potentials and release energy in the process.
tives of the vitamin riboflavin. These coenzymes have

V (Volt)
(−0.30 V)
−0.4
NADH E-FMN
(−0.32 V)
−0.2
Fe-S
kcal

0 Fe-S FAD Succinate (+0.03 V)

CoQ
Cyt b Fe-S Cyt c1 Cyt c (+0.25 V)
+0.2 (+0.05 V)
(+0.07 V) Cyt
+0.4 aa3

Cu
+0.6

+0.8 O2 (+0.82 V)
Direction of electron flow

Fig. 14.5. The respiratory chain of mitochondria showing direction of flow of electrons and energy relationships. The arrows ()
indicate the sites of ATP generation (E  NADH dehydrogenase, Fe-S  iron-sulphur centres, CoQ  coenzyme Q or ubiquinone,
cyt  cytochrome).
306 Textbook of Medical Biochemistry

chemically related structures. Both have a flavin mono- respectively. CoQ10 is the homologue that occurs in humans.
nucleotide (FMN) unit, which contains the reactive site. It can accept electrons from FMNH2 or from FADH2
FAD has an additional sugar group and an adenine base, and transfers them to cytochromes.
which completes its structure (Chapter 18). FAD and Generally, ubiquinone carries two hydrogen atoms, but
FMN react with two protons plus two electron, in alter- can also act in one-electron transfers by forming a free-
nating between the reduced and the oxidized state: radical intermediate, called semiquinone intermediate.
The reduced form is called ubiquinol (Fig. 14.6).
FAD  2H  2e  FADH2
FMN is prosthetic group of NADH dehydrogenase. It is Cytochromes
bound firmly to the enzyme protein and does not func- Cytochromes are the final class of components that par-
tion as a diffusible co-substrate. FAD, on the other hand, ticipate in electron transport. They are integral mem-
is linked to another flavoprotein, succinate dehydrogenase, brane proteins (with the exception of cytochrome c), and
and functions as a diffusible co-substrate. are named so because they are coloured cellular compo-
nents (cyto cell, chrome colour). They are conjugated pro-
Iron-sulphur Centres teins, containing an iron-porphyrin as a prosthetic group. In
They are also known as non-haem iron proteins or the most cases, this iron-porphyrin is the haem group, also
iron-sulphur clusters. Several types of iron sulphur (FeS) found in oxygen transport proteins, e.g. haemoglobin and
centres exist, but in each case the iron atoms are coordi- myoglobin. In contrast to these proteins, however, in cyto-
nated to inorganic sulphur atoms (and the sulphur of chromes the haem iron cannot bind oxygen (or carbon
cysteine side chains in proteins). Within an FeS cluster, monoxide or any other potential ligand), but rather acts as
an electron is carried by the iron atom, which, upon accept- a reversible electron carrier. Iron in cytochromes (but not
ing the electron, changes from the Fe3 (ferric) state to in haemoglobin or myoglobin) undergoes physiological
the Fe2 (ferrous) state. As the electron is passed to another oxidation–reduction between the ferrous (2) and ferric (3)
electron carrier, the iron atom of the FeS cluster changes states, as electrons are shuttled from one protein to another.
back again to the ferric state. Why cytochromes can carry only electrons, but can-
As explained in the next section, one group of iron- not bind oxygen? This is shown diagrammatically in
sulphur protein participates in the transfer of electrons Figure 14.7. In cytochromes, the iron group of the haem
from FMN to ubiquinone (CoQ), and the other from is anchored on both sides, on one side by histidine and
cytochrome b to cytochrome c1 (Fig. 14.5). on the other by methionine. In haemoglobin, it is only
anchored on one side by histidine, enabling oxygen to
Ubiquinone interact with the free side of Fe.
Ubiquinone, also known as coenzyme Q, is a mobile dif- The main varieties of cytochromes in mitochondria
fusible hydrogen carrier, which can move from donor to are the a, b, and c types of cytochromes, the classification
acceptor molecules during electron transport. It is a ben- being based on the respective absorption spectrum and
zoquinone with a hydrocarbon tail of (usually) 10 isoprene the type of porphyrin structure present. The cytochrome
(branched 5-carbon) units (Fig. 14.6), which makes it of b type contains haem, whereas the others have por-
strongly hydrophobic and confines it to the lipid bilayer phyrins with different side chains. Cytochromes a and a3
of the inner mitochondrial membrane. It was also named contain “haem a” rather than “ordinary haem”. In this the
ubiquinone because of its ubiquitous or widespread dis- porphyrin ring of haem is modified: (i) a methyl group
tribution. of haem is oxidized to a formyl group, and (ii) a hydro-
The names coenzyme Q and ubiquinone and the phobic isoprenoid chain is attached to one of the vinyl
abbreviations CoQ and UQ are used interchangeably. A groups. (See chapter 16 for haem structure.)
subscript(n) indicates the number of isoprene units; for The cytochrome families are subclassified in terms
example, CoQ6 or CoQ10 contain 6 or 10 isoprene units of the historical order of their discovery (b1, b2, b3 ...) or

O OH
H3CO CH3 H3CO CH3
2e−, 2H+
CH3 CH3

H3CO (CH2 CH C CH2)n H H3CO (CH2 CH C CH2)n H

O OH
2e−, 2H+
Ubiquinone–oxidized form Ubiquinol–reduced form (QH2)

Fig. 14.6. Structure of ubiquinone, also called coenzyme Q (CoQ) or UQ. The isoprene units are shown in colour.
 Electron Transport, Oxidative Phosphorylation and Mitochondrial Membrane Transporters 307

Cytochrome c, e− carrier Haemoglobin, O2 carrier NADH

O2
NADH ubiquinone Rotenone, BAL,
His N S Met His N Complex I
reductase (Site 1) barbitruates
Complex II
Ubiquinone Succinate
Haem group Haem group
Succinate-ubiquinone
Fig. 14.7. Haem is prosthetic group in both cytochrome C and reductase
haemoglobin, but performs different roles. It serves as oxygen
Ubiquinol-cytochrome c Antimycin A
carrier in haemoglobin and electron carrier in cytochrome C Complex III
reductase (Site 2)
(His  histidine; Met  methionine).
Cytochrome c
the wavelength (nm) of a characteristic spectral absorp-
tion peak (b562, b566). The electrons are transported from
coenzyme Q to cytochromes in the order: b, c1, c, a and Cytochrome oxidase
Cyanide, carbon
a3. The last two can directly react with molecular oxygen. Complex IV monoxide,
(Site 3)
hydrogen sulphide
Besides haem iron, the aa3 are associated with copper,
which acts as an electron carrier by a valence change O2
between the cuprous (Cu) and cupric (Cu2). As
described later, it (aa3) participates in the last reaction Fig. 14.8. The structured complex of functionally related elec-
and transfer of electrons to molecular oxygen. tron carriers. Inhibitors of ETC ( ) are also shown. Sites 1, 2 and
3 refer to the sites of proton pumping (“phosphorylation sites”).

Respiratory chain contains flavoproteins, iron-sulphur As the electron pair flows from NADH to complex I, it
proteins, ubiquinone, cytochromes and protein-bound is accepted together with a hydrogen ions, H, such that
copper. Each component participates in electron trans- two electrons and two H are accepted in total. As a result
fers by changing oxidation state between the oxidized FMN is converted into FMNH2. The electrons are then
form and the reduced form. transferred within the complex I to iron-sulphur clusters,
and then passed onto ubiquinone, which is thereby con-
verted to ubiquinol (Fig. 14.6). Ubiquinone has a long,
G. Pathway of Mitochondrial Electron flexible, lipid soluble arm, and so it can readily move through
Transport the inner membrane to transfer the electrons to the next
enzyme in the sequence, i.e. the Complex III (the
Only four components of the respiratory chain are freely Complex II will be dealt with later).
diffusible (NADH, ubiquinone, cytochrome c and oxy-

gen), and the rest are organized as constituents of large The, complex I contains various electron carriers that
protein complexes (Fig. 14.8). There are four complexes work sequentially to carry electrons down the chain to
(I to IV) embedded in the inner mitochondrial mem- ubiquinone, which then transfers them to the complex III.
brane. Exact detail of the complexes is not known. What
is known is: (i) that a complex contains many polypeptide or
protein subunits and several iron centres, (ii) that these com- Complex III
ponents can be readily reduced or oxidized, and (iii) that Complex III (ubiquinol-cytochrome c reductase complex)
they transfer electrons as they flip between the reduced once again uses iron atoms to shuffle electrons within its
state and the oxidized state. structure. It contains cytochrome b, an iron-sulphur centre
(called Rieske’s protein), and cytochrome C1.
Complex I QH2  Cyt b  FeS  Cyt c1  Cyt c
Oxidation of NADH begins with Complex I (also called
Complex III
NADH-ubiquinone reductase or NADH dehydrogenase com-
Figure 14.9 elaborately depicts the electrons passing
plex), which comprises 28–41 protein subunits (depend-
from ubiquinol, through the cytochrome b, FeS and
ing upon the species), FMN as a prosthetic group and
cytochrome c1 components of this complex, and the
about 7-iron-sulphur (FeS) centres. It transfers electrons
accompanying ferric-ferrous interconversions.
from NADH to ubiquinone via FMN and FeS centre:
Note that cytochrome c is a peripheral membrane protein
Substrate  NADH FMNH2  FeS  Ubiquinone bound to the outer membrane surface, which transfers
Complex I its electrons to Complex IV.
308 Textbook of Medical Biochemistry

Ubiquinol 2Fe3+ 2Fe2+ 2Fe3+ 2Fe2+

2.Cyt b 2.Fe-S 2.Cyt c1 2.Cyt c

Ubiquinone 2Fe2+ 2Fe3+ 2Fe2+ 2Fe3+

2H+
Complex III

Fig. 14.9. Flow of electrons from ubiquinol to cytochrome c via the large, multiprotein complex III, consisting of cytochrome b, an
iron sulphur protein and cytochrome c1.

 Succinate + FAD  Fumarate + FADH2

Large protein complexes are arranged asymmetrically in
SDH contains iron-sulphur centres, in addition to the
the membrane. Ubiquinone (Q) is present at junction of com-
plex I and III and cytochrome c lies at junction of III and covalently bound FAD. Electrons from FADH2 are trans-
IV. Q is in the lipid bilayer and cytochrome c is peripheral ferred to ubiquinone (via cytochrome b560 and FeS) and
membrane protein bound to outer surface of IMM. reduce it to ubiquinol. The latter then proceeds to reduce
Complex III.

Complex IV Succinate  FADH2  Cyt b560  FeS  CoQ

Complex IV (cytochrome oxidase complex) contains 13 Complex II
different polypeptides, two haem groups, and two cop-
per ions. The complex IV transfers electrons to O2, the Evidently, succinate dehydrogenase (and other mito-
final acceptor, to form water: chondrial flavoproteins) bypasses the Complex I and
O2 pass their electrons directly to ubiquinone. This results in
Cyt c  Cyt aa3  H2O generation of only two ATPs when FADH2 is used as
Complex IV the substrate; three are generated if NADH is used as
the substrate. The reason for this is that one ATP each is
The complex contains two cytochromes (a and a3), generated at Complex I, III, and IV.
which are associated with haem iron and copper.
Cytochrome a is paired with a copper atom, CuA and 
cytochrome a3 is paired with a different copper atom, The respiratory chain contains four large multi-protein
CuB. During electron transfer, the iron atoms of the haem complexes. Coenzyme Q shuttles electrons from com-
cycle between the ferric and ferrous states while the cop- plexes I and II to Complex III; and cytochrome c shuttles
per atoms cycle between cuprous and cupric. The cyto- electrons from Complex III to Complex IV, which then
chrome oxidase reaction is complex: it transfers four transfers them to oxygen, the final acceptor.
electrons to molecular oxygen to form two molecules of
water. Oxygen is tightly bound between haem a3 and
copper during its reduction, and it is released only after H. Inhibitors of ETC
its complete reduction to water.
4e  O2  4H  2H2O A number of site-specific inhibitors are known to block
transport of electrons along the respiratory chain (Fig. 14.8).
Because of high affinity of cytochrome oxidase for
This action inhibits oxygen consumption and second-
molecular oxygen, the oxidative phosphorylation is near
arily halts synthesis of ATP from ADP and Pi. Some com-
maximum even at low oxygen pressure.
mon inhibitors and their sites of action are as follows:
Complex II Rotenone: It blocks transfer of electrons from NADH to
Complex II (succinate-ubiquinone reductase or succinate ubiquinone. It is a plant product that is extremely toxic
dehydrogenase) contains four polypeptide chains; the and is used by the American Indians as a fish poison.
first two constitute the bonafide succinate dehydrogenase However, it is not very toxic to humans because of its
(SDH), a Krebs cycle enzyme, which catalyzes the follow- poor intestinal absorption. Humans, therefore, can safely
ing reaction: eat the poisoned fish.
 Electron Transport, Oxidative Phosphorylation and Mitochondrial Membrane Transporters 309

Barbiturates: Besides rotenone, some barbiturates (e.g. Chemiosmotic Hypothesis

amytal) inhibit electron flow through Complex I. Proposed by the British biochemist Peter Mitchell (1961),
this hypothesis is widely accepted. According to the hypoth-
Antibiotics: An antibiotic, British antilewisite (BAL) also
esis, oxidative phosphorylation occurs in two steps:
acts at Complex I; and another antibiotic, antimycin A,
inhibits Complex III.  Generation of electrochemical gradient across the
IMM (Step I).
Cyanide, carbon monoxide azide, and hydrogen sulphide:
 Utilization of this gradient to fuel ATP synthesis (Step II).
These inhibitors act at complex IV. They bind to the iron
protoporphyrin in cytochrome aa3 whose sixth coordina-
Step I
tion position is not occupied by an amino acid side chain
The electrochemical gradient is built by pumping of pro-
but is reserved for oxygen. Cyanide and azide bind to the
tons out of the mitochondrial matrix. The proton pumping
ferric form of the iron; carbon monoxide binds to the fer-
is fueled by the energy released by exergonic redox reactions of
rous form.
ETC (Fig. 14.10). This creates a proton gradient of approx-
Hydrogen sulphide also is a potent inhibitor. Carbon
imately 1.4 pH units across the IMM (inside alkaline). It also
monoxide not only inhibits electron flow, but also has
builds a membrane potential of 100–200 mV1 (inside
high affinity for haemoglobin, which adds to its clini-
negative). The concentration and electrical potential add
cally important toxicity.
up to a steep electrochemical gradient for protons.

Electron transport inhibitors block oxidation–reduction Step II
reactions at specific locations along the ETC. The electrochemical gradient is a potential source of
energy (4–6 kcal/mol) for generation of ATP. The enzyme
Inhibition of the electron flow by these agents creates responsible for utilizing this gradient for ATP synthesis is
crossover points. The electron carriers present before the known as ATP synthase. It is also known as the ATPase,
block become reduced and those after the block become because the isolated enzyme is capable of catalyzing
oxidized. Since absorption spectra of the oxidized and hydrolysis of ATP to ADP and inorganic phosphate.
the reduced forms of electron carriers differ from each ATPase activity
other, these changes can be detected by spectrophotometer. ATP ATP-Pi
Figure 14.11 gives a magnified view of the ATP syn-
thase, which consists of two components: F0 and F1. The
II. Oxidative Phosphorylation F0 is embedded in the IMM, whereas the F1 protrudes
into the mitochondrial matrix (Fig. 14.11). F1 unit (F1
There are several hypotheses proposed to explain how stands for “coupling factor 1”) is a protein complex of
energy released during flow of electrons along the respi- subunit structure 33 and a molecular weight of
ratory chain is used for ATP generation. Intermembranous
space

A. Hypotheses for ATP Generation

H+ I H+ Matrix
Q
Chemical Coupling Hypothesis
H+ III H+
This hypothesis, proposed by Edward Stater (1953), pos- H+ ATP
C
tulates that the energy released from ETC causes forma- H+ IV H+
tion of high energy covalent intermediates. These ADP Fo
intermediates are subsequently cleaved to release their
F1
energy content, which is used for the synthesis of ATP.

Conformational Coupling Hypothesis

The conformational coupling hypothesis put forth by
Paul Boyer (1964), proposes that the energy of electron Fig. 14.10. Protons are translocated from the mitochondrial
transport is used for altering conformation of certain matrix to intermembranous space, fueled by the redox reac-
proteins that are located in the IMM. The proteins with tions of the respiratory chain in step I. The three complexes
altered conformation have high energy content, which is shown (I, II, IV) are actually sites of proton pumping (Q  ubi-
subsequently used for ATP generation. quinone, C  cytochrome c).
310 Textbook of Medical Biochemistry

IMM (150 V) 1. An intact membrane is required for ATP synthesis by

Intermembranous + − Matrix oxidative phosphorylation.
space
+ − 2. Respiring mitochondria generates a proton gradient.
+ −
3. Certain compounds (such as DNP) stop ATP synthe-
sis without inhibiting electron transport from NADH,
or succinate, to oxygen. These compounds prevent
ADP + Pi
building up of the proton gradient, which stops/
halts synthesis of ATP. Thus, they act as uncouplers of
H+ oxidative phosphorylation (discussed later).
ATP + H2O 
The exergonic reactions of the electron transport chain
Proton cause translocation of protons out of the mitochondrial
channel F1 ATPase
(F1 unit) matrix to form an electrochemical gradient whose free
(F0 unit)
energy drives ATP synthesis.

Fig. 14.11. Protons are allowed to flow back into the mito- B. ATP Production and P : O Ratio
chondrion through a specific proton channel in F0. This proton
channel is coupled to an ATP synthesizing enzyme (the F1 unit).
It is well established experimentally that a pair of elec-
380 dDa. It can be visualized in the electron microscope trons originating in NADH generates three molecules of
as small buttons on the inner surface of the inner mito- ATP via the oxidative phosphorylation pathway, and that
chondrial membrane. The F1 unit is attached to the F0 from FADH2 generates two. It is said that the P : O ratio
unit, an integral membrane protein having a pore called is three for electrons from NADH and two for those
proton channel. from FADH2. P referring to a high-energy phosphate
The proton channel is crucial for ATP generation. This bond being synthesized and O referring to an atom of
is because the IMM is impermeable to protons, and so oxygen being reduced. Thus P : O ratio is defined as the
the extruded protons in the intermembrane space can re- number of ATPs generated for each oxygen consumed. Refer
enter the mitochondrial matrix through this proton to Box 14.1 for more information on relationship
channel. As these protons move inwards, the energy between ATP generation and oxygen consumption.
inherent in the electrochemical gradient is liberated as
concentrated packets. These energy packets are readily Sites of ATP Production
used by the F1-unit to fuel ATP synthesis. There are three energy conserving segments that release
sufficient amount of energy to result in synthesis of an
 ATP molecule each. At each of these segments, called
The two major components of ATP synthase are F1 (ATP phosphorylation sites (indicated by arrows in Fig. 14.5),
synthesizing) attached to F0, which is proton channel quantum of energy released exceeds 10 kcal/mole.
spanning the IMM. In mitochondria this complex uses the Because ATP is synthesized only when protons flow, this
energy released by electron transport to drive ATP synthe- results in halting ATP synthesis.
sis, but in isolation it hydrolyzes ATP (ATPase activity).
1. The Site I is between NADH and ubiquinone (specifi-
Action of the ATP synthase is inhibited by an antibi- cally during electron flow through the FeS complex).
otic, oligomycin. The latter binds with F0 portion of this 2. The Site II is between ubiquinol and cytochrome c
enzyme and closes the proton channel. As a result, re- (specifically during electron flow from cytochrome b
entry of protons is blocked and the energy inherent in to cytochrome c1).
the electrochemical gradient cannot be used for ATP gen- 3. The Site III is between cytochrome c and oxygen (i.e.
eration. As more and more protons are actively pumped the cytochrome oxidation reaction).
out, the proton concentration in the intermembrane
space rises. It becomes difficult to pump protons against 
the rising gradient and finally electron transport also
P : O ratio is defined as the number of ATPs synthesized per
oxygen reduced, is three with NADH and two with FADH2.
stops. At this stage, the ATP synthase is turned off.
Success of chemiosmotic hypothesis: The chemiosmotic The three sites correspond to Complexes I, III and IV
hypothesis has been widely accepted because it can pro- of the ETC. Since Complex I is bypassed with FADH2, only
vide explanations for the following phenomena: two ATPs are generated (P : O ratio-2).
 Electron Transport, Oxidative Phosphorylation and Mitochondrial Membrane Transporters 311

BOX 14.1
Acceptor Control of Respiration
Though ATP generation follows electron transport, the rate of electron flow is dependent on ATP generation itself. Under nor-
mal circumstances, as the electron transport proceeds, ADP and phosphate are removed from cytosol and intramitochondrial
concentration of ATP builds up concomitantly. When the ADP is depleted from cytosol, the rate of electron flow (measured by
oxygen consumption) falls. This is because of the limiting action of ADP on rate of oxygen consumption. This is known as “rest-
ing respiration”. Conversely, when ADP concentration rises in cytosol (for example, after some energy requiring process which
degrades ATP to ADP), phosphorylation of ADP increases. Rate of oxygen consumption increases in this instance because of ADP-
induced oxygen consumption. This is followed by (coupled) phosphorylation of ADP to ATP. Such dependence of the rate of
oxygen consumption on the intracellular concentration of ADP (the phosphate acceptor), is called acceptor control of respiration.
Active
respiration

Oxygen Resting
consumption respiration

Time

In the case of poisoning by certain uncouplers (e.g. pentachlorophenol), ATP generation is halted and concentration of ADP
in cytosol rises. Rise in ADP concentration causes increase in oxygen consumption, which implies speeding up of the electron
transport activity.

C. Energetics OH
NO2
If one starts from NADH, with its redox potential of
–0.32 V, and ends with oxygen, redox potential 0.82 V,
then one spans a total of 1.14 V (Fig. 14.5). As discussed NO2
earlier, this comes to a theoretical availability of about Fig. 14.12. Structure of 2,4 dinitrophenol, a highly diffusible
52.6 kcal/mole (equation iv). Because three ATP mole- chemical that uncouples oxidative phosphorylation by trans-
cules are synthesized for each pair of electrons channeled porting protons across the IMM.
to oxygen, oxidative phosphorylation recovers 21.9 kcal
preventing the building up of proton gradient. Though an
(each ATP  7.3 kcal) of the 52.6 kcal available as ATP, for
uncoupler impedes ATP generation, it has no effect on elec-
an approximate efficiency of 48%.
tron transport. The latter process continues as before, or is
21.9 rather enhanced due to acceptor control of respiration, dis-
Efficiency   100  48%
52.6 cussed later. The energy released by electron flow is mostly
The remaining 52% of energy is lost as heat. dissipated (i.e. entropy). Some portion of it generates heat:
Uncouplers: Being linked through a proton gradient, the this explains thermogenic effect of some uncouplers (Case 14.1).
oxidation and the phosphorylation are said to be coupled The other uncouplers include pentachlorophenol,
processes. They can be uncoupled from each other by dinitrocresol, and trifluorocarbonylcyanide. The latter
certain compounds, called the uncouplers. Primary action dissipates the proton gradient 100 times faster than DNP.
of these compounds is to increase permeability of the 
inner mitochondrial membrane to protons. As a result, The uncouplers allow electrons transport to proceed
relatively free movement of protons across the IMM without ATP synthesis. They uncouple by carrying H ions
occurs, which prevents building of the electrochemical across the IMM and hence dissipate the proton (electro-
gradient. Since this gradient is essential for ATP genera- chemical) gradient.
tion, failure to build it stops ATP generation.
An example of uncoupler is 2,4-dinitrophenol (Fig. Physiological uncouplers and ionophores: Some endoge-
14.12). The compound can freely move across the IMM due nous compounds, including free fatty acids, bilirubin
to its lipophilic nature. It can carry a proton along with itself. and possibly thyroxine, also can act as uncouplers at
As a result, free movement of protons across the IMM occurs, concentrations well above the physiological range. They
312 Textbook of Medical Biochemistry

are called physiological uncouplers. In case of bilirubin, Cytosol Mitochondrial matrix

which deposits in the basal ganglia of infants with severe ADP
hyperbilirubinaemia (see Chapter 16), a role of the un- Adenine nucleotide
coupling effect in the resulting brain damage is possible. ATP translocase
Valinomycin is a transport antibiotic or an ionophore H2PO4−
that makes the inner mitochondrial membrane permeable Phosphate tanslocase
to potassium (the term ionophore refers to compounds that OH− H+
promote the transport of ions across biological mem- Pyruvate
branes; Chapter 7). It also acts as an uncoupler. Monocarboxylate
OH− carrier
This is because the translocation of potassium ions
dissipates the membrane potential, which is an essential Malate
Dicarboxylate
component of the proton-motive force.
α-Ketoglutarate carrier
Citrate
Significance of Uncoupling Tricarboxylate
Uncoupling under natural conditions plays an important Citrate carrier
role in hibernating animals, and in animals of polar regions, Aspartate
for maintaining their body temperature. These animals Amino acid
contain brown adipose tissue which is specialized to carry Glutamate antiport
out oxidation uncoupled from phosphorylation. When fats
are oxidized in the brown adipose tissue, the energy liber- Fig. 14.13. Energy of electrochemical gradient being used
ated is not trapped as ATP but rather released as heat. for ATP generation.
Presence of active brown adipose in some individuals
is believed to burn excess calories, thereby preventing them
from gaining weight even after indulging in overeating. B. Phosphate Translocase System
The phosphate translocase system is functionally related to
the adenine nucleotide translocase. It cotransports a phos-
III. Mitochondrial Membrane phate ion along with H into the mitochondrial matrix
Transporters (Fig. 14.13). Phosphate is then used for the generation of
ATP from ADP. Thus, a combined action of these two
The carrier proteins in IMM, also known as mitochondrial translocase systems permits entry of phosphate and ADP
membrane transporters, are highly specific in their action. into the mitochondrial matrix, where they combine to
They permit movements of several molecules that are form ATP.
important for mitochondrial function, such as ATP, H,
pyruvate, malate, citrate, etc. A few transport proteins
(Fig. 14.13) and their actions are described here.
C. Monocarboxylate Carrier
This carrier transports pyruvate produced in cytosol,
A. Adenine Nucleotide Translocase (ANT) mainly through the glycolytic sequence, into mitochon-
dria where it is further metabolized.
The adenine nucleotide translocase permits inward
movement of ADP into the mitochondrial matrix, and a
simultaneous outward movement of ATP into the
D. Dicarboxylate Carrier
intermembranous space. These movements are impor-
It moves malate from site of its production (i.e. the mito-
tant because ADP (and phosphate) have to enter the
chondrial matrix) into cytosol.
mitochondrion as substrates for oxidative phosphoryla-
tion, and ATP has to pass from the mitochondrion to the
cytosol, where it is used for energy-dependent processes. E. Tricarboxylate Carrier
Structurally, ANT is an integral membrane protein that
extends across the IMM. It transports citrate, the first intermediate of TCA cycle,
Atractyloside is toxic glycoside which specifically into cytosol. This carrier is very specific: it does not trans-
inhibits the ANT system. Inhibition of ANT hampers the port even a closely related molecule, i.e. isocitrate.
transport of ATP, which leads to serious consequences Combined action of the dicarboxylate and tricarbox-
since ATP is required to drive a number of cellular activities. ylate carriers play vital role in lipogenesis (Chapter 11).
 Electron Transport, Oxidative Phosphorylation and Mitochondrial Membrane Transporters 313

2. Malate is then transported across the IMM by the


Specific transporters in IMM mediate the transmem- dicarboxylate carrier.
brane movements of ADP, ATP, Pi and several metabolites. 3. Within the mitochondrial matrix, the reducing equiv-
alents are transferred from malate to NAD to form
NADH. The enzyme that catalyzes this reaction is
F. Shuttle Systems mitochondrial malate dehydrogenases.
Thus, by the concerted action of the dicarboxylate car-
Certain shuttle systems are known to operate in mitochon-
rier and two enzymes (i.e. cytosolic malate dehydrogenase
dria that transport reducing equivalents from cytosol into
and mitochondrial malate dehydrogenase), NADH reaches
mitochondria. The reducing equivalents (in the form of
the mitochondrial matrix (Fig. 14.14a). The mitochon-
NADH) are generated in the cytosol (during glycolysis).
drial NADH now contains the reducing equivalents that
They have to be transported to mitochondrial matrix for oxi-
were originally present in the cytosolic NADH.
dation and generation of ATP. However, the IMM is imper-
Rest of the shuttle systems is concerned with transport
meable to NADH. Therefore, specific shuttle systems,
of the oxaloacetate back into the cytosol. It requires par-
namely malate aspartate shuttle and glycerol phos-
ticipation of amino acid antiport system which exchanges
phate shuttle, accomplish their transport (Fig. 14.14).
aspartate for glutamate across the IMM.
 Glycerol phosphate shuttle: In skeletal muscle and
Shuttle systems move the NADH produced in the cytosol brain, another type of shuttle, called glycerol phosphate
into the mitochondrion. shuttle, is present. It transfers the hydrogen first to
dihydroxyacetone phosphate forming glycerol phos-
Malate aspartate shuttle: It operates in liver and heart phate (in cytosol), and then to the FAD prosthetic group
muscles in the following steps: of the mitochondrial glycerol phosphate dehydrogenase (Fig.
1. The reducing equivalents are transferred from NADH 14.14b). The latter is an integral protein of the inner
to oxaloacetate to form malate. The reaction is cata- mitochondrial membrane, which regenerates its FAD by
lyzed by the cytosolic enzyme malate dehydrogenase. direct transfer of electrons to the respiratory chain. Only

(a) NAD+ Cytosol IMM Mitochondrial matrix

NADH 1 NAD+ NADH
3
Malate Malate
2
Oxaloacetate α-Ketoglutarate α-Ketoglutarate E
Oxalo- T
acetate C
4
6

Glutamate Glutamate
5
Aspartate Aspartate

1. Cytosolic malate dehydrogenase

2. Dicarboxylate carrier
3. Mitochondrial malate dehydrogenase
4. Transaminase
5. Amino acid antiport
6. Transaminase

(b) Cytosol Mitochondrial matrix

NAD+ Glycerol Glycerol

1 2 FAD
phosphate phosphate

Dihydroxy-
NADH Dihydroxyacetone
acetone FADH2
phosphate E
phosphate
IMM T
C
1. Cytosolic glycerol phosphate dehydrogenase
2. Mitochondrial glycerol phosphate dehydrogenase

Fig. 14.14. Transfer of reducing equivalents from cytosol into mitochondrial matrix. (a) Malate aspartate shuttle, (b) Glycerol
phosphate shuttle (IMM  mitochondrial matrix).
314 Textbook of Medical Biochemistry

two ATPs are therefore produced for each pair of reduc- fertilization. The polypeptide chains encoded by mtDNA
ing equivalents transferred. include seven subunits of NADH-Q reductase, one subunit
of QH2-cytochrome c reductase, three subunits of cytochrome
 oxidase and two subunits of ATP synthase. All other mito-
Malate aspartate shuttle (generates three ATPs) is more chondrial proteins are encoded by nuclear genes and syn-
important in liver and heart; and the glycerol phosphate thesized in cytoplasmic ribosomes.
shuttle (generates two ATPs) is more important in muscle Mitochondrial DNA is 10 times more susceptible to
and brain.
mutations than nuclear DNA. Some of these mutations
result in absence of specific polypeptide chains. Faulty
operation of the electron transport pathway results and
IV. Enzymes Participating in the ATP production is decreased. These disorders are
known as mitochondrial myopathies and they most com-
Biological Oxidation monly affect the tissues with high rate of oxidative phos-
phorylation. The mitochondrial myopathies are tissue
The enzymes involved in biological oxidation catalyze specific; some affect the heart, others the skeletal muscles.
oxidation-reduction reactions, hence they belong to the Many are accompanied by lactic acidosis, because the
class I, i.e. oxidoreductases (Chapter 6). They are subclassified inability to reduce NADH normally results in channeling
as: (a) dehydrogenases, (b) oxidases, and (c) hydroperoxidases. of pyruvate toward lactic acid production (Chapter 9).
Leber’s hereditary optic neuropathy (LHON) is a mitochon-
A. Dehydrogenases drial myopathy, characterized by sudden onset of blindness
in adults, associated with degeneration of the optic nerve.
They are most common of all oxidoreductases. They cata- 
lyze transfer of hydrogen between a substrate and a coen- Mutations in mitochondrial DNA cause inherited disorders
zyme, most commonly NAD, FAD, NADP or FMN. of oxidative phosphorylation, e.g. Leber’s hereditary optic
These enzymes are named according to the substrate neuropathy.
from which they remove the hydrogen (Table 14.1).

B. Oxidases Exercises

They catalyze transfer of hydrogen removed from a sub- Essay type questions
strate to oxygen. Mostly this results in formation of 1. What is redox potential and what is its significance?
water. Cytochrome oxidase, the terminal component of Describe various components of electron transport
ETC, is an example. chain and discuss the oxidation of NADH and FADH2.
2. What do you understand by the term P : O ratio? Indicate
diagrammatically the sites of ATP generation in the
C. Hydroperoxidases mitochondrial respiratory chain and their inhibitors.
3. What is oxidative phosphorylation? Discuss in detail
They use hydrogen peroxide or an inorganic peroxide as the chemiosmotic hypothesis of ATP synthesis and
a substrate. They are needed for degradation of hydrogen explain how the process of oxidative phosphoryla-
peroxide that may be produced in the reactions catalyzed tion is arrested by various inhibitors.
by the dehydrogenases. Technically, catalase is also a per- 4. What are uncouplers of biological oxidation? State
oxidase: it degrades hydrogen peroxide to molecular oxy- their mechanism of action.
gen and water.
Write short notes on
H2O2  H2O  O2 1. Uncouplers
2. P : O ratio
3. Redox potential
V. Mitochondrial Myopathies 4. ATP synthesis
5. Physiological uncouplers
Mitochondrial DNA consists of a circular DNA molecule 6. Brown adipose
of 16,569 base pairs which encodes 13 polypeptides, 22 7. Ubiquinone
tRNAs, and the RNAs of mitochondrial ribosomes (12S 8. Cytochrome aa3
and 16S). The mtDNA is maternally inherited since mito- 9. Chemiosmotic theory
chondria from sperm does not enter the ovum during 10. Malate aspartate shuttle
 Electron Transport, Oxidative Phosphorylation and Mitochondrial Membrane Transporters 315

CLINICAL CASE
CASE 14.1 Labourer exposed to a toxic wood preservative
A labourer felt dizzy and feverish while spraying penta- Note: Rate of electron flow is reflected by O2 consump-
chlorophenol, a popular wood preservative that kills ter- tion rate.
mites, fungi and bacteria. While being taken to the hospital
he lost consciousness on the way. He was admitted in the (a) (b)
hospital emergency. On examination, his body tempera-
ture was 39.9ºC, and respiratory rate was 60 per minute.

consumption
consumption
He died in a comatose state, after two days. Pentachlorophenol DNP

Oxygen
Oxygen
On autopsy, the subcutaneous and the omental fats
were markedly depleted. There was pulmonary conges-
tion, and the marrow examination showed evidence of
enhanced erythropoiesis.
Exposure to the wood preservative was thought to be Time Time
responsible for the patient’s condition. Effect of the chemical (c)
was investigated by performing the following tests on a prep- DNP
aration of rat liver mitochondria. The mitochondria were

consumption
suspended in a suitable buffer, to which ADP and succinate Pentachlorophenol

Oxygen
were added. The preparation was incubated in the oxygen
electrode chamber and the rate of oxygen consumption was
measured. Addition of pentachlorophenol to this prepara-
tion caused a marked increase in the oxygen consumption
(Experiment a). A similar increase was observed following Time
addition of 2, 4 dinitrophenol (Experiment b) to this prepara-
tion. However, when the dinitrophenol (DNP) was added fol- Q.1. State the possible mechanism of action of the uncou-
lowing the addition of pentachlorophenol, no further increase pling effect of the pentachlorophenol.
in oxygen consumption resulted (Experiment c). Q.2. Interpret the test results and elucidate the action of
Experiment a–c. Effect of the pentachlorophenol and pentachlorophenol on mitochondrial respiration.
dinitrophenol (DNP) on oxygen consumption by rat liver Q.3. Explain the clinical and the autopsy findings of this
mitochondria. patient in molecular terms.