electron transport chain

1. Electron Transport Chain and
Oxidative phosphorylation
1

2. Introduction
⚫Respiration is the oxidative breakdown of organic compound (lipid,
proteins, carbohydrates) to release energy.
⚫Main aim of this various metabolic reactions is to production of
ATP.
⚫An electron transport chain(ETC)is a series of complexes that
transfer electron from electron donors to electron acceptor via
redox(both reduction and oxidation occurring simultaneously)
reaction, and couples this transfer of protons(H+ ions) across a
membrane.
⚫Oxidative phosphorylation is the process of coupling the
electron
transport chain withATP synthase(complex-v).
⚫ETC occur in plasma membrane of prokaryotes, and inner
mitochondrial membrane of eukaryotes.

3.  Mitochondria contain
two membranes, an
outer membrane
permeable to small
molecules and ions and
an impermeable inner
membrane.
 The inner membrane
contains components of
the respiratory chain for
transfer of electrons to
O2 and also contains
ATP synthase, an
enzyme that
synthesizesA
TP
.

4.  Electron Carrier in the Respiratory
Chain
⚫NADH- Nicotidamide adenine dinucleotide soluble molecule
used by dehydrogenase.
⚫Flavoproteins- contain FAD or FMN which can be reduce to
FADH2 and FMNH2.
⚫Ubiquinone- also known as coenzymeQ, lipid-soluble
metabolite that function in the ETC.
⚫Cytochromes -protein that contain a heme prosthetic group.
⚫Iron-sulfur protein- role in oxidation-reduction reactions of
electron transport.

5. Enzyme complex of ETC
 The electron transport system consists of five large protein
complex:
 1.Complex 1; NADH-ubiquinone oxidoreductase (NADH
dehydrogenase)
 2.Complex 2; succinate dehydrogenase (citrate cycle enzyme)
 3.Complex 3;ubiquinone – cytochrome c oxidoreductase
 4.complex 4;cytochrome c oxidase
 5.ATP synthase complex;it consisting of a F0 and a head F1.

7.  Ubiquinone

8. Cytochromes
 Protein with characteristic
strong absorption of visible light
due to their iron containing
Hemeprosthetic group.
The heme cofactor of a and b
cytochrome are tightly but not
covalently bound to their
associated protein whereas the
heme of c type cytochrome are
covalently attached to Cys-
residues.
Cyt-c associates through
electrostatic interaction with the
outer surface of the inner
membrane.

9. Iron-sulfur Proteins
Iron-sulfur prosthetic groups
consist of non-heme iron complex
d with sulfur. There are three very
common types of iron-sulfur
center:[2fe-2s],[4fe-4s] and rieske
iron-sulfur center.
These iron-sulfur center consist
of equal number of iron and
sulfide.
Rieske fes-s

10. ComplexI;NADH-ubiquinone oxidoreductase
 Complex 1 is a largest of the four protein complex in the
mitochondrial electron transport system.
 It consisting of 45 polypeptide chain .
 L-shape structure.
 Complex 1 is to pass 2e obtained from the oxidation of NADH
to Q using a coupled reaction mechanism that result in the net
movement of 4H+ across the membrane .
 Complex 1 contain covalently bound flavin
mononucleotide(FMN) that accepts the two electron from
NADH.
 It transfer a electron from series of six Fe-s centre and further
transfer to Q .

11. 1. It has three critical role :
Serves as a mobile electron carrier that transport electron
laterally in the membrane from complex 1 to complex 3 .
Q is the entry point into the electron transport system for
electron pair (2e).
Q(semiquinone) has the important task of converting 2e
transport system into a 1e transport system which passes
electron one at a time to the mobile electron carrier
cytochrome c.
This conversion process is accomplished by the Q cycle.

13. ComplexII :succinate dehydrogenase
 4 subunit
 It consisting FAD, Fe-s prosthetic group.
 Succinate dehydrogenase an inner mitochondrial membrane
bound enzyme, is an integral component of the complex2.
 It convert succinate to fumrate during kreb cycle.
 The 2e are released during conversion of succinate to fumarate
are transferred to FAD, then to an iron- sulfur center and
finally to coenzyme Q.
 Coenzyme Q draws electron into the respiratory chain, not
only from NADH but also from FADH2.
 It does not pump proton during transport of electron across the
inner mitochondria membrane.

15. ComplexIII;cytochrome c oxidoreductase
 Complex1 and complex2 transfer electron to the complex3 via
coenzyme Q.
 Within complex3, the electron released from coenzyme Q
follow two path.
 In other path, electron are transported via Rieske iron-sulfur
centre and cytochrome1, directly to cytochrome c.
 In other path, electron move through b-type cytochrome and
reduce oxidized coenzyme Q.
 During transport each pair of electron from coenzyme Q to
cytochrome c complex3 pump four proton across the inner
mitochondria membrane.
 The mechanism involve in the proton pumping is called the
proton- motive Q cycle.

17. Q- Cycle
 The mechanism of the participate of ubiquinone in the electron
transport process was proposed by Peter Mitchell and termed
as a proton motive Q-cycle.
 Four steps of the Q Cycle
1. oxidation of QH2 at the Qp site result in transfer of one
electron to Rieske Fe-S center. Which is transferred to
cytochrome C1 and then passed off to Cyt-c. The second
electron is transferred to cytochrome bL, The oxidation of QH2
in this first step contribute 2H+ to the inner- membrane space.

18. 2. The oxidized Q molecule moves from the Qp site to the Qn
site through a proposed substrate channel within the protein
complex. This stimulates electron transfer from bL to bH
which then reduced Q in the Qn site to from the semi Quinone
intermediate.
3. A new QH2 molecule bind in the vacated Qp site and is
oxidized in the same way as step 1 such that one electron is
transferred to cytochrome C1 and then to a new molecule of
cyt-c. Oxidation of this second QH2 molecule translocated
another 2H+ into the inter membrane space (4H+ total ) and the
resulting Q molecule released into the membrane.
4. The second electron from the QH2 oxidation in the step3 is
passed directly from bL to bH and then used to reduce the
semiquinone intermediate already sitting in the QN site which
uses 2HN to regenerate a QH2 molecule.
5. The Q cycle require that 2H+ from the matrix be used to
regenerate QH2, even though 4H+ are translocated.

19. 2H+ 2H+
Step-1 Step-2
2H+

20. ComplexIV: Cytochrome C oxidase
 The mitochondrial complexIV protein consist of two
monomers of ~200kDa that each contain 13 polypeptide, two
copper center (CuA and CuB) and two heme group
(cytochrome a and cytochrome a3) .
 Cys C dock on the p site of the membrane to complex 4 near
CuA which accepts the electron leading to oxidation of the
heme group in Cyt C (Fe+2>Fe+3).
 The reduced CuA passes the electron to an iron atom in the
heme of cytochrome a which then transfers it to cytochrome
a3.
 Finally, the electron passed to CuB which donate it to oxygen.

22. H+ H+ H+

23. Electrochemical proton gradient
 Transfer of electron through the electron transport chain is
accompanied by pumping of proton across inner mitochondrial
matrix to inter membrane space.
 A total of 10H ion are translocate from the matrix across the
inner mitochondria membrane per electron pair flowing from
NADH to O2. This movement of H generates:
 Ph gradient across the inner mitochondria membrane (with the
Ph higher in the matrix than in the inter membrane space).
 Voltage gradient (membrane potential) across the inner
mitochondria membrane (with the inside negative and outside
positive).
 The Ph gradient and voltage gradient together constitute
electrochemical proton gradient .

24. The electrochemical proton gradient exerts a proton
motive force (pmf).
A mitochondria actively involved in aerobic respiration
typically has a membrane potential of about 160mV and a
Ph gradient of about 1ph unit (higher on the matrix side).
The total proton motive force across the inner
mitochondrial membrane consists of a large force due to
the membrane potential and a smaller force due to the H+
concentration gradient (ph gradient).

25.  Structure and function ofA
TP synthase
complex.
⚫When Mitchell proposed the chemiosmotic theory there was
already evidence that a large protein complex in the inner
mitochondria membrane was responsible forA
TP synthesis .
⚫Originally called complexV and later purified as an ATP
synthase complex.
⚫Mitochondrial ATP synthase complex consists of two large
structural component :
⚫Large structural component called F1 which encodes the
catalytic activity.
⚫Another subunit called F0 which is function as the proton
channel crossing the inner mitochondrial membrane.

27. Three functional unit of ATP synthase
complex
1. The rotor turned 120 degree for every H that cross the
membrane using the molecular “carousel” called C ring.
2. The catalytic head piece contain the enzyme active site
in each of the three beta subunit and contain three alpha
subunit.
3. The stator consist an of the alpha subunit imbedded in
the membrane which contain two half channels for
proton to enter and exit the F0 component .

28. Proton flow through Fo alter the conformation of
F1 subunit
Nucleotide binding studies revealed that it was affinity of
the beta subunit for ATP, not rate of ATP synthesis, that
was alter by proton flow through F0 component.
Paul boyer proposed the binding change mechanism of
ATP synthesis to explain how conformational change in
beta subunit controlA
TP production .

29. The binding change mechanism
The gamma subunit directly contact all three beta subunit
however, each of these interaction giving rise to three
different beta subunit conformation .
The ATP binding affinity of the three beta subunit
conformation are define as : T tight, L loose and O open .
The binding change mechanism model predict that one
full rotation of the gamma subunit should generate three
ATP.

30. The three alpha beta dimer have three different state :-
1. O state that bindA
TP ,ADP and Pi very weakly.
2. L state that bindADP and Pi loosely .
3. T state that bindADP and Pi very tightly and giveATP
In logical intermediate stage, rotation of the gamma with
hexamer convert the L state to a T state, the T state to an O
and the O state to an L state. The L state can accept new
charge of substrate, the T state can formATP.

32. Inhibitor of electron transport chain
These compound prevent the passage of electron binding
to a component of the chain, blocking the oxidation
reduction reaction .
Rotenone a plant product inhibit the transfer of electron
through comlpex1. It is used as fish poison and as an
insecticide .
Barbiturates also act as same site and inhibit the electron
transport through complex 1 .

33. Piericidin , an antibiotic block the transfer of electron at
complex I by competing with Q. The electron from
complex are transfer to a piericidin instead of Q.
Carboxin is inhibited the complex II.
Antimycin A, also an antibiotic block electron transport at
the level of complexIII.
Cyanide, azide and carbon monoxide bind with complex
IV and inhibit the terminal transfer of electron to oxygen.

34. Uncoupling agent and ionophores
Uncoupling agent uncouples oxidation from
phosphorylation .
They allow the oxidation of NADH and FADH2 and
reduction of O2 to continue at high level but not permit
ATP synthesis. Thus, electron transport continues
unabated, butA
TP synthesis stop.
Most common uncoupling agent are:-
1. 2,4-dintrophenol(DNP)
2. Dicoumarol and FCCP
3. Thermogenin

35. DNP is hydrophobic molecule that can easily diffuse
across the membrane and in the process, carry proton one
a time from the inner mitochondrial space (high H)to the
matrix (lowH).
DNP is functioning as uncoupling agent because it
uncouples redox energy available from the electron
transport system fromA
TP synthesis.
Uncoupler such as DNP have been used as “diet pills”
because they stimulate the body to oxidize fat in response
to a chronic state of low energy charge.

36. The energy released by the oxidation of NADH in the
presence of DNP is converted to heat.
Dicoumarol and FCCP act in the same way.
Thermogenin is the physiological uncoupler found in
brown adipose tissue that function to generate body heat,
particularly for the new born and during hibernation in
animal

37. IONOPHORES
Ionophore uncouple electron
phosphorylation by dissipating the
transfer from oxidative
electro chemical
gradient across the mitochondrial membrane

Valinomycin, an antibiotic is an example of ionophores it
addition make a inner mitochondrial membrane permeable
for k+.

38. Shuttle systems
is a primary source of NADH
The glycolytic pathway
formation
NADH synthesize during glycolytic process finally
transfer the electron to electron transport chain
NADH can not cross the inner mitochondrial memebrane.
So, two different shuttle system helping the transfer of
electron from NADH to the ETC.

39. Malate – aspartate shuttle
The principle mechanism for the movement of NADH
from the cytoplasm into the mitochondrial matrix
Electron are carried into the mitochondrial matrix in the
form of malate .
Cytoplasmic malate dehydrogenase reduce oxaloacetate
to malate while oxidizing NADH to NAD.
Malate then enter the mitochondrial matrix, where the
reverse reaction is carried out by mitochondrial malate
dehydrogenase and the regeneration of NADH occurs.

41. Glycerol 3 –phosphate shuttle
• mitochondrial electron transport chain by being use to reduce
dihydroxy acetonephosphate to glycerol 3- phosphate
 Glycerol 3- phosphate is reoxidise by electron transfer to Q to
from QH2,which allows these electron enter the electron transfer
chain.