Electron Transport

System dan
Fosforilasi
Oksidatif
Jumeri M. Wikarta, Ph.D

Overview of Cellular Respiration

Step 1: Glycolysis
Glucose + 2ADP
2ATP

2 pyruvate +

Glycolys
is
Occurs in the cytosol

Hi [ATP]

Glucose metabolized to
2 pyruvate + 2 ATP
High [ATP] inhibits
phosphofructokinase
(PFK)
High [ADP] stimulates
PFK
Pasteur Effect:

Increase in the rate of

carbohydrate
breakdown that occurs
when switched from
aerobic to anaerobic
conditions
Fig. 16-3

Step 2: Citric Acid Cycle

Mitochondri
a

Citric Acid
Cycle

Citric Acid Cycle

a.k.a. Krebs Cycle, TCA Cycle

Occurs in mitochondrial
matrix

Pyruvate reacts with CoA to

form Acetyl CoA

NAD+, FAD+ reduced to NADH,

FADH2,

NADH, FADH2 enter the

electron transport chain

 Substrate level phosphorylation

Oxidative phosphorylation

Step 3: Electron transport chain and oxidative

phosphorylation

Overview of the Electron

Transport Chain ( Respiratory
Chain )

Electron transport and oxidative

phosphorylation

Complete oxidation of glucose by molecular oxygen can be

described as:
C6H12O6 + 6O2 6CO2 + 6H2O
Go=-2823 kJ/mol
Can be broken down into two half-reactions with the transfer
of electrons
C6H12O6 + 6H2O 6CO2 + 24H+ +24e6O2 + 24H+ + 24e- 12H2O
12e- from the oxidation of glucose are not transferred directly
to O2, go to NAD+ and FAD to form 10NADH and 2FADH2
These are reoxidized, passing their electrons to the
electron- transport chain to reduce O2 to H2O causing
the mitochondrion to create a proton gradient.
This pH gradient is used to drive the synthesis of ATP via
oxidative phosphorylation.

Chemiosmotic Theory
Electron Transport: Electrons carried by reduced coenzymes
are passed through a chain of proteins and coenzymes to drive
the generation of a proton gradient across the inner
mitochondrial membrane
Oxidative Phosphorylation: The proton gradient runs
downhill to drive the synthesis of ATP
Electron transport is coupled with oxidative
phosphorylation
It all happens in or at the inner mitochondrial membrane

Oxidative Phosphorylation
Oxidative phosphorylation is the process by which the
energy stored in NADH and FADH2 is used to produce ATP.
A. Oxidation step: electron transport chain
1
2
1
FADH2+O2
2

NADH+H++O2

NAD++H2O
FAD+H2O

B. Phosphorylation step
ADP+Pi

ATP

Oxidative
Phosphorylati
on

H+ transport results in an
electrochemical gradient

Proton motive force:

energy released by flow
of H+ down its gradient is
used for ATP synthesis

ATP synthase: H+
channel that couples
energy from H+ flow with
ATP synthesis

Fig. 16-32

Summar
y
Glucose
ATP

Fig.169

Yeast ethanol
metabolism
EtOH
ADH

Glucose
ATP

acetaldehyde
acetic acid

CoA

Electron transport chain

inhibitors and substrates
rotenone

Antimycin A
Sodium azide

Ascorbate + TMPD
Glutamate, malate

Fig.1619

Inhibitors and
uncouplers of oxidative
phosphorylation
Inhibitors

Atractyloside: ADP/ATP
antiporter

Atractyloside

Oligomycin:ATP synthase

oligomycin

Uncouplers

DNP shuttles H+ across inner DNP

membrane, dissipates gradient

CaCl2 stimulates oxidative

phosphorylation and ATP
production

Ca2+

Fig. 16-32

Electron Transport
Inhibitors

Uncouplers Disrupt the

Coupling of Electron Transport

Summary of Cellular
Energetics
Glucose

High [ATP]
(Pasteur effect)

Glycolysis

N-ethylmaleimide
Pyruvate
NADH
FADH2

EtOH

Acetyl CoA
Malate

Citric Acid Cycle

Succinate
Uncouplers
Ca+2, DNP

Fig. 16-2

NADH + FADH2

Rotenone

Electron transport chain

Energy released used to pump H+
creating an elecrochemical gradient

O2

Flow of protons down the gradient fuels ATP synthase

Atractyloside

Antimycin A
Ascorbate + TMPD
Sodium Azide

ADP + Pi

Oligomycin
ATP

H2O

Oxidative
Phosphorylatio
n

Standard reduction
potentials of the major
respiratory electron
carriers.

See movie

Electron transport, also known as aerob

respiration, is the last stage of aerobic
metabolism.
After glycolysis and TCA cycle, 10 NADH
and 2 FADH2 are generated from the
oxydation of one glucose molecule.

dehydrogenase

AH2 + NAD
+ H+
Or
dehydrogenase

BH2 + FAD

A + NADH

B + FADH 2

 When there is sufficient O2 supply, NADH

and
FADH2 enter electron transport chain to
become reoxidized

Large amount of energy is recovered,

when
electrons are passed from NADH and
FADH2
to O2.

This is accomplished by a series of carrier

protein in the inner mitochondrial
membrane .

Mitochondrial Electron Transport

Folding in the mitochondria inner
membrane provides a large surface
area.
Electron-transport chain components
are arranged in packages called
respiratory assemblies.
assemblies

Electron Transport and Oxidative

Phosphorylation:
In ETS, electrons are transferred from
NADH
and FADH2 to O2 step by step.
In the mean time, energy released from
electron flow is coupled to ATP synthesis.
Here, phosphorylation of ADP is coupled
with
The oxidation of NADH or FADH2.

ADP + Pi

ATP

NADH + H+ + 1/2 OATP

2
synthase
ADP + Pi

FADH2 + 1/2 O2

ATP

ATP synthase

NAD+ + H2

FAD + H2O

Theenergyreleasedduringtheelectronflow
iscoupledtoATPsynthesis.

Composition of the Electron

Transport Chain
Four large protein complexes.
Complex I - NADH-Coenzyme Q reductase
Complex II - Succinate-Coenzyme Q reductase
Complex III - Cytochrome c reductase
Complex IV - Cytochrome c oxidase

Many of the components are proteins with

prosthetic groups to move electrons.

Electron Transport
Four protein complexes in the
inner mitochondrial membrane
A lipid soluble coenzyme (UQ,
CoQ) and a water soluble
protein (cyt c) shuttle between
protein complexes
Electrons generally fall in
energy through the chain from complexes I and II to

Complex
I

Complex
IV

Complex
III
cyt
c1

CoQ

cyt b
FADH2

Fe
S

cyt
c

(Cu)
cyt
a/a3

Complex II
NADH

matrix

O
2

Cofactors of the electron transport

chain

Fe-S clusters
Coenzyme Q (ubiquinone)
Flavin mononucleotide
FAD
Cytochrome a
Cytochrome b
Cytochrome c
CuA
CuB

Electron Transport Chain

Mitochondria
outer membrane relatively permeable
inner membrane permeable only to
those things with specific transporters
Impermeable to NADH and FADH2
Permeable to pyruvate

Compartmentalization
Kreb's and -oxidation in matrix
Glycolysis in cytosol

Outer Membrane Freely

permeable to small molecules
and ions. Contains porins with 10
kDa limit
Inner membrane Protein rich
(4:1 protein:lipid). Impermeable.
Contains ETR, ATP synthase,
transporters.

Cristae Highly folded inner membrane structure. Increase

surface area.
Matrix- cytosol of the mitochondria. Protein rich (500 mg/ml)
Contains TCA cycle enzymes, pyruvate dehydrogenase, fatty and
amino acid oxidation pathway, DNA, RNA, ribosomes
Intermembrane Space composition similar to cytosol

Mitochondrial transport
The inner membrane is impermeable to
hydrophilic substances. Has special transport
systems for the following:
1. Glycolytically produced cytosolic NADH.
2. Mitochondrially produced metabolites (OAA,
acetyl-CoA) for cytosolic glucose formation and
fatty acid biosynthesis.
3. Mitochondrially produced ATP must go to cytosol
where ATP-utilizing reactions take place.
Example: cytoplasmic shuttle systems transport
NADH across inner membrane.

 Important characteristic of the

electrontransport chain:
Electron carriers are arranged in order
of increasing electron affinity, from low
to high.
This results in the spontaneous flow of
electrons from carrier to carrier.

Eo, standard reduction potentials.

The more positive the Eo value a molecule has,
the
better it serves as an electron acceptor.
O2 has the highest Eo (0.82V), - highest affinity
for electrons and is located in the end of the
chain.

Eo for NAD is 0.32, lowest in the system, its

the
poorest electron carrier, located in the
beginning.

-0.4
-0.2

NADH
(-0.32)
NAD+

Path of
Electrons
Complex I

0.0
succinate
(FADH2)
0.2
(0.18)
fumarate

Q
Complex II

Complex III
cyt c

(0.25)

0.4

Complex IV
0.6
0.8
1.0

Table 17.2 !

1/2 O2 + 2 H+

(0.82)

HO
2

Components of the
electron transport chain
Complex I
Electrons pass from
NADH FMN Fe-S cluster ubiquinone

(flavin mononucleotide)

(coenzym

Fe-S cluster: iron cycles between 3+

and 2+

2 H+

FMN

Complex I

FMNH2

2 electrons

Fe-S

QH2

2 electrons

2 H+
NADH

H+

NAD+

Complex I

NADH-CoQ Oxidoreductase (NADH dehydrogenase)

Electron transfer from NADH to CoQ
More than 30 protein subunits - mass of 850 kD
1st step is 2 e- transfer from NADH to FMN
FMNH2 converts 2 e- to 1 e- transfer
6-7 FeS clusters.
Four H+ transported out per 2 e-

NADH + H+
NAD+

FMN

Fe2+S

CoQ

FMNH2

Fe3+S

CoQH2

 CoQ ubiquinone
Highlighted region serves as an anchor to inner
mitochondrial membrane.

O
H3CO

CH3
CH3

H3CO

(CH2
O

CH

CH2)10

Reduction of CoQ
Oxidized form
Ubiquinone (CoQ)

Reduced form
Ubiquinol (CoQH2)

OH

H3CO

CH3

H3CO

CH3

H3CO

H3CO

2e 2H+

OH

 Complex II
Entry point for FADH2.
Succinate dehydrogenase (from the
citric acid cycle) directs transfer of

electrons from succinate to CoQ via FADH2.

Acyl-CoA dehydrogenase (from -

oxidation of fatty acids) also transfers

electrons to CoQ via FADH2.

Complex II

Succinate-CoQ Reductase
Contains the succinate dehydrogenase (from TCA cycle!)
four subunits
Two largest subunits contain 2 Fe-S proteins
Other subunits involved in binding succinate dehydrogenase
to membrane and passing e- to Ubiquinone
FAD accepts 2 e- and then passes 1 e- at a time to Fe-S
protein
No protons pumped from this step

Succinate
Fumarate

FAD

Fe2+S

CoQ

FADH2

Fe3+S

CoQH2

 All electrons from FADH2 and NADH must pass through

CoQ.

innermembrane
space
I
Fe-S
FMN

II

Fe-S
FAD

NADH

NAD+

CoQ

FAD

Succinate
Fatty acyl
CoA

matrix

 Complex III (cytochromes b, c1 and c).

Electron transfer from ubiquinol to cytochrome c.

cytochrome c

heme prosthetic group

Cytochromes are electron-transfer

proteins that contain a heme
prosthetic group.
The iron atom in heme also cycles
through
reduced form (Fe2+) and the oxidized
form (Fe3+).
Red muscles are rich in mitochondria,
which
contains electron transport system and

 Complex IV
Combination of cytochromes a and a3,
10 protein subunits, 2 types of
prosthetic groups: 2 heme and 2 Cu.
Electrons are delivered from
cytochromes a and a3 to O2.
Several chemicals can inhibit the pathway at
different locations.
Cyanide and CO can block e transport between
a/a3 and O2.

Flow of electrons

-0.4
NADH
-0.2

Path of
Electrons

NAD+
Complex I

0.0
0.2

succinate
(FADH2)
fumarate

Complex II

Q
Complex III
cyt c

0.4
Complex IV
0.6
1/2 O2 + 2 H+
0.8
1.0

HO
2

Energy is not released at once, but in

incremental

Energy Yield
The amount of energy can be
calculated in
terms of Go .

Go = - nF Eo

n = electron number,
F = faraday constant =
96.5kJ/volt .mole
Eo = Eo
- Eo

Energy Yield
NADH + H+ + 1/2 O2

NAD+ + H2O

Go = - 220 kJ/mol
FADH2 + 1/2 O2

FAD + H2O

Go = - 152 kJ/mol
Note: ADP + Pi

ATP

Go = + 31 kJ/mol

Energy yield from one FADH2 is less than

one
NADH.

Oxidative phosphorylation
The electron-transport chain moves electrons
from NADH and FADH2 to O2.
In the mean time, ADP is phosphorylated to ATP.
The two processes are dependent on each other.
ATP cannot be synthesized unless there is energy
from electron transport (Go= +31 kj/mol).
Electrons do not flow to O2, unless there is need
for ATP.

3 ATP are generated when two electrons

are transported from NADH to O2.

The oxidation of FADH2 only produces 2 AT

Coupling of electron-transport
with ATP synthesis
Chemiosmotic coupling mechanism
Electron-transport causes
unidirectional movement of H+ into
the inner membrane space.
The results in a H+ gradient being
produced.
The gradient then drives the
synthesis of ATP.

Outer mitochondrial membrane

H+

H+

H+

H+
H

H+
H+

H+

H+
H+

H+

Inner mitochondrial membrane

H+ H+

H+
Electron
Electron
Transport
Transport
Chain
Chain

ATP
ATP
synthase
synthase
complex
complex

ADP + Pi

H+

ATP

Components of ATP synthase

These are knob-like projections into the matrix
side of the inner membrane.

Two units
F1 contains the catalytic site for ATP synthesis.
F0 serves as a transmembrane channel for H +
flow.

F1-F0 complex serves as the molecular apparatus

for coupling H+ movement to ATP synthase.

Components of ATP synthase

H+
Cytosole
H+

H+

H+
H+

H+

H+

H+
H+

H+

F0

Matrix

F
1

ADP + Pi

ATP

ATP is transported from the matrix of

mitochondria to cytosole by ATP-ADP
translocase.
ATP and ADP cannot diffuse through the
mitochondria membrane freely.

The exit of ATP is coupled with the

entry of ADP into mitochondria.

Regulation of oxidative
phosphorylation
Electrons do not flow unless ADP is
present for phosphorylation
Increased ADP levels cause an increase in
the activity of various enzymes including:
glycogen phosphorylase
phosphofructokinase
citrate synthase

Uncoupling of electron-transport
and oxidative phosphorylation
In some special cases, the coupling of the two
processes can be disrupted.
Large amounts of O2 are consumed but no ATP
is produced.
Used by newborn animals and hibernating
mammals.
Occurs in brown fat- which contain
thermogenin (uncoupling protein).
Thermogenin allows the release of energy as
heat instead of ATP.

Energy production from

glucose
Glycolysis

2 ATP 2 ATP

2 NADH
3 ATP/NADH
Citric Acid Cycle

2 GTP1 ATP/GTP
2 ATP

6 NADH
3 ATP/NADH

2 FADH2
2 ATP/FADH2

38 ATP

(in heart)
* 4 ATP in muscle and brain.
36 ATP / glucose

6 ATP*

18 ATP
4 ATP

Mitochondria
Glycolysis
Glucose
Glucose

22Pyruvate
Pyruvate

22NADH
NADH

22Acetyl
AcetylCoA
CoA

22NADH
NADH

66NADH+
NADH+
22FADH
FADH2
2

22GTP
GTP

Oxidative
Oxidative
phosphorylation
phosphorylation

22 ATP
ATP

32-34
32-34 ATP
ATP

22 ATP
ATP

Recycling of cytoplasmic NADH(?)

Different methods are used to recycle
NADH. This accounts for the different
energy productions from glucose.
Glycerol-3-phosphate shuttle

Used by skeletal muscles and the brain

Malate-aspartate shuttle

Used by the heart and liver

Transport of NADH across inner mito

membrane

Glycerol phosphate shuttle

1.

Simpler but less energy efficient than the malateaspartate shuttle.

2.

Supplies electrons in a manner similar to succinate

dehydrogenase

3.

Approx. 2 ATP per NADH, about 0.7 ATP less than the
malate-aspartate shuttle.

4.

Advantage-irreversible so it operates efficiently even

when cytoplasmic NADH is low relative to NAD +.
Malate-aspartate shuttle is reversible - driven by
concentration gradients.

5.

Reoxidation of Cytosolic NADH

The glycerol 3-phosphate shutt

NAD+

cytoplasmic
glycerol-3-phosphate
dehydrogenase

NADH + H+

Glycerol phosphate
shuttle

Malate-aspartate shuttle
2 phases
Phase A
1. Cytosolic NADH reduces OAA to malate (malate DH).
2. Malate--ketoglutarate carrier transports malate from
cytosol to mitochondrial matrix, exchanged for -KG
3. In the matrix, NAD+ reoxidizes malate to make OAA and
yield NADH.
Phase B
4. Transaminase converts OAA to Asp and Glu to -KG.
5. The glutamate-apartate carrier transports Asp from the
matrix in exchange for Glu.
6. Transaminase in cytosol converts Glu to Asp.
3 ATPs for every NADH but loses 0.3 ATP because each NADH
enters matrix with proton (yield 2.7 ATP).

Malate-aspartate shuttle
Matrix
L-aspartate

L-aspartate

-ketoglutarate

Glycolysis

cytoplasmic
aspartate
aminotransferase

L-glutamate
oxaloacetate

cytoplasmic malate
dehydrogenase

NADH + H+

L-malate
NAD+

-ketoglutarate
mitochondrial
aspartate
aminotransferase

L-glutamate
mitochondrial
malate
dehydrogenase

L-malate

oxaloacetate

NAD+

3 ATP
NADH
+ H+

 The malate-aspartate shuttle

ATP Synthesis and glucose

metabolism

C6H12O6 + 6 O2 + 36 Pi +36 ADP + 36 H+

6 CO2 + 36 ATP + 42 H2O