Oxidative Phosphorylation

Part I
Cellular respiration - is the process by
which cells consume O2 and produce ● Breakdown of glucose to CO2
CO2. starting from glycolysis where it is
broken down to pyruvate.
RECAP: ● Pyruvate is prepared for the citric
acid cycle via prior conversion to
acetyl-CoA which carries the two
carbon fragments from pyruvate.

2. Oxidation to CO2 in the citric

cycle

● Acetyl-CoA then enters the citric

cycle where it is further oxidized to
CO2. And in the process, electron
carriers NADH and FADH2 transfers
some of the energy from the
oxidation of acetyl to CO2.

3. Electron transport or “respiratory

chain” which drives the
production of ATP

● The electron carriers are shuttled in

the next step stage of cellular
respiration which is the oxidative
phosphorylation. And the electron
transport chain as well.

OXIDATIVE PHOSPHORYLATION

● The stage where the mechanism of

energy production in the
mitochondria is at the heart.
It occurs in 3 major stages: ● Mitochondria is called the
“powerhouse of the cell” because
1. Production of acetyl-Coa from the transport of electrons from one
fuels such as glucose, fatty acids, electron carrier to another ultimately
amino acid to oxygen as the final electron
 acceptor will be the reason why we
are able to synthesize ATP in the
mitochondrial matrix.
STRUCTURE OF THE MITOCHONDRIA
- Composed of a double membrane
due to its evolutionary origin
● Outer membrane
○ Separates the mitochondria
from the rest of the
cytoplasm
○ porous, allows passage of
metabolites, particularly the
ATP that the mitochondria
synthesizes from ADP
● Intermembrane space
○ similar to cytosol in
composition
○ the process of electron
transport causes this space
to have a high concentration
of H+ ions (or protons) or a
low pH environment
● Inner membrane
○ relatively impermeable to
metabolites; it sorts of traps
ions and molecules inside
the matrix
○ membrane-bound complexes
of respiratory chain (in order
to transport substances in
and out of the inner
membrane, you would need
to do so via membrane-
bound complexes)
● Matrix
○ PDH (pyruvate
dehydrogenase) complex
and all the enzymes of the
citric acid cycle are found
(dissolved) and recycled in
the matrix with the exception
of succinate
dehydrogenase which is
embedded in the inner
membrane; some enzymes
are shuttled out of the
mitochondria
- Lower [H+] or high pH
HOW DOES THE MITOCHONDRION
BEHAVE AS THE POWERHOUSE OF
THE CELL?
● The oxidative phosphorylation
step of the cellular respiration for
eukaryotes happens in the
mitochondria while for
prokaryotes, which lacks
 mitochondria, only happens in the
cytosol (cytoplasm)
It is a sequence of processes influencing
the other.
1. Flow of electrons coming from
reduced substrates such as NADH
and FADH2 from the citric acid
cycle and glycolysis serves as
electron carriers which are the
starting point of the ETC
(respiratory chain) where these
electrons are being passed along
across these several complexes and
many electron carriers ultimately
transferring electrons to oxygen.
2. Oxygen, as the final electron
acceptor, will be reduced to water
and that completes the redox
reaction of oxidizing fuels.
● This transfer of electrons is a
“downhill” in terms of
thermodynamic driving force
from high to low energy and
releases a lot of free energy
(a very favorable,
spontaneous process) which
we could coupled to the
“uphill” (endergonic) pumping
of protons from the matrix
into the inner mitochondrial
space of your mitochondrion.
*So, protons are transported from the
matrix environment which is kept at a
low concentration of H+ ions into the
intermembrane space which is kept at a
high H+ concentration or low pH. This is
against the natural flow from high to low
concentration. Instead, we are
transporting protons from low to high
which is against the concentration
gradient. Therefore, it is uphill and
requires a lot more energy from the
“downhill” flow of electrons via ETC or
the respiratory chain.
3. Oxidative Phosphorylation
● The proton gradient or the
concentration difference of H+ ions
created across the inner membrane
will then be used to drive the ATP
 synthesis that is carried out by the
mitochondrial inner membrane
● The enzyme ATP synthase exploits
or harnesses the energy coming
from the proton gradient created
across the inner membrane in order
to form ATP from ADP
TYPES OF ELECTRON CARRIERS
1. Nicotinamide Nucleotides
● NAD+ (Nicotinamide adenine
dinucleotide)
● NADP+ (other form of NAD+ and is
phosphorylated in one of the
carbons shown in the black arrow at
the bottom of this figure)
● The role of NAD is to remove 2
hydrogen atoms from a substrate
and these two will come out as 1
hydride ion (equivalent to 1 proton
and 2 electrons) added to the NAD.
This NAD will be reduced to NADH
and will proceed to the respiratory
chain. The other hydrogen atom is
released as proton in solution
○ So whenever NAD+ is
reduced, the products are
1 NADH which contains the
hydride ion carrying the
pair of electrons plus a
proton that is released in
the solution (NADH + H+)
● These nicotinamide nucleotides
associate reversibly with
dehydrogenases so they are only
held by weak interactions that may
detach from dehydrogenase
enzymes catalyzing such redox
reactions so they are soluble in
aqueous environments.
● NADH will be the one to carry
electrons from catabolic reactions
just like glycolysis and citric acid
cycle and even the oxidation of
lipids to the respiratory chain in
the mitochondrion
● NADPH appears primarily in
anabolic reactions or those that
carry out the biosynthesis of larger
molecules and are found mainly in
the cytosol.
● So, the cell keeps separate pools
of NADH mainly in the
mitochondria and NADPH in the
cytosol
2. Flavin Nucleotides (FAD and FMN)
● FAD (Flavin adenine dinucleotide)
● FMN (Flavin mononucleotide)
ubiquinone is converted to a free
radical called semiquinone (QH)
and gaining two electrons, the
ubiquinone is transformed to its
fully reduced form, ubiquinol
(QH2)
- Can carry protons and
electrons important in the
function of the respiratory
chain that is to transport
electrons across complexes
and pumping protons out of
the matrix

● These two are tightly bound

(sometimes covalently) to
flavoproteins. Unlike NAD+ and
NADP+, FAD and FMN are not
released in the solution. Instead,
they remain at the active site of an
enzyme (flavoprotein)
● Accepts one or two hydrogen
atoms 4. Iron-sulfur proteins
● Can carry protons and electrons

3. Ubiquinone, Coenzyme Q or “Q”

● Lipid-soluble benzoquinone with
isoprenoid side chain
Note: Quinone is a
substance with two double
bonds (2 C=O) in the
hexagon hydrocarbon ring; it
is what happens when you
oxidized phenol ● Contains an iron atom at the center
● It is a small molecule that is lipid- of the structure
like which is able to travel across ● In iron-sulfur proteins, the iron atom
a phospholipid bilayer of the inner is found in association with inorganic
membrane sulfur atoms that are either a stand-
● Capable of doing 1 to 2 electron alone or bound to a cysteine residue
transfers. So transferring one ● As an electron carriers, the iron
electron or gaining one electron, atoms, specifically, can switch
 between the ferrous 2+ oxidation ● In general, the process involves
state to ferric 3+ oxidation state the transfer of electrons starting
and vice versa as it gains or from electron carriers, NADH and
releases one electron FADH2, that is shuttled from
● It facilitates one-electron transfers various pathways to O2 as the
only final electron acceptor
● It involves four complexes: I, II, III, IV

5. Cytochrome proteins

COMPLEX I: NADH TO UBIQUINONE

● Contains an iron atom found at

the center of a heme group similar
to the hemoglobin and myoglobin
structures
- So the heme group is a
prosthetic group attached to
the protein or enzyme and
has a protoporphyrin planar
ring structure same with
myoglobin and hemoglobin
● Have three classes: cytochromes
a,b, and c which depends on its
structure
- They are named as such ● Also known as the
based on the way they NADH:ubiquinone oxidoreductase
absorb visible light or NADH dehydrogenase
- Strongly colored proteins ● Entry point of NADH in the
respiratory chain
RESPIRATORY CHAIN OR ELECTRON ● Gets the electrons from NADH and
TRANSPORT CHAIN transfers it ultimately to ubiquinone
 and this ubiquinone is reduced to SUMMARY:
ubiquinol (QH2)
○ The transfer is not ● Complex I carries out two functions:
straightforward as it is
passed on to a series of 1. The matrix arm performs the
molecules. catalytic function of converting
○ First, the electrons go to NADH to NAD+
FMN which is embedded in a 2. Reduction of ubiquinone to ubiquinol
flavoprotein part of the arm of - Transfer of electrons
this complex.
○ The matrix arm which is COMPLEX II: Succinate dehydrogenase
protruding into the matrix will
be the one to react with the
NADH catalytically in
performing the functions of
reoxidizing NADH to NAD+
and getting those pair of
electrons
● So, electrons will travel from FMN
through a series of iron-sulfur
centers ultimately to ubiquinone
as the final electron acceptor for
Complex I. Ubiquinone will be
reduced to ubiquinol as it carries
those pairs of electrons and the
ubiquinol (QH2) will travel to
Complex III.
● The redox reaction of converting
NADH to NAD+ is coupled with
the pumping of four (4) protons
(H+) for every NADH that transfers
electrons out of the matrix and ● Entry point of FADH2 in the
into the intermembrane space. respiratory chain (but FAD is part
○ The Matrix is called the N of the complex itself and not as a
side or negative side free coenzyme dissolved in the
because it has the lower matrix)
concentration of protons and ● Complex II is also succinate
therefore, a negative charge dehydrogenase, exactly the same
or pole in relation to the enzyme in this step of the citric acid
intermembrane space cycle where succinate is converted
which is called the P side into fumarate and likewise FAD
(higher concentration of H+ carries electrons reduced to FADH2
ions)
 ● The enzyme is membrane-bound, all
the other enzymes in the citric acid
cycle are dissolved to the matrix.
● FAD is treated as an electron carrier
but in reality, it stays as a part of this
dehydrogenase enzyme. FAD, as an
electron carrier, is just an
intermediary. It will transfer electrons
through a path of iron-sulfur centers
and one cytochrome b all the way to
ubiquinone, again, as a final COMPLEX III: cytochrome bc1 or
acceptor for this complex. And this ubiquinone:cytochrome c
will be reduced to ubiquinol (QH2) oxidoreductase
and will be shuttled to Complex III.
● No pumping of protons (the only
complex in ETC)

SUMMARY:

Functions:

● A substrate, such as a succinate will

transfer electrons to FAD making
FADH2 and FADH2 will transfer
electrons to ubiquinone forming
ubiquinol (QH2) which will be
shuttled to Complex III

The major difference between Complex I

and II is that the latter does not involved
the pumping of protons so the delta G
associated with the redox reaction of
● Ubiquinol (QH2) from complexes I
succinate to fumarate (or FAD to FADH2)
and II will travel inside the inner
is not exploited or coupled to the
membrane where it is soluble and
“uphill” or pumping of protons out of the
go to complex III and pass its
matrix.
electrons to cytochrome c as the
final acceptor of Complex III

● Stoichiometry:

- For every 1 ubiquinol, 2

cytochrome c equivalence are
 required. This is because cyt c, as a
cytochrome, has the iron atom which
can only accommodate 1 electron
transfer at a time (it switches
between Fe2+ and Fe3+)

Complex III Stages:

- Quite more complicated than
Complexes I and II in that there is a
cycle involved in the regeneration
of ubiquinol to ubiquinone. This
allows cyt c to be used up twice and
carry the pair of electrons even
though it only accommodates only
one electron at a time.

1. Stage I
● The first electron from ubiquinone is
given up to one cyt c which carries
only one electron
● The ubiquinol will combine with one
ubiquinone forming a semiquinone
(the intermediate of a ubiquinol and
a ubiquinone)