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Oxida ve phosphoryla on
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Overview of oxida ve phosphoryla on. The electron

transport chain forms a proton gradient across the inner
mitochondrial membrane, which drives the synthesis of ATP
via chemiosmosis.

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Why do we need oxygen?

You, like many other organisms, need oxygen to live.

As you know if you’ve ever tried to hold your breath
for too long, lack of oxygen can make you feel dizzy or
even black out, and prolonged lack of oxygen can
even cause death. But have you ever wondered why
that’s the case, or what exactly your body does with
all that oxygen?

As it turns out, the reason you need oxygen is so your

cells can use this molecule during oxida ve
phosphoryla on, the final stage of cellular respira on.
Oxida ve phosphoryla on is made up of two closely
connected components: the electron transport chain
and chemiosmosis. In the electron transport chain,
electrons are passed from one molecule to another,
and energy released in these electron transfers is used
to form an electrochemical gradient. In chemiosmosis,
the energy stored in the gradient is used to make ATP.

So, where does oxygen fit into this picture? Oxygen

sits at the end of the electron transport chain, where
it accepts electrons and picks up protons to form
water. If oxygen isn’t there to accept electrons (for
instance, because a person is not breathing in enough
oxygen), the electron transport chain will stop running,
and ATP will no longer be produced by chemiosmosis.
Without enough ATP, cells can’t carry out the
reac ons they need to func on, and, a er a long
enough period of me, may even die.

In this ar cle, we'll examine oxida ve phosphoryla on

in depth, seeing how it provides most of the ready
chemical energy (ATP) used by the cells in your body.

Overview: oxida ve phosphoryla on

The electron transport chain is a series of proteins

and organic molecules found in the inner membrane
of the mitochondria. Electrons are passed from one
member of the transport chain to another in a series
of redox reac ons. Energy released in these reac ons
is captured as a proton gradient, which is then used to
make ATP in a process called chemiosmosis. Together,
the electron transport chain and chemiosmosis make
up oxida ve phosphoryla on. The key steps of this
process, shown in simplified form in the diagram
above, include:

Delivery of electrons by NADH and FADH2 .

Reduced electron carriers (NADH and FADH2 )
from other steps of cellular respira on transfer
their electrons to molecules near the beginning of
the transport chain. In the process, they turn back
into NAD+ and FAD, which can be reused in other
steps of cellular respira on.

Electron transfer and proton pumping. As

electrons are passed down the chain, they move
from a higher to a lower energy level, releasing
energy. Some of the energy is used to pump H+
ions, moving them out of the matrix and into the
intermembrane space. This pumping establishes an
electrochemical gradient.

Spli ng of oxygen to form water. At the end of

the electron transport chain, electrons are
transferred to molecular oxygen, which splits in
half and takes up H+ to form water.

Gradient-driven synthesis of ATP. As H+ ions flow

down their gradient and back into the matrix, they
pass through an enzyme called ATP synthase,
which harnesses the flow of protons to synthesize
ATP.

We'll look more closely at both the electron transport

chain and chemiosmosis in the sec ons below.

The electron transport chain

The electron transport chain is a collec on of

membrane-embedded proteins and organic molecules,
most of them organized into four large complexes
labeled I to IV. In eukaryotes, many copies of these
molecules are found in the inner mitochondrial
membrane. In prokaryotes, the electron transport
chain components are found in the plasma membrane.

As the electrons travel through the chain, they go

from a higher to a lower energy level, moving from
less electron-hungry to more electron-hungry
molecules. Energy is released in these “downhill”
electron transfers, and several of the protein
complexes use the released energy to pump protons
from the mitochondrial matrix to the intermembrane
space, forming a proton gradient.
Click to see a free energy diagram

Image modified from "Oxida ve phosphoryla on: Figure 1", by OpenStax

College, Biology (CC BY 3.0).

All of the electrons that enter the transport chain

come from NADH and FADH2 molecules produced
during earlier stages of cellular respira on: glycolysis,
pyruvate oxida on, and the citric acid cycle.

NADH is very good at dona ng electrons in redox

reac ons (that is, its electrons are at a high energy
 level), so it can transfer its electrons directly to
complex I, turning back into NAD+ . As electrons
move through complex I in a series of redox
reac ons, energy is released, and the complex
uses this energy to pump protons from the matrix
into the intermembrane space.

FADH2 is not as good at dona ng electrons as

NADH (that is, its electrons are at a lower energy
level), so it cannot transfer its electrons to complex
I. Instead, it feeds them into the transport chain
through complex II, which does not pump protons
across the membrane.

Because of this "bypass," each FADH2 molecule

causes fewer protons to be pumped (and contributes
less to the proton gradient) than an NADH.
More about complexes I and II

Beyond the first two complexes, electrons from

NADH and FADH2 travel exactly the same route.
Both complex I and complex II pass their electrons to
a small, mobile electron carrier called ubiquinone (Q),
which is reduced to form QH2 and travels through the
membrane, delivering the electrons to complex III. As
electrons move through complex III, more H+ ions are
pumped across the membrane, and the electrons are
ul mately delivered to another mobile carrier called
cytochrome C (cyt C). Cyt C carries the electrons to
complex IV, where a final batch of H+ ions is pumped
across the membrane. Complex IV passes the
electrons to O2 , which splits into two oxygen atoms
and accepts protons from the matrix to form water.
Four electrons are required to reduce each molecule
of O2 , and two water molecules are formed in the
process. More about complexes III and IV

Overall, what does the electron transport chain do for

the cell? It has two important func ons:
 Regenerates electron carriers. NADH and FADH2
pass their electrons to the electron transport
chain, turning back into NAD+ and FAD. This is
important because the oxidized forms of these
electron carriers are used in glycolysis and the
citric acid cycle and must be available to keep
these processes running.

Makes a proton gradient. The transport chain

builds a proton gradient across the inner
mitochondrial membrane, with a higher
concentra on of H+ in the intermembrane space
and a lower concentra on in the matrix. This
gradient represents a stored form of energy, and,
as we’ll see, it can be used to make ATP.

Chemiosmosis

Complexes I, III, and IV of the electron transport chain

are proton pumps. As electrons move energe cally
downhill, the complexes capture the released energy
and use it to pump H+ ions from the matrix to the
intermembrane space. This pumping forms an
electrochemical gradient across the inner
mitochondrial membrane. The gradient is some mes
called the proton-mo ve force, and you can think of it
as a form of stored energy, kind of like a ba ery.

Like many other ions, protons can't pass directly

through the phospholipid bilayer of the membrane
because its core is too hydrophobic. Instead, H+ ions
can move down their concentra on gradient only with
the help of channel proteins that form hydrophilic
tunnels across the membrane.

In the inner mitochondrial membrane, H+ ions have

just one channel available: a membrane-spanning
protein known as ATP synthase. Conceptually, ATP
synthase is a lot like a turbine in a hydroelectric power
plant. Instead of being turned by water, it’s turned by
the flow of H+ ions moving down their
electrochemical gradient. As ATP synthase turns, it
catalyzes the addi on of a phosphate to ADP,
capturing energy from the proton gradient as ATP.

Image modified from "Oxida ve phosphoryla on: Figure 3," by Openstax

College, Biology (CC BY 3.0).

This process, in which energy from a proton gradient

is used to make ATP, is called chemiosmosis. More
broadly, chemiosmosis can refer to any process in
which energy stored in a proton gradient is used to do
work. Although chemiosmosis accounts for over 80%
of ATP made during glucose breakdown in cellular
respira on, it’s not unique to cellular respira on. For
instance, chemiosmosis is also involved in the light
reac ons of photosynthesis.

What would happen to the energy stored in the

proton gradient if it weren't used to synthesize ATP or
do other cellular work? It would be released as heat,
and interes ngly enough, some types of cells
deliberately use the proton gradient for heat
genera on rather than ATP synthesis. This might
seem wasteful, but it's an important strategy for
animals that need to keep warm. For instance,
hiberna ng mammals (such as bears) have specialized
cells known as brown fat cells. In the brown fat cells,
uncoupling proteins are produced and inserted into
the inner mitochondrial membrane. These proteins are
simply channels that allow protons to pass from the
intermembrane space to the matrix without traveling
through ATP synthase. By providing an alternate route
for protons to flow back into the matrix, the
uncoupling proteins allow the energy of the gradient
to be dissipated as heat.

ATP yield

How many ATP do we get per glucose in cellular

respira on? If you look in different books, or ask
different professors, you'll probably get slightly
different answers. However, most current sources
es mate that the maximum ATP yield for a molecule
of glucose is around 30-32 ATP2,3,4 . This range is
lower than previous es mates because it accounts for
the necessary transport of ADP into, and ATP out of,
the mitochondrion. More details

Where does the figure of 30-32 ATP come from? Two

net ATP are made in glycolysis, and another two ATP
(or energe cally equivalent GTP) are made in the citric
acid cycle. Beyond those four, the remaining ATP all
come from oxida ve phosphoryla on. Based on a lot
of experimental work, it appears that four H+ ions
must flow back into the matrix through ATP synthase
to power the synthesis of one ATP molecule. When
electrons from NADH move through the transport
chain, about 10 H+ ions are pumped from the matrix
to the intermembrane space, so each NADH yields
about 2.5 ATP. Electrons from FADH2 , which enter
the chain at a later stage, drive pumping of only 6 H+ ,
leading to produc on of about 1.5 ATP.
With this informa on, we can do a li le inventory for
the breakdown of one molecule of glucose:

Stage Direct products Ul mate ATP

(net) yield (net)

Glycolysis 2 ATP 2 ATP

2 NADH 3-5 ATP

Pyruvate 2 NADH 5 ATP

oxida on

Citric acid 2 ATP/GTP 2 ATP

cycle

6 NADH 15 ATP

2 FADH2 3 ATP

Total 30-32 ATP

Click here for a diagram showing ATP produc on

One number in this table is s ll not precise: the ATP

yield from NADH made in glycolysis. This is because
glycolysis happens in the cytosol, and NADH can't
cross the inner mitochondrial membrane to deliver its
electrons to complex I. Instead, it must hand its
electrons off to a molecular “shu le system” that
delivers them, through a series of steps, to the
electron transport chain.

Some cells of your body have a shu le system that

delivers electrons to the transport chain via
FADH2 . In this case, only 3 ATP are produced for
the two NADH of glycolysis.

Other cells of your body have a shu le system

that delivers the electrons via NADH, resul ng in
the produc on of 5 ATP.
In bacteria, both glycolysis and the citric acid cycle
happen in the cytosol, so no shu le is needed and 5
ATP are produced.

30-32 ATP from the breakdown of one glucose

molecule is a high-end es mate, and the real yield
may be lower. For instance, some intermediates from
cellular respira on may be siphoned off by the cell
and used in other biosynthe c pathways, reducing the
number of ATP produced. Cellular respira on is a
nexus for many different metabolic pathways in the
cell, forming a network that’s larger than the glucose
breakdown pathways alone.

Self-check ques ons

1. Cyanide acts as a poison because it inhibits

complex IV, making it unable to transport
electrons. How would cyanide poisoning affect 1)
the electron transport chain and 2) the proton
gradient across the inner mitochondrial
membrane?
Choose 1 answer:

The electron transport chain would speed up,

A
and the gradient would become stronger

The electron transport chain would stop, and

B
the gradient would decrease

Both the electron transport chain and the

C
gradient would stay the same

The electron transport chain would be re-

D routed through complex II, and the gradient
would become weaker
 Hint

2. Dinitrophenol (DNP) is a chemical that acts as an

uncoupling agent, making the inner mitochondrial
membrane leaky to protons. It was used un l
1938 as a weight-loss drug. How would DNP
affect the amount of ATP produced in cellular
respira on? Why do you think it is now off the
market?*
Choose 1 answer:

It would increase ATP produc on, but could

A also cause dangerously high body
temperature

It would decrease ATP produc on, but could

B also cause dangerously high body
temperature

It would decrease ATP produc on, but could

C
also cause dangerously low body temperature

It would increase ATP produc on, but could

D
also cause dangerously low body temperature

Hint

A ribu on and references

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Maulana Akmal 6 years ago

more

how does the nadh from glycolisys gets into

the matrix so its electron could be used?
1 (13
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5 years
m…
Eric Ballesteros ago

It's actually called Malate-aspartate

shu le
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na26262 4 years ago

more

if the volume of the intermembrane space

was increased, what effect would this have
on the func on of a mitochondrion?
(7
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4 years
m…
Richard Wu ago

Hm.... A cell stays small to allow

easier transport of molecules and
charged par cles from organelles. If
the intermembrane space of the
mitochondria was increased, I
would think that respira on would
be less efficient, because now the
electrons have to cross a larger
space and lose much more energy.
So.... That's my guess and it would
probably be wrong,
2
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Ashley Jane 3 years ago

more

Where do the hydrogens go? I mean in

glycolysis, one glucose is oxidised into two
pyruvic acid and two NADHs. But technically
there should be net two protons le in
cytosol and that's where I am puzzled. Are
the protons tansported into mitochondria
ma x and later pumped out by ETC or
intermembrane space to form
electrochemical gradient, or are they le in
cytosol? According to the amont of water
molecules generated in chemiosmosis, all the
hydrogen from the glucose should be used to
form water, so do protons go into the
mitochondria or mitochondria has extra
protons itself?

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tyersome 3 years ago

more

Remember that all aqueous

solu ons contain a small amount of
hydronium (H₃O⁺) and hydroxide
(OH¯) due to autoioniza on.

This means that processes in cells

can use water to get rid of or grab
"protons" (H⁺) as needed.

This could have a small local effect

on the pH, but cells are well
buffered. Also, in the end the
various processes that create and
use H⁺ end up balancing each other
out.

The only situa ons where we really

need to keep an account of H⁺ are
processes involving membranes
(e.g. oxida ve phosphoryla on)
 where H⁺ gradients are created and
used.

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Herukm18 5 years ago

more

What does substrate level phosphoryla on

means?
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Ellie Bartle 3 years ago

more

Substrate level is the 'direct'

forma on of ATP in glycolysis and
the Krebs cycle, basically any ATP
not formed during the electron
transport chain.
(4
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Chaarvee Gulia 3 years ago

more

I don't quite understand why oxygen is

essen al in this process. I get that oxygen
serves as an electron acceptor at the end of
the electron transport chain, but why is
having this electron acceptor so important?
Why would ATP not be able to be produced
without this acceptor (oxygen)?
(4
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3 years
mo…
Satwik Pasani ago

It is sort of like a pipeline. If you

block the exit, the flow through the
en re pipeline stalls and nothing
moves. Oxygen is what allows the
 chain to con nue and keep
2
producing ATP.
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richie56rich 5 years ago

more

How much H2O is produced is the electron

transport chain?
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2
…
years
Ivana - Science trainee ago

C6H12O6 + 6O2 → 6CO2 + 6H2

6 molecules of water :D

This is the overall formula for cell

respira on, however, you ask how
many water molecules are
produced specifically during
electron transport chain?

Knowing that in the end, you get 6

molecules of water from one
glucose molecule.
In the electron transport chain,
hydrogen atoms are being added to
available oxygen in order to
produce water.

So you are confused about 6Co2,

too? well, you know that a er
glycolysis, you get 3C molecules
and bot undergoes Krebs cycle.
Now, what you get from one Krebs
cycle and proceeding ETC is
doubled.
 While it is true that via Krebs cycle
we get 2 CO2 (giving 4 in total), do
not forget decarboxyla on prior to
entering Krebs cycle (in between
Glycolysis and Krebs cycle) there
are is 1CO2. 1x2 = 2, 2+4 = 6

only 2NADH are reduced during

ETC, giving a total number of 2, and
also one FADH2 giving 1. It is 3x2
=6

as for Oxygen, it comes from the

red blood cells which carry oxygen
to absolutely every cell. Once in
the cell, it enters mitochondria and
acts as final acceptor of electrons. if
not, water would not be produced.

h p://www.dbriers.com/tutorials/2
012/04/the-electron-transport-
chain-simplified/
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yejikwon00 3 years ago

more

Where did all the hydrogen ions come from?

If NADH becomes NAD+, it releases H+ and
if FADH2 becomes FAD and would release
2H+. So are the hydrogen ions released by
those electron carriers are going to be used
for the gradient and also for the water
forma on? Or are the Hydrogen ions that
just came back through the ATP synthase
going to be used for forming H2O??
1 (4
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tyersome 3 years ago

more

The individual reac ons can't know

where a par cular "proton" came
from.
 Remember that all aqueous
solu ons contain a small amount of
hydronium (H₃O⁺) and hydroxide
(OH¯) due to autoioniza on.

This means that processes in cells

can use water to get rid of or grab
"protons" (H⁺) as needed.

This does have a small local effect

on the pH, but forming that NADH
or FADH₂ used protons so it all
balances out in the end.
(6
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Dallas Huggins 5 years ago

more

The new Campbell Biology textbook updated

the ATP yield totals to be 26-28 (instead of
30-32). What does this mean for your table
on the 'breakdown of one molecule of
glucose'? Where did the net yield go down?
(5
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Peony 5 years ago more

well, seems like scien sts have

recently discovered that the old
ATP yield is not quite accurate, and
the most recent data shows that it
should be around 26-28
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SanteeAlexander 5 years ago

more

I thought it was 38 ATPs from the previous

videos
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 Richard Wu 4 years ago
more

Well, I should think it is normal

unless something is wrong with the
electron transport chain. Like the
ques ons above. Cyanide, and that
weight control pill all cause the
normal respira on to func on
abnormally.
(0
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Abdul Mannan 2 years ago

more

How much electron NADH & FADH2

donates?
(2
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2
…
years
Ivana - Science trainee ago

FADH2: 1proton and 1 electron

NADH: 1proton and 1 electron
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Oxida ve phosphoryla on and the electron transport

chain
Fermenta on and anaerobic respira on

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