Biochemistry- Oxidative phosphorylation

Question 1

What is the electron transfer potential of a compound?

Answer 1

This measures the energy present in the electrons carried, and can be measured by the standard redox potential, which is a measure of how readily a compound donates an electron in comparison with H2.

Question 2

What is the electron transfer potential of NADH and FADH2 converted into in oxidative phosphorylation?

Answer 2

The phosphoryl transfer potential of ATP.

Question 3

What does a negative standard redox potential mean?

Answer 3

This means that the reduced form of the compound has a lower affinity for electrons than H2 (and is therefore more likely to give them up). A positive redox potential means the opposite.

Question 4

What can be calculated if we know the redox potential for substrates in a redox reaction?

Answer 4

The standard free energy change for the reaction.

Question 5

What are standard redox potentials measured in?

Answer 5

Volts

Question 6

What has the lowest redox potential?

Answer 6

Oxygen

Question 7

What is oxidative phosphorylation? Give a brief overview.

Answer 7

It is the coupling of respiration to ATP synthesis.
It consists of two stages: electron transport and ATP synthesis.
Electrons from NADH and FADH2 flow down a chain of molecules (a respiratory chain) to oxygen. During the flow the energy within the electrons is used to pump hydrogen ions out of the mitochondrial matrix into the intermembrane space.
The gradient of protons is used to drive ATPsynthase, which uses the energy stored in the gradient to synthesise ATP.

Question 8

How many complexes are involved in the electron transport chain?
Where are they located?

Answer 8

4 multi-subunit complexes.

They are located on the inner mitochondrial membrane

Question 9

What happens to electrons in the electron transport chain?

Answer 9

They are handed from higher to lower redox potentials Eventually are passed on to oxygen which is the terminal oxygen acceptor, to form water.

Question 10

Where do electrons derived from NADH and FADH2 enter the chain?
What happens to them from there?

Answer 10

Electrons from NADH enter the chain at complex 1 and those from FADH2 enter the chain at complex 2. The electrons are then transferred to an intermediate electron transporter called coenzyme Q or ubiquinone. This transfers them onto complex 3, which then transfers them onto cytochrome c, which then transfers them onto complex 4, which then transfers them onto oxygen to form water.

Question 11

Is ubiquinone hydrophobic or hydrophilic? What does this allow it to do?

Answer 11

It is hydrophobic. This allows it to shuttle quickly in the inner mitochondrial membrane.

Question 12

Where is cytochrome c located?

Answer 12

In the intermembrane space.

Question 13

What do cytochromes contain which allows them to pick up electrons?

Answer 13

They contain a haem group which contains an Fe2+ ion. This can pick up electrons.
The haem group is used to pick up electrons, rather than oxygen, as it does in haemoglobin.

Question 14

Which of the electron transfer chain complexes can pump hydrogen ions from the mitochondrial matrix to the intermembrane space?
How is it possible for them to do this?

Answer 14

1, 3 and 4.
This is possible because when the electrons are transferred onto them, they drop from a higher to a lower redox potential. This releases energy which can be used to pump protons across the inner membrane.

Question 15

What is complex 2?

Answer 15

It is the enzyme from the TCA cycle which is located on the inner mitochondrial membrane and uses FAD as a cofactor. It reduces FAD to FADH2, and these electrons are then transferred to complex 2.

Question 16

What is the effect of the complexes pumping hydrogen ions into the intermembrane space?

Answer 16

An electrochemical gradient of hydrogen ions is built up. As a result, the hydrogen ions want to flow back into the matrix, but they cannot simply diffuse. Therefore the energy which was previously in the electrons is stored as a potential energy in this electrochemical gradient. Protons can flow back through the ATPsynthase, and this flow is coupled to ATP synthesis, as the ATP synthase uses the energy from the flow of protons to phosphorylate ADP to ATP.

Question 17

Give two other names for ATPsynthase?

Answer 17

mitochondrial ATPase

F1F0ATPase

Question 18

Describe the structure of ATPsynthase?

Answer 18

It has an F1 subunit, which is the head subunit and is located in the mitochondrial matrix.
The F0 subunit is hydrophobic and is located in the inner membrane of the mitochondria. This subunit contains a proton channel.
Together, these two subunits form a motor.

Question 19

Which subunits in the ATPsynthase form the static part of the rotor?
What is this called?

Answer 19

a, b, alpha, beta and delta subunits form the static part of the rotor, called the stator.

Question 20

Which subunits in the ATPsynthase can move?

Answer 20

c, gamma and epsilon- they can rotate.

Question 21

What happens to the ATPsynthase when protons flow through it?

Answer 21

This causes the rotor to move. the movement of the rotor against the alpha and beta subunits causes a change in conformation of the head group, and it is this change in conformation which can be exploited by the enzyme to synthesise ATP.

Question 22

Which molecules inhibit the transfer of electrons from complex four to oxygen?
What is the effect of this?

Answer 22

cyanide, azide and CO.

The electron transfer chain becomes clogged up with electrons, so ATP cannot be synthesised.

Question 23

Name a molecules which uncouples electron transport from phosphorylation.
How does it do this?

Answer 23

DNP

It forms a proton channel which disrupts the proton gradient, and stops the synthesis of ATP.

Question 24

Name a natural mechanism which uncouples electron transfer from phosphorylation?
What does this do?

Answer 24

Non-shivering thermogenesis.
This maintains body temperature in hibernating animals, newborn animals (including humans) and cold-adapted mammals.
Brown adipose tissue (brown because of the presence of lots of mitochondria) contains uncoupling protein (UCP), also known as thermogenin.
This creates a channel for protons to move back into the matrix from the intermembrane space, bypassing the ATPsynthase. Protons are still pumped into the intermembrane space, but some move back into the matrix via thermogenin.
As a result, less ATP can be synthesised, and there is a diffusion of energy present in the proton gradient, which manifests as heat.

Question 25

What is the PO ratio?

Answer 25

P stands for phosphate
O stands for oxygen
It is a measurement of the coupling of ATP synthesis to electron transport.
i.e. the number of molecules of inorganic phosphate incorporated into ATP, per atom of oxygen used.

Question 26

What does the PO ration depend on?

Answer 26

The substrate which is oxidised (NADH or FADH2).

Question 27

Why does the PO ratio differ for NADH and FADH2?

Answer 27

Because the electrons derived from NADH drives 3 proton pumps (1,3 and 4) while the electrons derived from FADH2 only drive 2 proton pumps (3 and 4). Therefore FADH2 contributes less to ATP synthesis.

Question 28

How are the electrons which are passed on to NADH in glycolysis transferred into the matrix of the mitochondria?

Answer 28

Via the glycerol-3-phosphate and the malate shuttles.
The NADH is not transported - just the electrons.
The pools of NADH in the cytoplasm and in the matrix of the mitochondria stay separate.

Question 29

Name two factors which the total yield of ATP from one glucose molecule depends on?

Answer 29

The precise values for the P/O ratio

The shuttle used to transport the electrons from glycolysis into the mitochondrial matrix

Question 30

Show how you would calculate the total yield of ATP from 1 glucose molecule, assuming the P/O ration for NADH is 2.5, and for FADH2 is 1.5.

Answer 30

Glycolysis: 2 ATP
TCA cycle: 2 GTP = 2 ATP
Glycolysis, PDC, TCA cycle = 10 x NADH + H+ = 25 ATP
TCA cycle = 2FADH2 = 3ATP

Therefore one glucose molecule yields 30-32 molecules of ATP, depending on what shuttle is used.

Question 31

What are OXPHOS diseases?

Answer 31

Diseases which involve components of oxidative phosphorylation.

Question 32

What causes OXPHOS diseases?

Answer 32

Mutations in mitochondrial or nuclear genes required for normal oxidative phosphorylation.

Question 33

Why may OXPHOS diseases may not be diagnosed until later in life?

Answer 33

Initially, a small number of normal mitochondria may provide enough ATP.
Spontaneous mutations can accumulate with age, and at some point, not enough ATP can be generated.

Question 34

Where do symptoms of OXPHOS diseases usually occur?

Answer 34

In tissues which have the highest energy demand, e.g. nervous system, heart, skeletal muscle, kidney