Mitochondrial respiratory chain

Question 1

Structure of mitochondria

Answer 1

Outer membrane (freely permeable to small molecules and ions)

Inner membrane (impermeable to small molecules and ions including H+)

Matrix

Cristae

Question 2

Structure of electron transport chain

Answer 2

Complex I (NADH-CoQ oxidoreductase)

Complex II (succinate dehydrogenase)

Complex III (cytochrome bc1 complex)

Complex IV (cytochrome x oxidase)

Question 3

Complex I

Answer 3

FMN becomes FMNH2 becomes QH2

NADH + H+ becomes NAD+

4H+ move in

Enzyme is NADH dehydrogenase

Question 4

NADH dehydrogenase

Answer 4

Initially electrons passed to FMN to produce FMNH2

Transfer to a series of iron sulphur clusters

Transfer to coenzyme Q or ubiquinone

Overall reaction is:
NADH + H+ + Q = NAD+ + QH2

It is a proton pump

Question 5

Complex II

Answer 5

Succinate dehydrogenase

Electrons of FADH2 pass on their electrons to complex II

Complex II passes them to ubiquinone

Other substrates for mitochondrial dehydrogenases also pass on their electrons but not through complex II

Question 6

Path of electrons entering the electron transfer chain

Answer 6

Electrons from NADH pass through a flavoprotein to a series of iron sulphur proteins in complex I then to Q

Electrons from succinate then pass through a flavoprotein and several Fe-S centres in complex II on the way to Q

G3P donates electrons to a flavoprotein on the outer face of the inner mitochondrial membrane where they pass to Q

Acyl-CoA dehydrogenase transfers electrons to electron transferring flavoprotein where they pass to Q via ETF

Question 7

Complex III

Answer 7

Ubiquinone: cytochrome c oxidoreductase

Second of three proton pumps in the respiratory chain

Question 8

Complex IV

Answer 8

Cytochrome oxidase

Third and final proton pump

Carries electrons from cytochrome c to molecular oxygen

Produces water

Question 9

Synthesis of ATP

Answer 9

Purpose of the whole process

Inner mitochondrial leaflet is generally impermeable to charged species but 3 specific systems in this membrane that:

1. Transport ADP and Pi into the matrix
2. Synthesise ATP
3. Transport ATP into the cytosol

Question 10

Synthesis of ATP equation

Answer 10

ADP3- + Pi2- + H+ = ATP4- + H2O

Actual substrates are the Mg2+ complexes of ADP and ATP

Question 11

Adenonsine nucleotide translocase

Answer 11

Integral protein of the inner mitochondrial membrane

Transport ADP3- from the intramitochondrial membrane space into the matrix

In exchange for an ATP4- molecule transported in the other direction

Known as an antiporter

Question 12

Phosphate translocase

Answer 12

A second membrane transport is essential for oxidative phosphorylation and synthesis of ATP

Transport both phosphate and hydrogen ions into the matrix: a symporter

Favoured by the transmembrane proton gradient

Question 13

ATP synthase

Answer 13

An F-type ATPase

Two functional domains:

1. F0, an oligomycin- sensitive proton channel
2. F1, an ATP synthase

Question 14

F0

Answer 14

Comprises three different types of subunit: a, b, c

Forms a complex of 13-15 subunits

Subunits c1-10 arranged in a circle

Question 15

F1

Answer 15

Comprises five different types of subunit: alpha3, beta3, gamma, delta, epsilon

Forms a complex of 9 subunits

The 3 beta subunits have catalytic sites for ATP synthesis

Question 16

Beta subunits

Answer 16

Are arranged alternately with alpha subunits like segments of an orange

Form a knob like structure held by the gamma and epsilon subunuts

Delta subunit interacts with the two b subunits of F0

Question 17

Theory of rotational catalysis

Answer 17

3 subunits take turns catalysing the synthesis of ATP

Any given beta subunit starts in conformation for binding ADP and Pi

Then changes conformation so the active site now binds the product ATP tightly

Then changes conformation to give the active site a very low affinity for ATP so ATP is released

Question 18

Summary of energy changes

Answer 18

Highly exergonic reaction

Energy released is coupled to the movement of H+ across the inner membrane

Electrochemical energy generated represents temporary conservation of the energy of electron transfer

Protons flow spontaneously down their electrochemical gradient releasing energy available to do work

Question 19

Energy yield from oxidation of 1 molecule of glucose

Answer 19

Glycolysis

• 2 NADH (3 or 5 ATP depending on shuttle used)
• 2 ATP

Pyruvate oxidation
- 2 NADH (5 ATP)

Acetyl-CoA oxidation in CAC

• 6 NADH (15 ATP)
• 2 FADH2 (3 ATP)
• 2 ATP or 2 GTP

Total yield of 30 or 32 ATP

Question 20

Uncoupling reagents

Answer 20

Normally e- flow and phosphorylation are tightly coupled

Uncouplers dissipate the pH gradient by transporting H+ back into the matrix of mitochondria bypassing the ATP synthase

Uncoupler severs the link between e- flow and ATP synthesis with energy being released as heat

Can occur naturally

Question 21

Uncoupling and brown fat thermogenesis

Answer 21

Brown adipose tissue:

• specialised for heat generation
• high numbers of mitochondria
• mitochondria contain thermogenin
• important in new borns, possible role in obesity/ diabetes

Question 22

DNP- an exogenous uncoupler

Answer 22

Weak acid that crosses membrane ‘ferrying’ H+ across

Each DNP molecule collects a proton from the IMS and moves through the membrane with it, depositing it in the matrix

Can then return through the membrane to collect another proton

Question 23

DNP effects

Answer 23

Leeds medical sudent took 2,4- DNP as weight loss aid

62 deaths due to DNP reported including body builders and slimmers

Toxicity arises from liver damage, respiratory acidosis and hyperthermia