lecture 27 - redox reactions in the mitochondrion

Question 1

How much NADH is produced in respiration?

Answer 1

For each mol of glucose,
• 2 mol of NADH are produced in glycolysis (cytosolic)
• 2 mol of NADH from oxidation of pyruvate (mitochondrial)
• 6 mol of NADH in the citric acid cycle (mitochondrial).
10 NADH

Question 2

What is the electron transport chain?

Answer 2

Protein complexes in inner mitochondrial membrane

4 Complexes, of which 3 are on NADH O2
pathway.
Complex II: Succinate dehydrogenase.

Question 3

How is NAD+ reduced to NADH?

Answer 3

endergonic process
E’
o = -0.32 V

Question 4

Remember to look over A level stuff with direction of redox reactions and all that xoxoxo

Answer 4

ok thanks love from future me xoxoxo

Question 5

WHAT IS THE PATHWAY OF

ELECTRONS FROM NADH —> O2?

Answer 5

Just like Glycolysis, this is not a single-step reaction:
Multiple steps allow conservation of free energy (as
ATP) at discrete points in pathway.
This reaction sequence is accomplished by the
respiratory chain which is an e-transport pathway.
The enzymes that catalyse the reactions of the
respiratory chain are all membrane-bound: comprises
a large fraction of the proteins of the inner membrane.

Question 6

What is complex i?

Answer 6

Complex I (NADH – UQ oxidoreductase)
-Catalyses transfer of 2e
-
from NADH to ubiquinone

NADH oxidized initially by a flavoprotein, containing flavine mono-nucleotide (FMN) as a prosthetic group
-Electrons then passed to Fe/S centres (non-haem iron proteins)

Non-haem iron proteins (FeS):
Each Fe/S centre covalently linked to Cys
residues in protein

-arrangement of FeS clusters forms the ‘electron wire’
inhibited by rotenone/NO

Question 7

What is complex ii?

Answer 7

(Succinate dehydrogenase)
Catalyses oxidation of succinate and reduction of UQ
1 flavine adenine dinucleotide (FAD which is reduced to FADH2) covalently bound to protein
-3 Fe/S centres

Question 8

What is Ubiqinone?

Answer 8

(Coenzyme Q10, CoQ10)
A lipid-soluble component, present in great excess over other constituents of chain, freely diffuses in membrane
oxidised form quinone, reduced form quinol

Question 9

What is Complex III?

Answer 9

(UQ-Cyt c oxidoreductase)
2 b-type cytochromes (b566 (=bL) and b562 (=bH)
 1 Fe/S centre (Rieske protein)
 cytochrome c1
 bH inhibited by antimycin

Each cytochrome reduced by 1 e-
Cytochromes:a haem prosthetic group bound to a protein.
The Fe atom coordinated in the porphyrin ring is reduced: Fe3+ –> Fe2+

Question 10

What is Cytochrome c?

Answer 10

The only component of the respiratory chain which is
not an integral part of the membrane.
Nevertheless, it is bound loosely to the OUTER side
of the inner membrane.

Question 11

What is complex IV?

Answer 11

(cytochrome c oxidase)
 cytochromes a, a3
 2 copper atoms (CuA, CuB)
Accepts 4 e- from cyt c (=4 separate turnovers
of cyt c)
When fully reduced, can then reduce O2, together with 4H+ –> 2H2O
Oxygen reduction on matrix side of membrane
Inhibitors: CN, CO, azides (N3)

Question 12

How can we identify Which Complexes Actually ‘Make’

ATP?: Coupling Ratios?

Answer 12

Measure O2 consumption by mitochondria in a closed
chamber with O2 electrode and observe
Mitochondria in Pi buffer
-Background respiration observed with respiratory substrate
(NADH).
-Addition of ADP large increase in respiratory rate. When
ADP is all phosphorylated, rate returns to background rate.
i.e. The e-transport and phosphorylation reactions are
COUPLED.
-If a known amount of ADP is added, the amount of O2 used during phosphorylation can be measured to give:
ADP: O2 ratio (P:O ratio)
-P:O ratio is a measure of the number of ATP molecules
synthesized per pair of e- passing down the respiratory chain.

Question 13

Describe other methods to study mitochondrial

complexes: Physical separation?

Answer 13

Weak detergents disrupt lipid-lipid interactions
Purification of macromolecular components (protein complexes)
catalysing specific partial reactions in e- transport pathway
Reconstitute in lipid vesicles