1. The respiratory chain and Oxidative phosphorylationBy
Dr. Manal Louis
Assistant professor
of Biochemistry

2. By the end of this lecture, the student should be able to:
1. Explain the chemiosmotic theory. 2. Determine the inhibitors of the resp. chain.

3. Electron Transport Chain

4. How is oxidation coupled to production of ATP?
Chemiosmotic Model
How is oxidation coupled to production of ATP?
Peter Mitchell Proposed chemiosmotic hypothesis

5. Chemiosmotic Model The theory postulates that the energy
obtained from oxidation is coupled to the
translocation of protons H+ from the
mitochondrial matrix to the inter-
mitochondrial space.
Complexes I, III, IV act as proton pumps.

6. So an electrochemical gradient is created across the inner mitochondrial membrane. (outside is +ve, inside is negative)

7. F0 F1
Scattered on the surface of the inner mitochondrial membrane are units of the ATP-synthase.

8. Step wise production of Energy in the form of ATP is a physiological behavior

9. Chemiosmotic Model
Several findings support the chemiosmotic theory:
Addition of protons (acid) to the external medium of mitochondria stimulates ATP production.
Oxidative Phosphorylation does not occur in case of solubilizing mitochondrial membranes.
Uncouplers.

10. P:O Ratio If we start from NADH = 3:1 If we start from FADH2 = 2:1
No. of ATP molecules produced
No. of Oxygen atoms consumed
If we start from NADH = 3:1
If we start from FADH2 = 2:1
Uncoupler = zero

11. Respiratory control
Mainly by availability of ADP. (ATP/ADP transporters may be rate limiting at certain times).
Availability of electrons: NADH/NAD ratio or FADH2/FAD.
3. Availability of O2.

12. Inhibitors of the respiratory chain
These compounds abolished the coupling between oxidation and phosphorylation through increasing the permeability of the IMM
Failure of formation of the electrochemical gradient
ATP formation stops while oxidation proceeds.
A- Uncouplers

13. What is the mechanism of uncouplers?
They act by transporting H+ to inside the mitochondria without passing through F₀F1
And energy is released as heat.
What is the mechanism of uncouplers?

14. Some endogenous compounds when increased: Bilirubin and Thyroxine.
A- Uncouplers
2,4 dinitrophenol
Dinitrocresol
Snake venoms phospholipases
Some endogenous compounds when increased: Bilirubin and Thyroxine.

15. 5-Thermogenine It is considered as a physiological uncoupling protein.
It is present in the brown adipose tissue of newly born, some people and hibernating animals.
It allows protons to pass the mitochondrial matrix without passing F0-F1 complex.
No ATP is formed and energy is released in the form of heat.

16. Inhibitors of the respiratory chain
B- inhibitors of ETC proper:
These inhibitors act on the three sites that produce the electrochemical gradient:
Complex I is inhibited by barbiturates and the insecticide rotenone.
Complex III is inhibited by the antibiotic, antimycin A.
Complex IV is inhibited by H2S, CO and cyanide
( ATP synthesis especially in the CNS causing Death).

17. Inhibitors of the respiratory chain
C-Inhibitors of phosphorylation:
Completely blocks F₀ so it inhibits ATP synthesis
Example: oligomycin (antibiotic)
D- ATP/ADP transporters inhibitors
This is achieved by the compound atractyloside

18. Derived from mother’s ovumA
Mitochondrial DNA
Circular DNA
Derived from mother’s ovumA
It contains 37 genes
13 proteins of ETC
22 tRNA
2 rRNA

19. Mitochondrial abnormalities
Mutations in mitochondrial DNA
Mitochondrial Myopathies
It usually occurs in mitochondria of muscles resulting in myopathies characterized by muscular weakness, cramping and wasting.
Inborn errors of mitochondrial DNA is only inherited from mothers.

20. MCQ Electron transport and phosphorylation
can be uncoupled by compounds that
increase the permeability of the inner
mitochondrial membrane to
(A) Electrons
(B) Protons
(C) Uncouplers
(D) O2
(E) ATP

21. An uncoupler of oxidative phosphorylation
such as dinitrophenol
(A) Inhibits electron transport and ATP synthesis.
(B) Allow electron transport to proceed without
ATP synthesis.
(C) Inhibits electron transport without impairment of ATP synthesis.
(D) Specially inhibits cytochrome b.
(E) Specially inhibits cytochrome c.

22. Reduced NADH generated in the cytoplasm is able to pass its electrons to oxygen through the mitochondrial electron transport system by which of the following mechanisms?
A) A specific transport system enables NADH to pass through the mitochondrial membrane. B) NADH reduces FAD to FADH2, which then passes through the mitochondrial membrane. C) NADH reduces dihydroxyacetone phosphate to glycerol phosphate, which can pass through the mitochondrial membrane. D) NADH reduces membrane-bound NADP to NADPH, which passes electrons to coenzyme Q. E) NADH oxidizes aspartate to oxaloacetate, which can enter the mitochondria.

23. Which of the following statements about the electron transport chain is correct?
a) The electron transport chain is made up of a chain of electron carriers with decreasing electron affinity. b) The electron transport chain is made up of a chain of electron carriers with increasing redox potential. c) The electron transport chain is made up of a chain of electron carriers with decreasing oxidising power. d) The electrons transferred from carrier to carrier in the electron transport chain gain heat.

24. How does atractyloside affect mitochondrial respiration? ...
If you isolate mitochondria and place them in buffer with a low pH, they begin to manufacture ATP. Why?

25. Loss of electrons from one substance is called:
a-Oxidation
b-Reduction
c-Dehydrogenation
d-Phosphorylation
e-Condensation

26. ATP- Synthase (Which is a membrane protein complex) is powered by: a- The flow of hydrogen ions b- ATP c- ADP d- Pi E- O2

27. Which of the following is oxidized by the respiratory chain?
a. NADPH
b. Acetyl CoA.
c. Oxygen.
d. NAD+.
e. NADH

28. Coenzyme Q is correctly described by all except:
Reduced by complex I & II
Transfers electrons to complex III
Is lipid soluble
Carry proton & electron
Accepts reducing equivalent from cytochrome aa3

29. The rate of Respiratory chain is mainly controlled by the concentration of:
a. ADP
b. NAD+
c. Inorganic phosphate.
d. NADH.
e. FADH2

30. When dinitrophenol is added to mitochondria, it:
decreases the flow of electrons through
cytochrome oxidase.
B. results in an increase in the intra-mitochondrial
ratio of ATP/ADP.
C. uncouples oxidative phosphorylation.
D. increases the rate of ATP formation.

31. Cyanide toxicity result from inhibition of:
NADH: Q reductase
Cytochrome oxidase
Cytochrome reductase
PDH
Malate dehydrogenase