Download Electron Transport Chain, Oxidative phosphorylation and Pentose

Answer keys: 59-362 Quiz
Pentose Phosphate pathway:
1. What are the end products of pentose phosphate pathway?
NADPH, CO2 and Fructose 6 phosphate and Glyceraldehyde 3 phosphate
2. What is the need for pentose phosphate pathway?
For producing NADPH the reducing currency and for the production of Ribose
5 phosphate for the synthesis of nucleic acids.
3. People with defective GPDH enzyme are resistant to malaria. Why?
They have higher reactive oxygen species (ROS) in the system (due to low production
of NADPH). Malaria parasite is very sensitive to ROS, therefore it does not survive
in these people.
4. What enzymes are responsible for the production of NADPH and pentose
phosphate?
Glucose 6 phosphate dehydrogenase
6-phosphogluconolactonase
6-phosphogluconate dehydrogenase
5. What are the other enzymes involved in pentose phosphate pathwaway? Why are
they needed in this pathway?
Isomerases & epimerases
Trans-aldolases and Trans-ketolases
These enzymes are needed to convert pentoses (produced in the oxidative step) back
to hexoses and glyceraldehyde 3 Phosphate which could to used for next cycle of
pentose phosphate pathway or for glycolysis.
6. Which of the following is a correct statement?
a.
b.
c.
d.
Pentose phosphate pathway can be directly linked to CAC.
Pentose phosphate pathway can be directly linked to glycolysis. X
Pentose phosphate pathway can be directly linked to glycogen metabolism.
None of the above is correct statement.
7. What are the substrates and end products of pentose phosphate pathway?
Substrates: Glucose 6 phosphate, NADP
End Products: NADPH, CO2, Glucose 6 Phosphate, and Glyceraldehyde 3
phosphate
Electron Transport Chain, Oxidative phosphorylation and Pentose phosphate
pathway
1. Write down the different forms of energy at various steps in the oxidative
phosphorylation i.e. NADH oxidation coupled to ATP generation.
Chemical energy –to-- electrochemical gradient (electrical energy) –to-- kinetic energy
(rotation of the shaft) –to-- chemical energy (ATP)
2. How many Co-enzyme Q10 molecules will be needed to oxidize one molecule of
NADH, or one molecule of FADH2.
One molecule of CoQ10
3. Fe++/Fe+++ plays major role in the transfer electron s from one molecule to other
during mitochondrial ETC and many complexes and proteins have either Fe-S centres
or heme rings, but Fe++ in each protein has different reduction potential. Why?
Because of different electronic environments by surrounding amino acids around the F++
atom.
4. How many cytochrome c molecules are reduced during transfer of electron from one
Ubiquinone (QH2) in complex III?
One cyt. C at a time but eventually two cyt. C by one QH2
5. Reduction of coenzyme Q takes place in
a) Complex I b) Complex II c) Complex III d) in all of above X
6. Rotenone and antimycin A inhibit NADH dehydrogenase (complex I) and Ubiqinone
oxidase (complex III) respectively. Which one is more potent and dangerous poison?
Antimycin A
7. If isolated intact mitochondria are placed in solution containing rotenone, antimycin
A, ADP, Pi, at pH 5.0. Will there be
i.
synthesis of ATP
Yes
ii.
Oxygen consumption
No
iii.
Reduction of ubiquinone
No
iv.
Reduction of Oxygen
No
8. Comparing the F0-F1 ATP synthase to an electrical generator in a hydroelectric plant,
make the right connection in the followings,
Hydroelectric plant
Mitochondrial energy plant
Water
ATP
Turbine
H+
Dynamo shaft
subunits of F0
Electricity
 sub-unit of F1
10. What are the two components of proton motive force (the force which drags proton
across inner mitochondrial membrane generating ATP) in mitochondria?
Chemical gradient (H+ concentration)
Electrical gradient (more +ive charges outside than inside)
11. How can you prove the answer in above question?
I.
II.
One can inject K+ or other cataion inside the mitochondria to equalize the
+ive charges on both side of mitochondria (thus no voltage across the
membrane). In this scenario only chemical gradient will be working leading to
drastically decreased amount of ATP generation.
One can inject H+ inside to equalize the proton concentration, and increase
the K+ out side mitochondria. In this scenario there will be electrical gradient
(voltage due to high + ive charges outside). There will be little or no ATP
generation, because ATP synthase is very specific to protons only ant it allows
only H+ to pass through F0 to make ATP.
12. How does the purification F1 and F0 differ?
F1 is easy to purify as it is loosely and peripherally bound to inner mitochondrial
membrane through subunit.
It is difficult to purify
F0 is intra-membrane protein tightly associated to the
mitochondrial membrane. One has to use has to use detergent for solubilizing F0.
13. Thermogenin generates heat by
i.
Exergonic reactions of glycolysis
ii.
Oxidizing NADH
iii.
Hydrolyzing ATP
iv. Uncoupling ET and oxidative phosphorylation X
14. Why should an ionophore (a protein that allows diffusion of cations through it)
block ATP generation?
Because it dissipates (kills) the Electrochemical gradient, thus there is no force to puch
protons through F0-F1 , thus no ATP generation.
15. What are the three different conformational stages of the alpha-beta unit of F1
O: Open state, L: Lose state and T: Tight state
16. Theory of energy oupling of ATP synthesis to NADH oxidation reactions (like
substrate level phosphorylation) generating high energy intermediates was ruled out.
What was the evidence against this theory?
1. There were no high energy intermediates detected during the oxidation reaction on
ETC complexes.
2. If mitochondrial members were permeabilized, there was no effect on Oxidative
reactions but ATP generation was stopped (i.e. oxidation and phosphorylation were
uncoupled).
17. The Boyer’s theory of conformational change following reduction of ETC
complexes leading to ATP formation was also ruled out. Why?
1. There was no generation of ATP on each of the electron transfer complexes as
oxidation was occurring.
2. If mitochondrial members were permeabilized, there was no effect on Oxidative
reactions but ATP generation was stopped (i.e. oxidation and phosphorylation were
uncoupled).
18. Boyer’s hypothesis was partly true and it is applied in oxidative phophorylation.
What are the steps where it is applied correctly?
Indeed, Boyer’s theory of conformational change was true for explaining Proton
pumping stage (Complex I, III and IV) and also for ATP synthesis by F1.
19. How many ATP are synthesized from one NADH molecule produced in cytosol in
brain and skeletal muscle?
Theoretically: 2, actually: 1.5
20. Transfer of electrons from NADH to Oxygen leads pumping of 10 protons out of
inner mitochondrial membrane. This energy is sufficient to derive synthesis of 3 ATP
molecules. But actually only about 2.5 ATP molecules are generated, why?
Following are the reasons for this observation.
a. Part of the electrochemical gradient generated by Electron transport chain complexes
is used for transporting metabolites in and out of mitochondria.
b. A small amount of H+ leak back into mitochondria.
21. How many moles of ATP will be generated from one mole of sucrose in the subject
taking DNP as diet pill?
8 ATP
DNP (dinitrophenol) acts as an un-coupler, destroys the H+ gradient, therefore, ATP
generation by Oxidative phosphoylation is stopped.
22. Glycolysis can be carried out anaerobically as none of the reactions of this pathway
require oxygen. None of the reactions in citric acid cycle require oxygen. Can citric acid
cycle proceed under anaerobic condition? Explain.
If there is no oxygen, electron transport chain will not work, NADH produced in CAC
cannot be oxidized to NAD+. Also there will be an increase in the concentration of
NADH that inhibits CAC enzymes.
23. How many grams of glucose will be catabolized during 100 meter climbing for a
person weighing 50 Kg?
Assume that
free energy change of ATP hydrolysis is
-50KJ/mole,
no loss of energy any where,
Maximum ATP generation from glucose possible theoretically.
Please solve this problem at home and check with Dr. Pandey during office hour.
24. List the three experiments that show that oxidation reactions in the intact
mitochondrial membrane are coupled to the phosphorylation of ADP (leading to the ATP
generation).
1. Oxygen consumption (oxidation) and ATP generation (phosphorylation) can be
monitored in an in vitro reaction by adding the appropriate substrates in reaction
mixture containing isolated mitochondria. If you add only NADH or succinate
(substrates for oxidation reaction), there is no consumption of oxygen unless you add
ADP and Pi (substrates for phophorylation reaction) indicating that these two reactions
are coupled.
2. If you block the oxidation reaction only by adding rotenone or antimycin A to the
reaction mixture, ATP generation (Phosphorylation reaction) is also blocked indicating
these two reactions are coupled.
3. If you add blockers of ATP synthase (Oligomycin or FCCP), the oxygen consumption
is also stopped indicating these two reactions are coupled.
25. If the yeast is grown in aerobic condition during beer brewing in presence of
a) rotenone
Some what increased amount of alcohol and lesser CO2
b) antimycin A
High alcohol and less CO2
c) KCN
High alcohol and less CO2
d) DNP
No alcohol and more CO2
What will be the status of CO2 and alcohol content in the beer in each condition?
Answers are given in front of the conditions above.
26. What will the effect of oligomycin on Oxygen consumption in a reaction mixture
containing isolated mitochondria?
Oligomycing inhibit ATP synthase, this means, H+ gradient is not utilized, and the
electrochemical gradient remains high, this leads to inhibition of complex I.III and IV
(no need to pump H+), therefore, oxygen consumption will be inhibited.
27. What is so special about the surface of alfa-beta dimmer subunit of F1 of ATP
synthase?
G of ATP synthesis/hydrolysis is close to 0 on the surface of F1.