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103 Cards in this Set

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1. Rank the following redox-active coenzymes and prosthetic groups of the electron-transport chain in order of increasing electron affinity: cytochrome a, CoQ, FAD, cytochrome c, NAD, FeS, Cu, cytochrome c1.
1. Cytochrome a @ Cu > cytochrome c1 @ cytochrome c > CoQ > FAD > NAD. Where are the FeS cluster(s) ranked? They are in the NADH-Q reductase, succinate Q and cytochrome reductases. They will assume an E˚’ close to what complex they are in. Their electron affinity or E˚’ is thus variable.
2. Why is the oxidation of succinate to fumarate only associated with the production of two ATPs during oxidative phosphorylation while the oxidation of malate to oxaloacetate is associated with the production of 3 ATPs?
The oxidation of succinate to fumarate catalyzed by succinate dehydrogenase reduces FAD to FADH2 . The FADH2 electrons enter the electron transport chain at CoQ bypassing the first oxidative phosphorylation (ATP forming) step, NADH -> CoQH2.Thus only 2 ATP forming steps in the electron chain from CoQ to O2 are utilized. In contrast the reaction malate to OAA produces NADH and therefore the electrons from NADH goes through the three ATP forming complexes in the electron transport chain.
3. What is the thermodynamic efficiency of oxidizing FADH2 so as to synthesize two ATPs under standard biochemical conditions?

E' of 1/2O2 + 2H+ 2e- ---> H2O = +0.816 V and
E' of FAD + 2 H+ + 2 e- ---> FADH2 = 0.0 V
∆E˚’ = +0.815V - (-0.0) = 0.815 V
∆G˚’ = -nF ∆E˚’ = -2 x 96.494 x 0.815 = - 157.3 kJ•mol-1
2Pi + 2ADP <===> 2ATP + 2 H2O
∆G˚’ = 2 x 30.5= +61 kcal •mol-1
Therefore, 61 / 157.3 = 0.388 or 38.8% efficiency.
4. Match the compound with it’s behavior: (1) rotenone, (2) dinitrophenol and (3) antimycin A.

(a) Inhibits oxidative phosphorylation when the substrate is pyruvate but not when the substrate is succinate.

(b) Inhibits oxidative phosphorylation when the substrate is either pyruvate or succinate.

(c) Allows pyruvate to be oxidized by mitochondria even in the absence of ADP.
1. answer is a. rotenone; rotenone inhibits complex I (pyruvate to Q) and not II (succinate to Q)

2. answer is c. dinitrophenol; uncouples oxidative phosphorylation from respiration. Thus pyruvate respiration goes on without ATP formation.

3. Answer is b. antimycin A; Antimycin A inhibits complex III, cytochrome c reductase.
5. How does atractyloside affect mitochondrial respiration?
Atractyloside inhibits the transport of ATP and ADP in the mitochondria by inhibiting the ATP/ADP translocase. Thus, it inhibits oxidative phosphorylation by decreasing the amount of ADP available for phosphorylation.
6. What change will occur when each of the following substances is added to mitochondria carrying out oxidative phosphorylation? (a ), antimycin a, (b), amytal, (c), 2,4-dinitrophenol, (d), succinate.
(a) antimycin a; inhibits complex III electron flow (CoQ -> Cyt c). Therefore NADH and FADH2 will accumulate while Cyt c and Cyt a3 oxidized forms will accumulate.

(b) amytal; inhibits complex I. Thus, NADH cannot be oxidized by CoQ and will accumulate.

(c) 2,4 Dinitrophenol; uncouples oxidative phosphorylation by dissipating the proton gradient across the inner mitochondrial membrane. O2 respiration increases without the production of ATP.

(d) succinate; it forms FADH2 which is oxidized by CoQ to form CoqH which is then oxidized by Cyt c which in turn is oxidized by O2. Two ATPs are formed per atom of Oxygen consumed.
7. What are the functions of the following components of the ATP synthase? F1, F0 and stalk proteins.
F1 is composed of 5 peptides, alpha3beta3,gamma,delta and epsilon.The alpha-beta subunits contain the ATP synthase active site. The gamma, epsilon peptides forms the gate coupling the stalk with F1 (see fig. 19-23f on page 711 in Lehninger or Fig. 11-39 in course pack (page 103) or in Lehninger, page 401). The Fo is composed of 4-5 peptides, is transmembrane and forms the proton channel. The stalk proteins,gamma and epsilon, interconnect F1 with F0. One protein binds oligomycin which blocks ATP synthesis by interfering with the utilization of the proton gradient.
9. Can ATP be produced by oxidative phosphorylation in the presence of antimycin? If so, what is the P:O ratio for the reduction of two-electron donors?
Yes. However you must use a reducing agent that will reduce cyt c (eg., ascorbate + TMPD). You will be able to use complex IV as the electron transport system and thus you will be able to synthesize one ATP per atom of O. thus P:O = 1.
10. Functional electron-transport systems can be reconstituted from purified respiratory electron transport chain components and membrane vesicles. For each of the following sets of components, determine the final electron acceptor. Assume O2 is present.
(a) NADH,Q, complexes I, III & IV
(b) NADH, Q, cytochrome c, Complexes II & III
(c) Succinate, Q, cytochrome c, Complexes II, III
& IV
(d) Succinate, Q, cytochrome c, Complexes II & III
(e) Succinate, Q, cytochrome c, Complexes I & III
10.
a) complex III. Cyt c is missing.

b) No reaction. Complex I is missing.

c) O2. System is complete.

d) Cyt c. Complex IV is missing.

e) No reaction. Complex II is missing.
Outline the mechanism for the reduction of O2 to H2O via cytochrome C oxidase. Describe the path of the electrons from the heme a-CuA cluster to the heme a3-CuB cluster and note the changes in the oxidation states of Fe and O. Note the formation of the superoxyl anion intermediate.
This answer is on page 702 (Fig. 19-14,also on page 99 in course pack). Reduced cytochrome c (ferrocytochrome c) transfers its electrons to the heme a- CuA cluster. The first electron goes to Cu++ then the next to Fe+++. Then the electrons from the heme a- CuA cluster are transferred to heme a3- CuB cluster. After reduction to the Cu+, Fe++ form in the heme a3- CuB cluster, O2 adds to Fe++ and the O2 of the complex then takes electrons from Cu and from Fe to form a Fe+++O2- -, Cu++ complex.This is known as a peroxy intermediate. Two protons and an electron are added and one oxygen atom is tranferred from the Fe to Cu. The electrons and protons combine with an Oxygen atom transferred to the Cu to give a Cu++-O-H2, Fe++++-O- - complex. An electron is added to the complex to form water with the oxygen associated with the the Cu which is oxidized to Cu++. Both waters are then released from the Fe and Cu with addition of another electron and two protons. The reduction of O2 is done this way by binding the partially oxidized intermediates tightly to both Fe and Cu in the heme a3- CuB cluster. Thus, no toxic partially reduced oxygen intermediates are released.
13. Calculate the Delta Go’ for the reaction, Succinate + FAD -------> fumarate + FADH2
Fumarate:succinate Eo’ =0.03
FAD:FADH2 Eo’ = 0.0

F = 96.494 kcal/V ; 1V = 1 Joule/Kcal
Delta Eo’ = E˚’ electron acceptor - E˚’ electron donor = 0 - (0.03) = - 0.03V

Therefore Delta Go’ = -nF Delta Eo’ = -2 x 96.494 kJ/V mol x (-0.03) = +5.79 kJ/mol.

The reaction is endergonic.
What is the standard Potential Electrode of the hydrogen redox pair at at pH 7.0 the emf of the electrode ?
the E’ 0 is -0.414 V.
The salt bridge in test cell, usually saturated
KCl, is the path for?
counter-ions.
The salt bridge in test cell is usually saturated?
KCl,
What is a standard of
reference that is assigned a standard reduction potential of 0.00 V.
H+ + e- ===> 1/2 H2
Reduction potential of a reaction also depends on?
the concentration of the reductant and
oxidant forms.

Thus the reaction of two reactants is,
An+ + B reduced <======> A reduced + B n+


Delta G = Delta G0’ = RT ln [A reduced ] [ Bn+ ] /[B reduced ] [ An+ ]


Now Delta G = -n F Delta E



Substitute Delta E for Delta G/- n ℑthen we have the Nernst Equation.

Delta E=

Delta E0’ – (RT/n F) ln ([A reduced ][ Bn+ ]/[B reduced ] [ An+ ])
The half reactions are written in this manner
A (n+) + n e- <==> A (red)

E = E0’ – (RT/n F) ln [A red]/[An+]

B (n+) + n e- <==> B (red)

E = E0’ – (RT/n F) ln [B (red)]/[B(n+)]In the reaction where A is the electron acceptor and B is the donor we have what rxn and what equation:
A (n+) + B (red) <==> A (red)+ B (n+)

and

Delta E0’ = E0’ (e- acceptor A) - E0’ (e- donor B)

If Delta E is positive then Delta G is negative and we have an _______ reaction.

Exergonic
Cellular oxidation of glucose to CO2 requires specialized electron carriers known as
Coenzymes.

Four examples are?
Pyridine nucleotides; Nicotinamide adenine dinucleotide (NAD+) and Nicotinamide
adenine dinucleotide 2’-phosphate (NADP)

Please note that reduction of NAD+ to NADH or NADP+ we have a two electron transfer
.
NAD(P)+ + 2 e- + 2 H+ ====>NAD(P)H + H+

NAD + NADH concentration is ~ 10-5 M and

cellular NAD/NADH ratio is >50 and

cellular NADPH/NADP ratio is > 50.
What redox carriers are tightly bound to proteins?
Flavin nucleotides
There are two flavins. They are?
FAD and FMN.
FAD and FMN are what type of electron carriers?
can be two electron carriers or one electron carriers as seen in their semi-quinone structure.
What is the Delta G0’ of the oxidation of NADH by O2 ?
NAD+ + H+ + 2 e- <=====> NADH E0’ = - 0.320 V

1/2 O2 + 2H+ + 2 e- <=====> H2O E0’ = + 0.816

Therefore the reaction is,

1/2 O2 + H+ + NADH <=====> H2O + NAD+
and Delta E0’ = 0.816 - (- 0.320) = 1.136 V

Delta G0’ = - nF Delta E0’ Thus, Delta G0’ = - 2 (96840) ( 1.136) = 220,020.48 J
Or 220.0 kJ.

For synthesis of ATP from ADP + Pi you need 30.5 kJ.

Therefore you could synthesize 220/30.5 or 7 ATPs from the oxidation of NADH by oxygen.

only 2.5 ATPs are synthesized when NADH is oxidized in the mitochondria by the electron transport chain with oxygen as the ultimate electron acceptor.
What is the sub-cellular organelle where the
electron transfer system that transports electrons and hydrogen from the NADH and FADH2 produced in the TCA cycle to O2 to form H2O.
The Mitochondria
What is in the mitochondrion?
The electron transfer (transport) system

The TCA cycle
The electron transfer (transport) system is a series of oxidative reactions through respiratory carriers in 4
different enzyme complexes known as?
complexes I-IV.
The electron transfer (transport) system is?
a series of oxidative reactions through respiratory carriers in 4 different enzyme complexes known as complexes I-IV.
3 parts of the anatomy of
Mitochondrion
Outer membrane:

Inner membrane:

Matrix
What is the Outer membrane of Mitochondrion permeability?
Freely permeable to small molecules and ions
What is the inner membrane of Mitochondrion permeability?
Impermeable to most small molecules and ions, including H+
What is needed to penetrate inner membrane of Mitochondrion?
Needs transporter
What does the inner membrane of Mitochondrion Contain?
Resp. electron carriers (complexes I-IV.)

ADP-ATP translocase

ATP Synthase (F0, F1)

Other membrane transporters
What does the Matrix of Mitochondrion Contain?
Pyruvate dehydrogenase complex

TCA enzymes

Fatty acid beta-oxidation enzymes

A.A oxidation enzymes

DNA, ribosomes

ATP, ADP, Pi, Mg2+, Ca2+. K+

Many soluble metabolic intermediates andother enzymes

The cristae are extensions of
the inner membrane.
The mitochondria has its own?
DNA
RNA
and can make proteins
The proteins made in the mitochondria (G/S/L) than
proteins made in the cytosol
Less than
What are the other electron acceptors in the electron transport chain?
FAD and FMN
FAD and FMN accept electrons and hydrogen from?
NADH as well as from succinate.
What electron acceptors in the electron transport chain are tightly bound to proteins and sometimes they may be covalently linked to the
Protein?
FAD and FMN
FAD and FMN in the electron transport chain are tightly bound to proteins and sometimes they may be covalently linked to?
The Protein
FAD and FMN in the electron transport chain are tightly bound to proteins and sometimes they may be covalently linked to the Protein. What happens then?
FADH2 then transfers its electrons to Ubiquinone or Coenzyme Q.
Unlike NAD+ reduction which is a two electron transfer, ubiquinone can
also undergo a one electron transfer where a semiquinone intermediate is formed.
Heme A with its isoprenoid tail is found in?
a-type cytochromes.
being mobile and freely diffusible in the lipid bilayer of the inner mitochondrial membrane, Ubiquinone can do what?
therefore transfer electrons from the soluble electron carriers to less mobile electron carriers in the membrane.
NAD+ reduction involves what type of transfer?
a two electron transfer
Ubiquinone involves what type of transfer?
undergo a one electron transfer where a semiquinone intermediate is formed or a two electron transfer
What is Ubiquinone?
a small molecule and is hydrophobic, being mobile and freely diffusible in the lipid bilayer of the inner mitochondrial membrane.

transfer electrons from the soluble electron carriers to less mobile electron carriers in the membrane.
Ubiquenone is involved in ______ and are reduced to _______?
in FAD and FMN electron acceptors

Ubiquinol
The 4 five-membered nitrogen Containing rings in cyclic structure are named?
porphyrins.
In porphyrins, the 4 nitrogen atoms are coordinated with a?
central Fe ion, either Fe2+ or Fe3+.
Iron Protoporphyrin IX is found in
b-type cytochromes
and in hemoglobin and Myoglobin.
What is found in b-type cytochromes and in hemoglobin and Myoglobin?
Iron Protoporphyrin IX
Heme C is covalently bound to cytochrome C protein through?
thioether bonds from Cys residues in the protein.
What is covalently bound to cytochrome C protein through thioether bonds from Cys residues in the protein?
Heme C
Heme A with its ________ is found in a-type cytochromes.
isoprenoid tail
What are the 3 types of cytochrome structures?
Heme A - isoprenoid
Heme B - hyrin
Heme C - Cys
Cytochromes are proteins with strong absorption of visible light because?
of their Fe-containing heme prosthetic groups.
All cytochromes have in common?
Same base structure
distinctive absorption spectra.
What cytochromes are tightly bound?
a and b cytochromes
What cytochromes are tight, covalently bound?
Heme C
Cytochrome a and b are integral proteins of?
the inner mitochondrial membrane
What cytochrome is soluble and associates with the membrane through electrostatic interactions with the outer surface of the inner membrane and why?
Mitochondrial Cytochrome c

Cause it’s has a tight, covalently bond
Mitochondrial Cytochrome C Facts
soluble

Cys side chain

associates w/ the membrane through electrostatic interactions with the outer surface of the inner membrane


Iron sulfur proteins. They are proteins in which one Fe atom is coordinated with inorganic sulfur atoms or with the sulfur atoms of Cys residues.
What are the three iron sulfur centers?
(a) a single Fe ion surrounded by S atoms of 4 Cys centers.

(b) as in (a) but two inorganic sulfur atoms are
between the Fe ions. This is known as a 2Fe-2S cluster or center.

(c) The middle is known as a 4Fe-4S center or cluster.
What is known as a 2Fe-2S cluster or center?
two inorganic sulfur atoms are between the Fe ions
Electrons pahway from NADH to O2?
from NADH or succinate through flavoproteins, ubiquinone, iron sulfur proteins,cytochromes and then to O2
Electrons move from NADH or succinate through flavoproteins, ubiquinone, iron sulfur proteins,
cytochromes and then to O2

How was this sequence determined?
Standard Reduction potential

Neg to positive
What are the 3 method used to determine the sequence of electron carriers?


standard reduction potentials of the individual electron carriers that have been determined

having the whole electron transport chain being reduced and oxidized

the use of inhibitors of the electron tranport chain.




First the standard reduction potentials of the individual electron carriers have been determined experimentally. It is expected that the carriers to function in order of
increasing reduction potential, from carriers of lower E’0 to carriers of higher E’0.
The order of 8 carriers based on increasing reduction potentials is:
NADH => Q => cytochrome b => cytochrome c1 => cytochrome c => cytochrome a => cytochrome a3 => O2.
What is the second method used to determine the sequence of electron carriers?
having the whole electron transport chain being reduced in the absence of
O2. Then introduce O2 in the system and see the sequence of oxidation of the
electron carriers (done spectrophotometrically).
How is the carrier getting oxidized first in the second method determining the sequence of electron carriers?
The carrier closest to O2 becomes oxidized first, the second closest oxidized next and so forth.
Explain the second method used to determine the sequence of electron carriers?
having the whole electron transport chain being reduced in the absence of
O2.

Then introduce O2 in the system and see the sequence of oxidation of the
electron carriers (done spectrophotometrically).
What is the Third method used to determine the sequence of electron carriers?
using inhibitors of the electron tranport chain.
Explain the third method used to determine the sequence of electron carriers?
In the presence of O2 electron donor carriers in the chain that function before the inhibited step becomes fully reduced. Those that function afterwards would remain oxidized.
In the Inhibitor method for determining sequence of electron carriers, what is oxidized and what is reduced?
Those before inhibition step is reduced and those after the
inhibition step remains
The electron carriers are in multienzyme complexes that are?
membrane-bound that can be physically separated.
The electron carriers are in?
multienzyme complexes
What are the multienzyme complexes that the electron carriers?
Complex I carries electrons from NADH to ubiquinone.

Complex II carries electrons from succinate to ubiquinone.

Complex III carries electrons from reduced ubiquinone to cytochrome c complex IV transfer electrons from cytochrome c to O2.
What does Complex II do?
Complex II carries electrons from succinate to ubiquinone.
What does Complex III do?
Complex III carries electrons from reduced ubiquinone to cytochrome c
What does Complex IV do?
complex IV transfer electrons from cytochrome c to O2.
What does the Complex I proton pump do?
transfers 4 protons in one direction, from the matrix to the inter- membrane space.
What happens when the Complex I proton pump transferring 4 protons in one direction, from the matrix to the inter- membrane space.?
The matrix becomes negative and the intermembrane space, positive.
Other than carrying electrons from NADH to ubiquinone, Complex I also acts as?
a proton pump
The rxn that takes place in Complex I?
NADH <~~~> ubiquinone oxidoreductase.
In complex I, Why does 4 protons from the matrix transfer in a vectorial manner to the intermembrane space?
due to electron transfer from NADH to Q to form QH2.
What happens in the complex I do during electron transfer from NADH to Q to form QH2?
4 protons from the matrix are transferred in a
vectorial manner to the intermembrane space
What is the site where is the site where 4
protons from the matrix are transferred in a
vectorial manner to the intermembrane
space due to electron transfer from NADH
to Q to form QH2?
matrix arm portion
What causes formation of a proton gradient in Complex I?
4 protons from the matrix are transferred in a vectorial manner to the intermembrane space
What is the Complex rxn?
NADH + H+ + Q →NAD+ + QH2 or

NADH + 5H+ N + Q →NAD+ + QH2 +4H+ P
The rxn that takes place in Complex II?
Succinate Dehydrogenase (SDH)
Succinate Dehydrogenase is the only TCA cycle enzyme that is?
membrane-bound, having 5 prosthetic groups and 4 different subunits.
In the structure of complex II, succinate dehydrogenase, Two subunits of SDH are located __________ while the 2 other subunits are?
the matrix of the mitochondria

membrane proteins.
What is contained in the complex II, succinate dehydrogenase?
The matrix proteins contain 3 2Fe-2S centers, FAD and a binding site for succinate.


The ultimate e- acceptor, ubiquinone, is bound to the membrane subunits.

A heme b molecule is present in the membrane subunits but is non-functional in the electron path.
The matrix proteins in the complex II,Contain?
3 ~~ 2Fe-2S centers, FAD and a binding site for succinate.
What is the ultimate e- acceptor, contained in the complex II and what is bound to?
ubiquinone, is bound to the membrane subunits.
A ____ molecule is present in the complex II
membrane subunits but is non-functional in the electron path.
heme b
The rxn that takes place in Complex III?
Ubiquinone:Cytochrome Oxidoreductase (also called cytochrome (bc1)Complex.
In Complex III, The reduced ubiquinone also known as ________ in this complex, transfers its electrons eventually to cytochrome c
Ubiquinol
In Complex III, The reduced ubiquinone also known as ubiquinol in this complex What does what to electrons?
transfers its electrons eventually to cytochrome c
How many structure does the complex III have?
(a) monomer structure
(b) dimeric unit
Electron transport in complex III is
known as?
the Q cycle.
Electron transport in complex III is known as the Q cycle and the equation
is?
QH2 + 2 Cyt c1 (ox.) + 2H+ N →Q + 2 Cyt c1 (red.) + 4H+P
What is Q cycle?
a process in the complex where a two e- carrier
becomes a single e- donor to cytochromes b562, b566, c1, and c.
What is the net result of the Q cycle?
one molecule of Ubiquinol is oxidized and 2 molecules of Cyt c are reduced and the reduced Cyt c moves to complex IV.