Oxidative Phosphorylation & Electron Transport Flashcards
What does the term oxidation mean?
Oxidation refers to the process of LOSING an electron.
What does the term reduction refer to?
Reduction refers to the GAIN of electrons.
What are monooxygenases?
Monooxygenases are enzymes that add ONE oxygen atom of O2 to a molecule.
What are dioxygenases?
Dioxygenases are enzymes that add both atoms of O2 to a substrate.
This increases the solubility of nonpolar compounds.
What are oxidases?
Oxidases are enzymes that catalyze REDOX reactions involving O2 as the electron acceptor.
Oxygen will be reduced to either:
a) Superoxide (1 electron)
b) Hydrogen peroxide (2 electrons)
c) Water (4 electrons)
What does reduction potential (Eo) refer to?
Reduction potential (Eo) is the measure of the ease with which a compound can be oxidized or reduced.
The more positive the value of Eo, the more readily it accepts electrons.
The more negative the value of Eo, the more readily it donates / loses electrons.
What is the equation that relates delta Go’ to the Nernst equation and what is it used for?
delta Go’ = - nF delta Eo’
This equation is used to determine how much energy a molecule / system possesses based on its reduction potential.
n = # of electrons
F = 96.485 kj/V x mol
Which direction do electrons flow?
Electrons will flow to any half reaction that has a higher or less negative reduction potential.
True or False: Oxygen is an effective electron sink.
True: Oxygen has a very positive reduction potential and is therefore an effective electron acceptor.
What does the generation of ATP via oxidative phosphorylation require?
To generate ATP via oxidative phosphorylation (the electron transport chain), an electron donor and an electron acceptor is needed.
Components of the electron transport chain.
ATP synthase.
An intact, inner mitochondrial membrane is needed as well.
NADH and/or FAD [2H] are electron donors and O2 is the electron acceptor.
How is the electron transport chain organized?
There are four protein complexes on the INNER mitochondrial membrane.
Lipid soluble coenzymes (UQ and CoQ) and a water soluble protein (cytochrome c) shuttle between the complexes.
Electrons FALL in energy as they are passed through the chain from Complex I and II to Complex IV.
Picture of the Electron Transport Chain
Energy Blance of Body
What are the five(5) major protein components of the electron transport chain (oxidative phosphorylation)?
Complex I (NADH-CoQ Oxidoreductase)
Complex II (Succinate dehydrogenase)
Complex III (Cytochrome b-c1 complex)
Complex IV (Cytochrome oxidase)
ATP Synthase
What are the two coenzymes that are embeded in the inner mitochondrial membrane and what role do they play in the electron transport chain?
Coenzyme Q
Cytochrome C
What is the function of CoQ10 and what three states can it exist in?
CoQ10 functions as an electron carrier from enzyme complex I and enzyme complex II to complex III.
CoQ10 can exist in one of three oxidation states:
1) Fully oxidized to ubiquinone
2) Semiquinone
3) Fully reduced to ubiquinol
What is the function of cytochrome C?
The heme group of cytochrome c accepts electrons from the b-c1 complex and transfers electrons to the cytochrome oxidase complex.
Cytochrome c is also involved in initiation of apoptosis. Upon release of cytochrome c to the cytoplasm, the protein binds apoptotic protease activating factor-1 (Apaf-1).
What is the purpose of the electron transport chain?
The function of the electron transport chain is to produce a transmembrane mitochondrial proton electrochemical gradient as a result of the redox reactions.
If protons flow back through the mitochondrial membrane, they enable mechanical work.
Why is the electron transport chain also called oxidative phosphorylation?
The entire process is called oxidative phosphorylation, since ADP is phosphorylated to ATP using the energy of hydrogen oxidation in many steps.
Diagram of the overall pathway of the transfer of electrons within the electron transport chain.
**NADH** → ***Complex I*** → **Q** → ***Complex III*** → **cytochrome *c*** → ***Complex IV*** → **O<sub>2</sub>** ↑ ***Complex II*** ↑ ***FADH***
What is the function of Complex I (NADH dehydrogenase)?
Complex I oxidizes NADH and reduces CoQ. It transfers two electrons from NADH to CoQ.
Ubiquinone is reduced to ubiquinol and difuses across the membrane and transfers four protons across the membrane to produce a proton gradient.
Complex I also transports protons from the mitochondrial matrix to the cytosolic side of the inner mitochondrial membrane.
The cytosolic side is more positive than the matrix side.
How are electrons oxidized and what is their path at Complex I?
NADH is oxidized to NAD+, by reducing Flavin mononucleotide (FMN) to FMNH2 in one two-electron step.
FMNH2 is then oxidized in two one-electron steps, through a semiquinone intermediate. Each electron thus transfers from the FMNH2 to an Fe-S cluster, from the Fe-S cluster to ubiquinone (Q).
Transfer of the first electron results in the free-radical (semiquinone) form of Q, and transfer of the second electron reduces the semiquinone form to ubiquinol.
During this process, four protons are translocated from the mitochondrial matrix to the intermembrane space.
What is the purpose of succinate dehydrogenase (Complex II)?
Complex II oxidizes succinate and reduces CoQ.
When succinate is converted to fumarate in the TCA, there is a concomitant reduction of bound FAD to FADH2 in succinate dehydrogenase.
The FADH2 transfers its electrons to Fe-S centers which then pass them on to CoQ.
NOTE: It is the only TCA cycle enzyme that is an intergral membrane protein of the inner mitochondrial membrane.
NOTE II: The oxidation of one FADH2 in the electron transport chain results in the synthesis of two ATPs. The oxidation of one NADH results in the production of three ATPs.
What is unique about succinate dehydrogenase (Complex II)?
Succinate dehydrogenase is an enzyme complex, bound to the inner mitochondrial membrane.
It is the only enzyme that participates in both the citric acid cycle and the electron transport chain.
What does the Complex III (Cytochrome c reductase / cytochrome bc1) do?
Complex III mediates electron transport from CoQ to cytochrome c.
This cycle is known as the Q cycle.
Complex III also transports two protons across the inner mitochondrial membrane.
What does Complex IV (cytochrome c oxidase) do?
Complex IV transfers electrons from cytochrome c to reduce oxygen in the mitochondrial matrix.
Cytochrom c oxidase and oxygen are the final destination for the electrons in the ETC.
The combined processes of oxygen reduction and proton transport require eight protons. Four are used to reduce two molecules of oxygen (2 per cycle) and four are transported from the matrix to the intermembrane space (2 per cycle)
Proton transport in Complex IV takes place via two channels that are lined with water molecules and polar amino acids.
Picture of the electron transport chain.
Picture showing relationship of electron transport chain and TCA.
What does the term “coupling” refer to?
The electron transport chain and oxidative phosphorylation are coupled by a proton gradient across the inner mitochondrial membrane.
The efflux of protons from the mitochondrial matrix creates a proton gradient. This gradient is used by the ATP synthase complex to make ATP via oxidative phosphorylation.
The FO component of ATP synthase acts as an ton channelthat provides for a proton flux back into the mitochondrial matrix. This reflux releases free energy produced during the generation of the oxidized forms of the electron carriers (NAD+ and Q). The free energy is used to drive ATP synthesis, catalyzed by the F1 component of the complex.<i></i>
Coupling with oxidative phosphorylation is a key step for ATP production. However, in specific cases, uncoupling the two processes may be biologically useful. The uncoupling protein, thermogenin—present in the inner mitochondrial membrane of brown fat—provides for an alternative flow of protons back to the inner mitochondrial matrix. This alternative flow results in thermogenesis rather than ATP production.<i> </i>
<i></i>Synthetic uncouplers (e.g., 2,4-dinitrophenol) also exist, and, at high doses, are lethal.<i></i>
What is the function of cytopchrome C?
Cytochrome C is a mobile electron carrier.
Electrons are passed from Complex III to cytochrome c.
It is associated with the inner mitochondrial membrane where it carries electrons in its reduced state to to cytochrome c oxidase.
What is ATP synthase and what does it do?
ATP synthase is a combined enzyme, proton pump, and rotating molecular motor.
It is made up of the F1 subunit and the F0 subunit.
What is the F1 subunit of the ATP synthase and what does it do?
The F1 subunit of the ATP synthase is the catalytic subunit of the ATP synthase; it catalyzes ATP production.
The F1 subunit is made up of five modules: Alpha, beta, gamma, delta, and epsilon.
The F1 portion of the ATP synthase is above the membrane, inside the matrix of the mitochondria.
The alpha and beta subunits are arranged in alternating patterns in the hexamer and are similar but not identical.
There are six ATP-binding sites on this subunit hexamer. Three of these, located on the beta subunit, ar ethe catalytic sites. The other three, located on the alpha subunit, are non-catalytic.
The release of newly synthesized ATP is what powers the subunits catalytic activity.
What is the F0 subunit and what is its funtion within the ATP synthase?
The F0 subunit serves as the attachment point for the F1 subunit.
It is made up three hydrophobic subunits (A, B, and C) and six additional subunits, d, e, f, g, F6, and 8.
The a and b subunits form part of the stator. This is a stalk-like structore that is anchored into the membrane and connects to the F1 subunit. There is also a ring composed of 1-15 subunits; this makes up the rotor of the motor.
Protons flowing through the a-c complex cause the c-ring to rotate in the membrane.
The flow of protons through this subunit drives the conformational changes that result in the binding of substrate on the ATP synthase, ATP synthesis, and release of product (ATP).
How exactly does the Fo subunit operate?
The proton-motive force across the inner mitochondrial membrane, generated by the electron transport chain, drives the passage of protons through the membrane via the FO region of ATP synthase.
A portion of the FO (the ring of c-subunits) rotates as the protons pass through the membrane. The c-ring is tightly attached to the asymmetric central stalk (consisting primarily of the gamma subunit), which rotates within the alpha3beta3 of F1 causing the 3 catalytic nucleotide binding sites to go through a series of conformational changes that leads to ATP synthesis.
The binding change mechanism involves the active site of a β subunit’s cycling between three states.In the “open” state, ADP and phosphate enter the active site. The protein then closes up around the molecules and binds them loosely — the “loose” state. The enzyme then undergoes another change in shape and forces these molecules together, with the active site in the resulting “tight” state (shown in pink) binding the newly produced ATP molecule with very high affinity. Finally, the active site cycles back to the open state, releasing ATP and binding more ADP and phosphate, ready for the next cycle of ATP production.
What would happen if compounds were partially reduced in the electron transport chain?
Hazardous compounds such as superoxides and free radicals would be produced.
What are some inhibitors of oxidative phosphorylation and what steps do they block?
Inhibitors of Complex I, II, and III block electron transport:
a) Rotenone inhibits NADH-UQ oxidoreductase in Complex I; this prevents the reduction of CoQ and the oxidation of the Fe-S clusters of NADH-UQ oxidoreductase.
b) Blocking the activities of Q cytochrome c oxidoreductase prevents passage of protons on to cytochrome c
c) Cyanide, azide, and carbonmonoxide inhibit Complex IV (Cytochrome C oxidase): This inhibits passing protons on to O2 to produce water.
d) ATP synthase inhibitors (Oligomycin and DCCD) block the movement of protons through the Fo subunit. No ATP synthesis occurs. This proves that electron transport and ATP synthysis is tightly coupled.
What do uncouplers do?
Uncouplers disrupt the coupling of electron transport and ATP synthesis.
They dissapate the proton gradient across the inner mitochondrial membrane that is created by the ETC.
Examples include: 2,4-DNP
They are hydrophobic and have a dissociable proton.
They carry protons across the inner membrane to the matrix. In otherwords, protons leak back in via the uncouplers and so ATP synthasis does not occur.
Increased O2 consumption and NADH oxidation.
The energy released in electron transport in this case is released as heat.
What happens when no pH or charge gradient exists?
Instead of ATP being synthesized by ATP synthase, it is hydrolyzed.
How does brown fat respond to cold temperatures?
Uncouple oxid. Phos. from ATP synthesis to generate heat.
In response to a temp drop, the release of hormones leads to the liberation of free fatty acids from triacylglycerols that in turn activate uncoupling protein 1 (UCP-1).
UCP-1 forms a pathway for the flow of H+ from the cytoplasm to the matrix.
The tissue with high UCP-1 called brown fat. (combination of greenish cytochromes and red hemoglobin in blood supply)
How many protons are required by ATP synthase to generate a molecule of ATP?
2.7 protons are required to make each ATP.
What is the ATP-ADP translocase?
ADP enters the mitochondrial matrix only if ATP exits.
ATP movement out of the matrix is favored due to the negative charge on the ATP; the cytosol is + relative to the matrix.
One ATP out requires the energy from a hydrogen.
Thus, making and transporting one ATP requires four hydrogens.
ATP-ADP translocase tightly couples the movement of of ATP out of the matrix with the movement of ADP into the matrix.
How many protons are pumped out of the matrix per electron pair?
NADH-Q reductase - 4
Q-cytochrome C oxidoreductase - 2
Cytochrome c oxidase - 4
Thus 2.5 molecules of cytoplasmic ATP are generated as a result of the flow of a pair of electrons from NADH to O2 and those from the oxidation of succinate, the yield is about 1.5 molecules of ATP per electron pair.
How are electrons of cytosolic NADH shuttled into the ETC?
a) Glycerophosphate shuttle: Two different glycerophosphate dehydrogenases (one in the cytosol and one on the outer face of the mitochondrial inner membrane) work together to carry electrons into the matrix of the mitochondrial matrix.
NADH produced in the cytosol transfers its electrons to dihydroxyacetone phosphate and it is reduced to glycerol-3-phosphate. It is reoxidized FADH2 and its two protons are passed to UQ to form UQH.
This yields 1.5 molecules of ATP.
This mode is ireversible and operates when NADH levels are low.
b) Malate-Aspartate shuttle: Oxaloacetate is reduced in the cytosol and thus acquires the electrons from NADH (which is oxidized NAD+). Malate is transported across the inner membrane where it is reoxidized by malate dehydrogenase converting NAD+ to NADH in the matrix. This NADH enters the ETC. The oxaloacetate is transaminated to form aspartate and is transported back to the cytosolic face of the mitochondrial matrix.
ATP synthase picture.
ATP synthase picture II.
ATP synthase picture III.
ATP binding to Fo subunits.
