Mary K. Campbell
Shawn O. Farrell http://academic.cengage.com/chemistry/campbell
Paul D. Adams • University of Arkansas

The Role of Electron Transport in Metabolism
• Electron transport is carried out by four closely related multisubunit membrane-bound complexes and two electron carriers, coenzyme ___ and cytochrome ___
• In a series of oxidation-reduction reactions, electrons from FADH
2 and NADH are transferred from one complex to the next until they reach _____
• O
2 is reduced to H
2
O
• As a result of electron transport, ____________ are pumped across the inner membrane to the intermembrane space, creating a pH gradient

• The production of ATP in the mitochondria is the result of oxidative phosphorylation
• The proton gradient establishes a voltage gradient
• The proton and voltage gradients together provide the mechanism to couple electron transport with
_______________________________________

• Electron transport from one carrier to another creates a proton gradient across the inner mitochondrial membrane
• The proton gradient is coupled to the production of ATP in aerobic metabolism

• A useful way to look at electron transport is to consider the change in free energy associated with the movement of electrons from one carrier to another
• If we have two electron carriers, for example NADH and coenzyme Q, are electrons more likely to be transferred from NADH to coenzyme Q, or vice versa?
• What we need to know is the _____________
_____________ for each carrier
• A carrier of high reduction potential will tend to be
_____________ if it is paired with a carrier of
_____________ reduction potential

• Standard reduction potentials provide a basis for comparison among redox reactions
• The sequence of reactions in the electron transport chain can be predicted by using reduction potentials.

• Complex I: NADH-CoQ oxidoreductase
• Electrons are passed from
NADH to FMN
NADH + H + E-FMN
NAD + + E-FMNH
2

• Electrons are then passed to the iron-sulfur clusters
• The last step of Complex I involves electrons being passed to coenzyme Q (also called ubiquinone)

• The transfer of electrons is strongly ____________ and sufficient to drive the phosphorylation of ADP

Complex II: Succinate-coenzyme Q oxidoreductase
Succinate + E-FAD Fumarate + E-FADH
2
• The overall reaction is exergonic (-13.5 kJ/mol), but not enough to drive ATP production
• No H + is pumped out of the matrix during this step

Complex III: CoQH
2
-cytochrome c oxidoreductase
CoQH
2
+ 2Cyt c[Fe(III)] CoQ + 2Cyt c[Fe(II)] + 2H +
• This reactions of this complex results in a decrease in free energy that is sufficient to drive the phosphorylation of ADP to ATP
• The flow of electrons from reduced CoQ, a quinone that can exist in 3 forms, is known as the _____________

Complex IV: Cytochrome c oxidase
• Catalyzes the final step in electron transport
2Cyt c[Fe(II)] + 2H + + ½O
2
2 Cyt c[Fe(III)] + H
2
O
• Complex IV contains cytochrome a, cytochrome a
3
, and Cu(II), which are also involved in the electron transport
• Complex IV is the link to __________ __________

The Energetics of Electron Transport Reactions

• All cytochromes contain a _____________ group
• Side chain differences depending on the heme

Connection between Electron Transport & Phosphorylation
• The energy-releasing oxidations give rise to proton pumping and a pH gradient across the _____________
_____________ membrane
• Differences in the concentration of ions across the membrane generates a _____________ _____________
• A coupling process converts the electrochemical potential to the chemical energy of ATP
• The coupling factor is ATP _____________, a complex protein oligomer, separate from the electron transport complexes
• Uncouplers inhibit the phosphorylation of ADP without affecting electron transport; examples are
________________, _____________, & _____________

• P/O ratio: the number of moles of P i consumed in
_____________ to the number of moles of oxygen atoms consumed in _____________
• Phosphorylation : ADP + P i
• Oxidation : 1/2O
2
+ 2H + + 2e -
ATP + H
H
2
O
2
O
• P/O = ______ when NADH is oxidized
• P/O = ______ when FADH
2 is oxidized

• The coupling of electron transport to oxidative phosphorylation requires a multisubunit membrane-bound enzyme, ATP synthase.
• This enzyme has a channel for protons to flow from the intermembrane space into the mitochondrial matrix.
• The proton flow is coupled to ATP production in a process that appears to involve a conformational change of the enzyme.

• Chemiosmotic coupling
• based on a ________ concentration gradient between the intermembrane space and the matrix
• a proton gradient exists because the various proteins that serve as electron carriers are not symmetrically oriented with respect to the two sides of the inner mitochondrial membrane
• these proteins take up protons from the matrix when they are reduced and release them to the intermembrane space when they are reoxidized
• the reactions of NADH, CoQ, and O
2
_______________________ all require

Evidence for chemiosmotic coupling suggested (Mitchell 1961):
• A system with definite inside and outside _____________
(closed vesicles) is essential.
• Submitochondrial vesicles can be prepared, which carry out oxidative phosphorylation and have an asymmetric orientation of respiratory complexes.
• A model system for oxidative phosphorylation can be constructed with proton pumping in the absence of electron transport; the model system consists of reconstituted membrane vesicles, mitochondrial ATP synthase, and a proton pump.
• The existence of the pH gradient has been demonstrated and _____________ _____________

• The mechanism by which the proton gradient leads to the production of ATP depends on ion channels through the inner mitochondrial membrane
• Protons flow back into the matrix through channels in the F
0 unit of ATP _____________
• The flow of protons is accompanied by formation of
ATP in the F
1 unit of ATP _____________
• The details of how phosphorylation takes place as a result of the linkage to the proton gradient are not explicitly specified by this mechanism

• The proton gradient leads to changes in conformation in a number of proteins, including ATP _______________
• There are 3 sites for substrate on ATP _____________, and 3 possible conformations:
- Open (O); a low affinity for substrate
- Loose-binding (L); not catalytically active, binds ADP & P i
- Tight-binding (T); catalytically active, binds ATP
• These sites interconvert as a result of proton flux through ATP synthase
• Proton flux converts L to T, which produces ATP
• Proton flux converts T to O, releasing ATP

Release of ATP from
ATP Synthase

• In chemiosmotic coupling, the proton gradient is the crux of the matter. The flow of protons through pores in the synthase drives ATP production.
• In conformational coupling, a change in the shape of the synthase releases bound ATP that has already been formed.

• Respiratory inhibitors can be used to determine the order of reactions in the electron transport chain
• Isolate intact _____________ from cells
• Provide an oxidizable substrate so that electron transport can occur
• Add a respiratory inhibitor
• Measure relative amounts of oxidized and reduced forms of various electron carriers
• Inhibitors have an effect on three sites in the electron transport chain

Sites of Action of Some Respiratory Inhibitors

• Shuttle mechanisms: transport metabolites between _____________ and _____________
• Glycerol phosphate shuttle:
• We know glycolysis in the _____________ produces
NADH
• NADH does not cross the _____________________ membrane, but glycerol phosphate and dihydroxyacetone phosphate do
• Through the glycerol phosphate shuttle, ____ ATP are produced in the mitochondria for each cytosolic NADH

• The Malate-Aspartate Shuttle:
• Has been found in mammalian kidney, liver, and heart
• Malate crosses the mitochondrial membrane, while oxaloacetate cannot
• The transfer of electrons from NADH in the cytosol produces NADH in the mitochondria
• In the malate-aspartate shuttle, _____ mitochondrial
ATP are produced for each cytosolic NADH

• Shuttle mechanisms transfer ___________, but not _____________, from the cytosol across the mitochondrial membrane
• In the malate-aspartate shuttle, 2.5 molecules of ATP are produced for each molecule of cytosolic NADH, rather than 1.5
ATP in the glycerolphosphate shuttle…
• This affects the overall yield of ATP in these tissues

ATP Yield from Complete Oxidation of Glucose
• In the complete oxidation of glucose, a total of
______ or ______ molecules of ATP are produced for each molecule of glucose, depending on the shuttle mechanism

The ATP Yield from Complete Oxidation of Glucose