Electron Transport and Oxidative

Phosphorylation

DR. REEM ARAFA

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An Overview
 Electron Transport: Electrons carried by reduced
coenzymes are passed through a chain of proteins and
coenzymes to drive the generation of a proton gradient
across the inner mitochondrial membrane

 Oxidative Phosphorylation: The proton gradient runs

downhill to drive the synthesis of ATP

 It all happens in or at the inner mitochondrial

membrane
 21.2 Reduction Potentials
High positive Eo' indicates a strong affinity for electrons and a hightendency to be
reduced
 Crucial equation: Go' = -nF Eo'

 Eo' = Eo'(acceptor) - Eo'(donor)

 Electrons are donated by the half reaction with the more

negative reduction potential and are accepted by the reaction with
the more positive reduction potential: Eo’ positive, Go' negative

 If a given reaction is written so the reverse is true, then the Eo'

will be a negative number and Go' will be positive
21.3 Electron Transport

 Four protein complexes in the inner mitochondrial

membrane

 A lipid soluble coenzyme (UQ, CoQ) and a water

soluble protein (cyt c) shuttle between protein
complexes

 Electrons generally fall in energy through the chain -

from complexes I and II to complex IV
 ETC and the oxphos

H
H++ HH++++
H
H++
H
H++H
H++
H
H
OUT Cyt C Cyt C
e- e- e- e-
e- Q e- e-

IN e- e- -- ++H
H
H H
++ ++
2OH H ++-- O2 OH
H
HH++++H
OH HH++ 2O
HH++HH O2
H
HH
+`
H HH++NAD
+`+`
+`
NAD++
ADP ATP

NADH
NADH TCA
Cycle

Peter S. Rabinwitch after Mandavilli et al, Mutation Research 509 (2002) 127–151
21.4 Complex I
NADH-CoQ Reductase
 Electron transfer from NADH to CoQ

 More than 30 protein subunits - mass of 850 kD

 Path:
 NADH  FMN  Fe-S  UQH2 Fe-S  UQH2

 4H+ transported out per 2e-

21.5 Complex II
Succinate-CoQ Reductase
 aka succinate dehydrogenase (from TCA cycle!)

 aka flavoprotein 2 (FP2) - FAD covalently bound

 four subunits, including 2 Fe-S proteins

 Path: succinate  FADH2  2Fe2+  UQH2

 Net reaction:
 succinate + UQ  fumarate + UQH2
 21.6 Complex III
CoQ-Cytochrome c Reductase

 CoQ passes electrons to cyt c (and pumps H+) in a unique

redox cycle known as the Q cycle

 Cytochromes, like Fe in Fe-S clusters, are one- electron

transfer agents

 UQH2 is a lipid-soluble electron carrier

 cyt c is a water-soluble electron carrier

21.7 Complex IV
Cytochrome c Oxidase

 Electrons from cyt c are used in a four-electron

reduction of O2 to produce 2H2O

 Oxygen is thus the terminal acceptor of electrons in the

ETC - the end!

 Complex IV also transports H+

Coupling e- Transport and
Oxidative Phosphorylation
This coupling was a mystery for many years

 Many biochemists wasted careers searching for the elusive

"high energy intermediate"

 Peter Mitchell proposed a novel idea - a proton gradient

across the inner membrane could be used to drive ATP
synthesis

 Mitchell was ridiculed, but the chemiosmotic hypothesis

eventually won him a Nobel prize
21.9 ATP Synthase

Proton diffusion through the protein drives ATP

synthesis!

 Two parts: F1 and F0 (latter was originally "F-o" for its

inhibition by oligomycin)

 Racker & Stoeckenius confirmed Mitchell’s hypothesis

using vesicles containing the ATP synthase and
bacteriorhodopsin
21.10 Inhibitors of Oxidative
Phosphorylation
 Rotenone inhibits Complex I - and helps natives of the
Amazon rain forest catch fish!

 Cyanide, azide and CO inhibit Complex IV, binding

tightly to the ferric form (Fe3+)

 Oligomycin is an ATP synthase inhibitors

 Cyanide
Antimycin A
Azide, CO Oligomycin
Rotenone H+

FCCP
(or DNP)
21.11 Uncouplers
Uncoupling e- transport and oxidative phosphorylation

 Uncouplers disrupt the tight coupling between electron

transport and oxidative phosphorylation by dissipating the
proton gradient

 Uncouplers are hydrophobic molecules with a dissociable

proton

 They shuttle back and forth across the membrane, carrying

protons to dissipate the gradient
 Antimycin A Azide Oligomycin
Rotenone H+

FCCP
(or DNP)
 21.12 ATP-ADP Translocase
ATP must be transported out of the mitochondria
 ATP out, ADP in - through a "translocase"

 ATP movement out is favored because the cytosol is "+" relative to the "-"
mitochondrial matrix

 But ATP out and ADP in is net movement of a negative charge out -
equivalent to a H+ going in

 So every ATP transported out costs one H+

 One ATP synthesis costs about 3 H+

 Thus, making and exporting 1 ATP = 4H+

21.13 What is the P/O Ratio??
i.e., How many ATP made per electron pair through the
chain?
 e- transport chain yields 10H+ pumped out per electron pair
from NADH to oxygen

 4H+ flow back into matrix per ATP to cytosol

 10/4 = 2.5 for electrons entering as NADH

 For electrons entering as succinate (FADH2), about 6 H+

pumped per electron pair to oxygen

 6/4 = 1.5 for electrons entering as succinate

Net Yield of ATP from Glucose
It depends on which shuttle is used!
 32 ATP per glucose if malate-asp shuttle used

 Inbacteria - no mitochondria - no extra H+

used to export ATP to cytosol, so:
 10/3 = ~3ATP/NADH

 6/3 = ~ 2ATP/FADH2