Energetics of Electron

Transport
Zeynep A. Durer, Ph.D.
 Learning Objectives

• Compare the energetics in anaerobic and aerobic cells.

• State the components of proton motive force.
• Understand how electron transfer potential is converted to phosphate
transfer potential in mitochondria.
• State the relation between redox potential and free energy.
• Know how free energy change of oxidation-reduction reactions are
calculated.
• Describe chemiosmotic hypothesis.
 References
• Biochemistry Berg, Ytmoczko, Stryer
• Physical Chemistry with Biological Applications
Laidler
• Lehninger Principles of Biochemistry by Nelson and
Cox
• The Molecules of Life Kuriyan, Koforti &Wemmer
A comparison of biological oxidation
with combustion.
Schematic representation of the controlled
stepwise oxidation of sugar in a cell, compared
with ordinary burning.
An outline of glycolysis.
 Generation of ATP in Anaerobic
Cells
• Aerobic cells  O2 final electron acceptor and oxidant
of foodstuff
• Anaerobic cells  some molecule other than O2 is the
oxidant.
Two fragments A and B; A oxidizes B
Coupled to oxidation: formation of ATP
Fermentation of glucose to lactic acid-anaerobic organisms
– Yeast: Glucose  ethanol
– Some bacteria Glucose  acetone or butanol
– Others Glucose  lactic acid
Two pathways for the anaerobic
breakdown of pyruvate.
 Glycolysis
• No oxygen is required
Balanced equation ΔG′
Eqn.1 C6H12O6  2 CH3CHOHCOOH -52
kcal/mole
(lactate)
Anaerobic breakdown of glucose in living cells takes
place coupled to ATP formation
Eqn.2 Glucose + 2HPO + 2ADP  2 lactate + 2ATP + 2H O
4
2-
2

-38 kcal/mole
 ATP Formation and Glucose
Breakdown are Coupled
• According to equation 1, ΔG’ = -52 kcal/mole
• According to equation 2, 2 moles of ATP
produced (52-38= 14 kcal/mole)
• 14,000  27%  the PROFIT in glucose
52,000 fermentation
 Priming the Pump
• TWO molecules of ATP are needed to ‘PRIME’ the pump.
– Glucose + ATP  Gl-1-ph + ADP + H+ (hexokinase)
– Fructose-6-phosphate + ATP  Fr-1,6-biph + ADP +
H+ (phosphofructokinase)
• Later
– 2ADP  2ATP
1,3-BPG + ADP  3-phosphoglycerate + ATP
(phosphoglycerate kinase)
– 2ADP  2ATP
phosphoenolpyruvate + ADP +H+  pyruvate + ATP
(pyruvate kinase)
 Glycolysis

• NET : 2 ATP
• Glucose + 2 ATP + 2 phosphate + 4 ADP 2
lactate + 2 ADP + 4 ATP
• Glucose + 2 phosphate + 2 ADP  2 lactate +
2 ATP
•  All intermediates (except glucose and
pyruvate-lactate) are esters of phosphoric acid
 Energy Conserving Steps
• First energy conserving step:
Glyceraldehyde 3-phosphate + NADox + Pi  1,3-BPG + NADred + H+
ΔG0 +1.5 kcal/mole
1,3 BPG + ADP  3-phosphoglycerate + ATP
ΔG0 -4.5 kcal/mole
2 moles of 3-phosphoglyceraldehyde  2 moles of ATP
• Second energy conserving step:
Intramolecular oxidation-reduction reaction
Pyruvate kinase
PEP + ADP + H+ ------------------- pyruvate + ATP
 Phosphate transfer potential of PEP is much higher than that of ATP.
 Energetics of Fermentation and
Respiration
• Glucose- Fermentation-- 2 lactate---Respiration-- 6CO2 + 6H2O
ΔG = -52 kcal
ΔG =-686 kcal
52,000  a little over 7%
686,000
– Anaeorobic organism wastes  90%
– Aerobic organism: lactate does not leave the cell. 93% of the energy of
glucose  respiration
– Anaerobic organism: more fuel/time, weight; uses 20X as much
glucose as aerobic cells to do the same amount of work
Cancer cells (facultative cells)- metabolic defect- use immense quantities
of glucose by glycolysis even though they are supplied with O 2 and are still
able to respire.
Oxidative Phosphorylation
 ATP yield from complete oxidation of glucose
Electron shuttle
Cytoplasm across membrane Mitochondrion

2 NADH 2 NADH
(or 2 FADH2)
2 NADH 6 NADH 2 FADH2

GLYCOLYSIS OXIDATIVE
2 2 Acetyl CITRIC ACID PHOSPHORYLATION
Glucose Pyruvate CoA (Electron Transport
CYCLE
and Chemiosmosis)

2 ATP 2 ATP about 34 ATP

by substrate-level by substrate-level by oxidative phosphorylation
phosphorylation phosphorylation

Maximum per glucose: About

38 ATP
 Oxidative Phosphorylation
• Process in which ATP is formed as electrons are transferred from
NADH or FADH2 to O2 by a series of electron carriers.
– 32 out of 36 (38 ) ATP***
• Krebs cycle 3 NADH formed  3 ATP each
1 FADH2 "  2 ATP each
• The oxidation of fuels and the phosphorylation of ADP are coupled by a
proton gradient across the inner mitochondrial membrane.

• *** More recent textbooks: 30 ATP in total

2,5 ATP/NADH
1,5 ATP/ FADH2
 Proton Motive Force
• Respiratory assembly -
electron carriers - proton
motive force generated.
• Transfer of electrons from
NADH ( FADH2 ) to O2 
pumping of protons out of
mitochondrial matrix.
i. pH gradient
ii. Transmembrane
potential
• ATP formed as protons flow
BACK to the matrix
Energetics of Electron Transport
Electron transfer potential (measured by E0, redox
potential) of NADH or FADH2

Phosphate transfer potential of ATP (measured by ΔG 0)

Chemical reactions that involve the transfer of an electron from
one molecule to another are referred to as redox reactions.
Energetics of Electron Transport

– Negative redox potential  lower affinity for

electrons than does H2 (0 volt)
– Positive redox potential  higher affinity for
electrons than does H2 (0 volt)

– Strong reducing agent (NADH)  Negative redox

potential
– Strong oxidizing agent (O2)  Positive redox
potential
 Measurement of Redox Potentials

• E0 ′ (Conjugate redox pair at 1M

concentrations and pH 7 is
connected with the standard (pH
0 ) electrode: 1 atm H2
equilibrated with 1 M H+)
• ΔG0 ′ = -nFΔE0′
– n= number of electrons transferred
– F= the faraday (96.48 kJ/mol V or
23.06 kcal/mol V
– Reduction potential in V
 Standard Reduction Potentials at
25oC
Half Reaction E′o (V)
1/2O2+ 2 H+ + 2 e- -----> H2O +0.82
NAD+ + 2 e- -----> NADH + H+ - 0.32
Pyruvate + 2 e- -----> Lactate -0.19
2 H+(aq) + 2 e- -----> H2(g) -0.42

*
Note: E′0 is the standard oxidation-reduction potential (pH 7, 25°C)
Note from the Table:E0 for H2/H+ is -0,42 V at pH 7 (10-7 M H+)
Standart hydrogen electrode (25C,1 M H+,1M H2) is 0

E′0 refers to the partial reaction written as

Oxidant + e- ----->reductant
 Free Energy Change of Oxidation-
Reduction Reactions
• The free energy change of an oxidation -reduction reaction can be readily
calculated from the diff. in reduction potential of the reactions.
• For example: Consider the reduction of pyruvate by NADH
– a) pyruvate+ NADH+H+ Lactate + NAD+
Reduction potential of the NADH/NAD+ couple is -0.32 V
OXIDANT REDUCTANT n E0'(volts)
NAD+ NADH+H+ 2 -0,32
whereas that of pyruvate :lactate couple is -0,19 volts
OXIDANT REDUCTANT n E0'(volts)
Pyruvate Lactate 2 -0,19
 Partial Reactions for
Pyruvate/Lactate Pair
– b) pyruvate+2H++2e- Lactate -0,19V
– c) NAD+ +2H++2e- NADH+H+ -0,32V
– b-c =a=-019+0.32=+0.13 V
• Now we can calculate the Go'= -n F Eo'
– n= number of electrons transferred
– F= Faraday constant : The energy change as 1 mole of electrons falls through a
potential of 1 volt . (23,06 kcal/Vmol)
– Eo'= Standart red.pot. in volts
– Go'= free energy in kcal/mol
Go'= (-2)(23.06)(+0.13)= -6 kcal/mol
 E ′ and G ′
o o

• A positive Eo' (negative Go') signifies an exergonic reaction

under standard conditions)
• Electrons will tend to pass from the most (-) carrier (NAD+) to
the more (+) carrier below it.
• The driving force of oxidative phosphorylation is the electron
transfer potential of NADH or FADH2 relative to that of O2
 Respiratory Chain

a) 1/2 O2+ 2H++2e- H2O +0,82 V

b)NAD++H+ +2e- NADH -0.32 V
c) 1/2 O2+NADH+H+ H2O+NAD+ +1.14 V
• a-b=c. Go=-nFEo′ =(-2)(23.06)(+1.14) =-52.6 kcal/mol
as a pair of electrons moves the entire length of the chain.
• All together 12 pairs of electrons pass down the respiratory chain to
oxygen during complete oxidation of 1 mole of glucose to CO 2 and H2O.
• A 1.14-volt potential difference between NADH and O2 drives electron
transport through the chain and favors the formation of a proton gradient
 Complete Oxidation of Glucose

– (12)(52600)=-634000 cal/mol
• The process of electron transport from NAD red. to oxygen
can account for -634 kcal/mol of glucose oxidized. If we recall
than G of combustion of glucose is -686 kcal/mol, it is clear
that almost all of the free energy decrease in biological
oxidation of glucose occurs during the enzymatic transport of
electrons from the FIRST ELECTRON ACCEPTOR along the
respiratory chain to molecular oxygen.
– 634 X 100  93%
684
 Conservation of Energy as ATP

• For a pair of electrons: -52.6 kcal

– 3 ATP formed per NADH**
3 ADP+Pi 3ATP Go=(3)(+7.3)=+21.9 kcal/mol
Coupled phosp.of 3 molecules of ATP (under cellular
conditions 11-12 kcal may be needed for each ATP
syntesized)
**Note: currently accepted value is 2,5 ATP/NADH
• Thus conserves (21.9 / 52.6 )X 100 or 42% of the total
free energy decline during transport of a pair of
electrons from NADH to Oxygen under the usual
standard conditions.
 Conservation of Energy as ATP

• The respiratory chain is really a molecular

device for delivering the 52000 cal. of energy
in a series of small packets (quanta), three of
which are energetically equivalent to ATP.
• Compare to fermentation 14/52 27%
 Oxidation and Phosphorylation are
Coupled by a Proton Motive Force

• NADH+1/2O2+H+ H2O +NAD+ Go=-52.6kcal/mol

• ADP+Pi+H+ ATP +H2O + Go= +7.3 kcal/mol
• The synthesis of ATP is carried out by a molecular assembly in the inner
mitochondrial membrane .This enzyme complex has been called the
mitochondrial ATP or H+ ATPase. How is the oxid. of NADH coupled to
the P of ADP ?
 Cheimiosmotic Hypothesis by Peter Mitchell
• Electron transport and ATP synthesis are coupled by
a proton gradient across the inner mitochondrial
membrane. In his model, the transfer of e- s through
the respiratory chain leads to the pumping of protons
from the matrix to the other side of the inner
mitochondrial membrane. The H+ concentration
becomes higher in the cytosolic side and an electric
potential with that side (+) is generated.

The Nobel Prize in Chemistry 1978

was awarded to Peter Mitchell
"for his contribution to the
understanding of biological energy
transfer through the formulation of
the chemiosmotic theory"
 Chemiosmotic Hypothesis
• Mitchell postulated that the proton-motive force drives the synthesis of ATP by
the ATPase complex. The primary energy conserving event in this model is the
movement of protons across the inner mitochondria membrane.
• Both a H+ gradient contribution and a membrane potential contribution
– pH 1.4 with lower than inside
– membrane potential 0.14 V ( outside positive)
 The Two Components of the
Electrochemical Proton Gradient

• The total proton-motive force across the inner mitochondrial membrane

consists of a large force due to the membrane potential (traditionally
designated ΔΨ by experts, but designated ΔV here) and a smaller force
due to the H+ concentration gradient (ΔpH). Both forces act to drive
H+ into the matrix.
• Copyright © 2002 Bruce Alberts, Dennis Bray, Julian Lewis, Martin Raff, Keith
Roberts, and James D. Watson
 Free Energy Stored in
Concentration Gradients
• The free-energy change in transporting a molecule from side 1, to side 2, is

• For a charged species, the unequal distribution across the membrane

generates an electrical potential that also must be considered because the
ions will be repelled by the like charges. The sum of the concentration and
electrical terms is called the electrochemical potential. The free-energy
change is then given by

• in which Z is the electrical charge of the transported species, ΔV is the

potential in volts across the membrane, and F is the faraday [23.1 kcal V -1
mol-1].
 Energy Associated with a Proton
Gradient

• under typical conditions for the inner mitochondrial membrane, the

pH outside is 1.4 units lower than inside [corresponding to log 10
(c2/c1) of 1.4] and the membrane potential is 0.14 V, the outside
being positive. Because Z = +1 for protons, the free-energy change
is (2.303 × 1.98 × 10-3 kcal mol-1 K-1 × 310 K × 1.4) + ( + 1 × 23.06
kcal mol-1 V-1 × 0.14 V) = 5.2 kcal mol-1 Thus, each proton that is
transported out of the matrix to the cytosolic side corresponds to 5.2
kcal/ mol of free energy.
 PMF and ΔG

• ΔμH = ΔΨ – 2.3 RTΔpH /F

Electrochemical Membrane pH gradient H+gradient (pmf)
Potential

ΔpH gradient generates an electrical potential (Δμ H ) of

0.14 V across the membrane. ΔpH (1.4 units lower
than in the matrix) has a negative value because it is
measured from inside to outside= +0.084 V.
G=-nFEo′= -5.2 kcal/mole
 Coupling of Oxidation and ATP
Synthesis

• 10 protons transported per electron pair

•  52 kcal /mole of electron pair for NADH
• 3 (or 2,5)ATP per NADH
• 10 / 3 = 3 -4 mole of protons/mole ATP
 Energy Balance Sheet

• Glucose +36/38ADP+36/38Pi+6O2 6CO2+

36/38ATP+ 4 H2O
32/36(38) oxid. phosph.***
Thermodynamic efficiency of ATP formation
• For 36 ATP G=263 kcal
– 263/686=38%
– For 38ATP 42%
*** 30 ATP according to more recent textbooks
NOTES
Relating standard free-energy difference (ΔG°)
to the equilibrium constant (K).
Simple overview of the citric acid
cycle.
Glycolysis and the citric acid
cycle provide the precursors
needed to synthesize many
important biological molecules.
The generation of an H+ gradient
across a membrane by electron-
transport reactions.