Molecular Biochemistry I

Electron Transfer Chain

Copyright © 1999-2004 by Joyce J.

Diwan.
All rights reserved.
 Electron Transfer

An electron transfer reaction:

Aox + Bred  Ared + Box
Aox is the oxidized form of A (the oxidant)
Bred is the reduced form of B (the reductant).

For such an electron transfer, one may consider two

half-cell reactions:
Aox + n e-  Ared e.g., Fe+++ + e-  Fe++
Box + n e-  Bred
 Aox + n e-  Ared
Box + n e-  Bred
For each half reaction:
E = E°' – RT/nF (ln [reduced]/[oxidized])
e.g., for the first half reaction:
E = E°' – RT/nF (ln [Ared]/[Aox])
E = voltage, R = gas const., F = Faraday, n = # of e .
When [Ared] = [Aox], E = E°'.
E°' is the mid-point potential, or standard redox potential,
the potential at which [oxidant] = [reductant] for the half
reaction.
For an electron transfer:
E°' = E°'(oxidant) – E°'(reductant) = E°'(acceptor) – E°'(donor)
Go' = – nFE°'
(E°' is the mid-point potential)

An electron transfer reaction is spontaneous (negative

G) if E°' of the donor is more negative than E°' of the
acceptor, i.e., when there is a positive E°'.
Consider transfer of 2 electrons from NADH to oxygen:
a. ½ O2 + 2H+ + 2e-  H2O
E°' = +0.815 V
b. NAD+ + 2H+ + 2e-  NADH + H+
E°' = 0.315 V
Subtracting reaction b from a:
 c. ½ O2 + NADH + H+  H2O + NAD+
E°'= +1.13 V
G =  nFEo' = – 2(96494)(1.13) = – 218 kJ/mol
 Electron Carriers

NAD+/NADH and FAD/FADH2 were introduced

earlier.
FMN (Flavin MonoNucleotide) is a prosthetic
group of some flavoproteins.
It is similar in structure to FAD (Flavin Adenine
Dinucleotide), but lacking the adenine nucleotide.
When free in solution, FMN (like FAD) can accept
2 e- + 2 H+ to form FMNH2.
 O O O
H H H H H
C N C C N C C N C
H
3CC C C NH H
3CC C C NH H
3CC C C NH

H
3CC C C CO H
3CC C C CO H
3CC C C CO
C N N C N N C N N
H H H H
CH
2  + CH
2  +
CH
2
e+
H e+
H
H
COH H
COH H
COH

H
COH H
COH H
COH

H
COHO H
COHO H
COHO

H
2C OP O
- H
2C OP O
- H
2C OP O
-

O
- F
MNH O
-
F
MN F
MN·O
H -
2

FMN, when bound at the active site of some enzymes, can

accept 1 e to form the half-reduced semiquinone radical.
The semiquinone can accept a 2nd e to yield FMNH2.
Since it can accept/donate 1 or 2 e, FMN has an important
role mediating e transfer between carriers that transfer 2e
(e.g., NADH) & those that can accept only 1e (e.g., Fe+++).
 O
C H 3O CH 3
CH3
CH 3 H 2C C C CH2
C H 3O (CH 2 CH C CH 2 ) n H
H
O
coenzym e Q isoprene

Coenzyme Q (CoQ, Q, ubiquinone) is very hydrophobic.

It dissolves in the hydrocarbon core of a membrane.
The structure of CoQ includes a long isoprenoid tail, with
multiple units having a carbon skeleton comparable to that
of the compound isoprene. Most often n = 10.
The isoprene tail of Q10 is longer than the width of a lipid
bilayer, but may be folded to yield a more compact shape.
 O
C H 3O CH 3

CH 3
C H 3O ( CH 2 CH C CH 2 ) n H
O coenzym e Q
2 e + 2 H +

O H
C H 3O CH 3
The quinone ring of
CH 3
coenzyme Q can be
C H 3O ( CH 2 CH C CH 2 ) n H
reduced to the quinol
O H coenzym e Q H
in a 2e reaction: 2

Q + 2 e + 2 H+  QH2.
 O O
C H 3O CH 3 C H 3O CH 3
e
CH 3 CH 3
C H 3O (CH 2 CH C CH 2 ) n H C H 3O (CH 2 CH C CH 2 ) n H
O O
c o e n zy m e Q c o e n zy m e Q • 
e + 2 H +
OH
C H 3O CH 3

CH 3
C H 3O (CH 2 CH C CH 2 ) n H
OH c o e n zy m e Q H 2

When bound to special sites in respiratory complexes,

CoQ can accept 1 eto form a semiquinone radical (Q·).
Thus CoQ, like FMN, can mediate between 1 e & 2 e
donors/acceptors.
Coenzyme Q functions as a mobile e carrier within
the mitochondrial inner membrane.
Its role in trans-membrane H+ transport coupled to e
transfer (Q Cycle) will be discussed later.
 CH3

CH3 HC S CH2 protein

N
H3 C CH3
N Fe N
 protein
OOC CH2 CH2 CH S CH2
N
CH3

CH2 CH3

CH2

COO Heme c

Heme is a prosthetic group of cytochromes.

Heme contains an iron atom in a porphyrin ring system.
The Fe is bonded to 4 N atoms of the porphyrin ring.
 CH3

CH3 HC S CH2 protein

N
H3 C CH3
N Fe N
 protein
OOC CH2 CH2 CH S CH2
N
CH3

CH2 CH3

CH2

COO Heme c
Hemes in the 3 classes of cytochrome (a, b, c) differ slightly in
substituents on the porphyrin ring system. A common feature is
2 propionate side-chains. Only heme c is covalently linked to
the protein via thioether bonds to cysteine residues.
 CH3

CH2 CH2 CH C CH2 3 H

CH3 HC OH

O
N
HC CH3
N Fe N

OOC CH2 CH2 CH CH2
N

CH2 CH3

CH2

COO Heme a

Heme a is unique in having a long farnesyl side-chain

that includes 3 isoprenoid units.
 PDB file 5CYT

In the RasMol display of

heme c at right, the
porphyrin ring system is
displayed as ball & sticks,
while Fe is displayed as
spacefill. Heme in cytochrome c

The heme iron can undergo a 1 e transition between

ferric and ferrous states: Fe+++ + e  Fe++
 X
The porphyrin ring is planar.
The heme Fe is usually bonded N N
to 2 axial ligands, above & Fe
below the heme plane (X,Y) in
addition to 4 N of porphyrin. N N

PDB file 5CYT Y

His
Axial ligands may be S or N
atoms of amino acid side-chains.

Axial ligands in cyt c are Met S

(yellow) and His N (blue).
Met
A heme that binds O2 may have
Heme in cytochrome c an open (empty) axial ligand
 Cytochromes

Cytochromes are proteins with heme prosthetic groups.

They absorb light at characteristic wavelengths.
Absorbance changes upon oxidation/reduction of the
heme iron provide a basis for monitoring the redox state
of the heme.
Some cytochromes are part of large integral membrane
complexes, each consisting of several polypeptides and
including multiple electron carriers.
Cytochrome c is instead a small, water-soluble protein
with a single heme group.
Cytochrome c PDB
5CYT

heme complex IV

 



cyt. c
Lys13 Lys 72

Positively charged lysine residues (in magenta) surround

the heme crevice on the surface of cytochrome c.
These may interact with anionic residues on membrane
complexes to which cyt c binds, when receiving or
donating an e.
 PDB file
Cys S 1A70
S Fe
Cys
S Fe S
S Fe S
Cys Cys
S Fe
S

Cys S S S Cys
Fe Fe 2-Fe iron-sulfur
center of ferredoxin
Cys S S S Cys
2 Fe colored orange;
Iron-Sulfur C enters elemental & Cys S yellow.

Iron-sulfur centers (Fe-S) are prosthetic groups containing

14 iron atoms complexed to elemental & cysteine S atoms.
Electron transfer proteins may contain multiple Fe-S centers.
4-Fe centers have a tetrahedral structure, with Fe & S atoms
alternating as vertices of a cube.
 Cys S
S Fe
Cys
S Fe S
S Fe S
Cys Cys
Iron-sulfur centers transfer S Fe
S
only one electron, even if
Cys S S S Cys
they contain two or more
Fe Fe
iron atoms, because of the
close proximity of the iron Cys S S S Cys
atoms. Iron-Sulfur C enters

E.g., a 4-Fe center might cycle between redox states:

Fe+++3, Fe++1 (oxidized) + 1 e  Fe+++2, Fe++2 (reduced)
 matrix

inter-
cristae membrane
space

Respiratory inner outer

membrane mitochondrion membrane
Chain:
Most constitutents of the respiratory chain are
embedded in the inner mitochondrial membrane (or
in the cytoplasmic membrane of aerobic bacteria).
The inner mitochondrial membrane has infoldings
called cristae that increase the membrane area.
 Matrix
H+ + NADH NAD+ + 2H+ 2H+ + ½ O2 H2O

2 e
I Q III IV

cyt c
Intermembrane Space
Electron transfer from NADH to O2 involves multi-subunit
inner membrane complexes I, III & IV, plus CoQ & cyt c.
Within each complex, electrons pass sequentially through
a series of electron carriers.
CoQ is located in the lipid core of the membrane. There
are also binding sites for CoQ within protein complexes.
Cytochrome c resides in the intermembrane space. It
alternately binds to complex III or IV during e transfer.
Composition of Respiratory Chain Complexes
No. of Prosthetic
Complex Name Proteins Groups

Complex I NADH 46 FMN,

Dehydrogenase 7 Fe-S cntrs.

Complex II Succinate-CoQ 5 FAD, cyt b560,

Reductase 3 Fe-S cntrs.

Complex III CoQ-cyt c 11 cyt bH, cyt bL,

Reductase cyt c1, Fe-SRieske

Complex IV Cytochrome 13 cyt a, cyt a3,

Oxidase CuA, CuB
 Matrix
H+ + NADH NAD+ + 2H+ 2H+ + ½ O2 H2O

2 e
I Q III IV

cyt c
Intermembrane Space

Mid-point potentials of constituent e carriers are

consistent with the e transfers shown being spontaneous.
Respiratory chain inhibitors include:
Rotenone (a rat poison) blocks complex I.
Antimycin A blocks electron transfer in complex III.
CN & CO inhibit complex IV.
Inhibition at any of these sites will block e transfer from
NADH to O2.
 NAD+
NADH
Complex I
catalyzes matrix
oxidation of
inner mitochondrial
NADH, with membrane Complex I
reduction of
coenzyme Q:
NADH + H+ + Q  NAD+ + QH2
Transmembrane H+ flux associated with this reaction is
discussed in the section on oxidative phosphorylation.
Complex I is L-shaped. A high-resolution crystal
structure is not yet available for this large complex that
in mammals includes at least 46 proteins.
 NAD+
An low-
NADH
resolution EM
structure of matrix
complex I is
displayed on a inner mitochondrial
membrane Complex I
website of
N. Grigorieff.

The domain where NADH interacts protrudes into

the mitochondrial matrix.
Coenzyme Q binds within the membrane domain.
Fe-S centers are in the NADH-binding domain & in a
connecting domain closer to the membrane segment.
The initial electron transfers are:
NADH + H+ + FMN  NAD+ + FMNH2
FMNH2 + (Fe-S)ox  FMNH· + (Fe-S)red + H+

After Fe-S is reoxidized by transfer of the electron to

the next iron-sulfur center in the pathway:
FMNH· + (Fe-S)ox  FMN + (Fe-S)red + H+

Electrons pass through a series of iron-sulfur centers in

complex I, eventually to coenzyme Q.
Coenzyme Q accepts 2 e and picks up 2 H+ to yield the
fully reduced QH2.
 COO COO
Q QH2
H C H C H
Succinate
Dehydrogenase of H C H via FAD H C
the Krebs Cycle is COO COO
also called complex II succinate fumarate
or Succinate-CoQ Succinate Dehydrogenase
Reductase. (Complex II)

FAD is the initial electron receptor.

FAD is reduced to FADH2 during oxidation of succinate
to fumarate.
FADH2 is then reoxidized by transfer of electrons
through a series of three iron-sulfur centers to Coenzyme
Q, yielding QH2.
 Complex II

OAA FAD
X-ray crystallographic
analysis of E. coli FeS
complex II indicates a
linear arrangement of CoQ
electron carriers within heme
membrane
complex II, consistent with domain
the predicted sequence of
electron transfers: PDB 1NEK

FAD  FeScenter 1  FeScenter 2  FeScenter 3  CoQ

In this crystal structure oxaloacetate (OAA) is bound in place of
succinate.
 Matrix
H+ + NADH NAD+ + 2H+ 2H+ + ½ O2 H2O

2 e
I Q III IV

cyt c
Intermembrane Space

Complex III accepts electrons from coenzyme QH2 that

is generated by electron transfer in complexes I & II.
The structure and roles of complex III are discussed in the
class on oxidative phosphorylation.
Cytochrome c1, a prosthetic group within complex III,
reduces cytochrome c, which is the electron donor to
complex IV.
 Matrix
H+ + NADH NAD+ + 2H+ 2H+ + ½ O2 H2O

2 e
I Q III IV

cyt c
Intermembrane Space

Cytochrome oxidase (complex IV) carries out the

following irreversible reaction:
O2 + 4 H+ + 4 e  2 H2O
The four electrons are transferred into the complex one
at a time from cytochrome c.
 membrane

Cytochrome oxidase dimer (PDB file 1OCC)

Intramembrane domains of cytochrome oxidase

(complex IV) consist mainly of transmembrane
-helices.
 Complex IV
binuclear center

heme a3 CuB

PDB 1OCC

Metal centers of cytochrome oxidase (complex IV):

heme a & heme a3,
CuA (2 adjacent Cu atoms) & CuB.
O2 reacts at a binuclear center consisting of heme a3
and CuB.
 PDB file 1OCC

Metal center ligands in

complex IV:
Heme axial ligands are His N
atoms.
Heme a is held in place
between 2 transmembrane
-helices by its axial His
ligands.
Liganding of Heme a
in Cytochrome Oxidase
 binuclear center
Heme a3, which sits adjacent to His ligands
CuB, has only one axial ligand.
Cu ligands consist of His N, &
heme a3
in the case of CuA also Cys S,
Met S, & a Glu backbone O. CuB
Electrons transferred from cyt c
enter complex IV through CuA PDB 1OCC
& heme a.
They then pass to the binuclear center where the
chemical reaction takes place.
O2 binds at the open axial ligand position of heme a3,
adjacent to CuB.
 binuclear center
O2 + 4 H+ + 4 e  2 H2O His ligands

Details of the reaction sequence

are still debated. heme a3
A Tyr-His complex adjacent to
the binuclear center is postulated CuB
to have a role in O-O bond
splitting. PDB 1OCC

The open axial ligand position of heme a3 makes it

susceptible to binding of CN, CO, or the radical signal
molecule ·NO.
All inhibit cytochrome oxidase activity.
Regulation of respiration by ·NO has been postulated.