section 14.2
Oxidative Phosphorylation
261
The central idea of chemiosmosis, that ATP synthesis
is driven by the proton-motive force generated during res-
piration, is well accepted and is supported by several lines
of experimental evidence. However, some aspects of the
chemiosmotic theory are still to be resolved. For example,
the precise location of proton-generating sites, the path-
ways of proton transport, the mechanism of proton flow,
and how many protons are ejected per site are not well
understood. It was originally proposed that two protons
were ejected at each site, but the number may be more.
Uncoupling Agents of Oxidative Phosphorylation
Uncoupling agents dissociate ATP synthesis and other
energy-dependent membrane functions from the trans-
port of reducing equivalents in the respiratory chain. Nor-
mally, these processes are tightly coupled. These uncou-
pling agents cause a several-fold stimulation of respiration
with unimpeded utilization of substrate and dissipation of
energy as heat. A classic example of an uncoupling agent
is 2,4-dinitrophenol (Figure 14-16);
A group of antibiotics (e.g., valinomycin, nigericin, and
gramicidin A) transport cations across the cell membrane.
Such agents, known as
ionophores,
are widely used to
probe membrane structure and function. Ionophores un-
couple oxidative phosphorylation. Valinomycin, a cyclic
peptide (Figure 14-17), forms a lipid-soluble complex
with K+ that readily passes through the inner membrane,
whereas K+ by itself does not. In the valinomycin-K+
complex, hydrophobic groups, present on the outside, fa-
cilitate transport of the complex in the lipid environment;
In te rm e m b ra n e
s p a c e
In n e r m e m b ra n e
M atrix
N 0
2
n o
2
O
n o
2
o
F I G U R E 1 4 -1 6
Uncoupling of oxidative phosphorylation by 2,4-dinitrophenol (2,4-DNP).
The anionic form of 2,4-DNP is protonated in the intermembrane space, is
lipid soluble, and crosses the inner membrane readily. In the matrix, the
protonated form dissociates, abolishing the proton gradient established by
substrate oxidation. The ionized form of 2,4-DNP is poorly soluble in the
membrane lipids and therefore is not easily transported across the
membrane (dashed arrow). It is lipophilic and capable of transporting
protons from one side of the membrane to the other (a
p r o to n o p h o re ),
thus
abolishing the proton gradient.
K+, located on the inside, interacts with the hydrophilic
groups.
The
matrix
side
of
the
inner
membrane
has
a negative potential,
so
the positively
charged
valinomycin-K+ complex is drawn inward. The com-
plex uncouples oxidative phosphorylation by decreas-
ing the membrane potential. Unlike valinomycin, it
exchanges
H+
for
K+
across
the
membrane.
Its
(CH 3)2CH
«
\
-c—
XCH
n
NH
D -V al
i
(CH3)2CH c
/C H D -H „
0
'2
o—c
(C H ,),C H -C H Lj ( al
1
"
NH
<^3 O
-O-CH-Q^
0 = A
L-L
^ C H
ic
CH3
O
„C
o'
^C
l
(CH 3)2CH
-"NH CH (CH 3)2
^CH
o
L -V al
V
3
O ^CH(CH3)2
CH
k °
\
NH
D-H yi
D -V al
CH -C H (C H 3)2
?~°
L -L a c
6
k -V a i
C.
O
/
CH
7
y
NH
O
D-H yi
8
^-CH
-C H —O -C
\CH(CHj)2
CH (CH 3)2
(a)
(b)
F I G U R E 1 4 - 1 7
Structure of valinomycin (a) and its complex with K+ (b). Valinomycin, which consists of three identical fragments of
D-valyl-L-lactyl-L-valyl-D-cr-hydroxyisovaleric acid (D-Val-L-Lac-L-Val-D-Hyi), is a mobile or channel-forming
ionophore and an uncoupler of oxidative phosphorylation. Note the hydrophobic exterior and the hydrophilic interior of
the complex. [Structure (b) is reproduced, with permission, from B. C. Pressman: Biological application of ionophores.
A n n u . Rev. B io ch em .
45,501 (1976). © 1976 by Annual Reviews Inc.]
