In higher organisms, the oxidation of pyruvate takes place
in subcellular, membranous organelles known as mitochondria.
Because mitochondria are responsible for the
synthesis of most of the ATP in nonphotosynthetic tissue,
they are often referred to as the powerhouses of cells.
Mitochondrial ATP synthesis is called oxidative phosphorylation
since it is linked indirectly to oxidative reactions.
In the complete oxidation of pyruvate, there are five
oxidation–reduction reactions. Three of these reactions are
oxidative decarboxylations. The electron acceptor (oxidizing
agent) for four of the reactions is NAD+
; the oxidizing
agent for the fifth is flavin adenine dinucleotide, or FAD.
Knowing the oxidation–reduction potentials of the reactants
in an oxidation–reduction reaction permits the ready
calculation of the standard free energy change for the reaction.
It may be shown that
|⇒ Equation 
is the number of electrons transferred in the reaction, F
is Faraday’s constant (23,060 cal/V-equivalent),
is the difference between the ΔE0'
value of the oxidizing agent and that of the reducing agent.
The reduced form of NAD+, NADH, is a strong reducing
agent. The ΔE0'
at pH 7.0 of the NAD+−NADH couple
is −340 mV, which is equivalent to that of molecular hydrogen.
is the potential when the concentrations of the
oxidized and reduced species of an oxidation–reduction
pair are equal. Reduced FAD, FADH2
, is a weaker reductant
than NADH, with an ΔE0'
(pH 7.0) of about 0 V. In
contrast, molecular oxygen is a potent oxidizing agent and
fully reduced oxygen,water, is a very poor reducing agent.
(pH 7.0) for the oxygen–water couple is +815mV.
The oxidation of NADH and FADH2
results in the reduction
of oxygen to water:
|⇒ Equation 
||H+ + NADH + ½O2 → NAD+ + H2O
|⇒ Equation 
||FADH2 + ½O2 → FAD + H2O.
In both cases two electrons are transferred to oxygen,
so that the n
in Eq. (1) is equal to 2. Under standard
conditions, the oxidation of 1 mol of NADH by oxygen
liberates close to 53 kcal, whereas the ΔG0'
for that of
is −38 kcal/mol. These two strongly exergonic reactions
provide the energy for the endergonic synthesis of
The details of carbon metabolism in the citric acid cycle
are beyond the scope of this article. In brief, pyruvate
is first oxidatively decarboxylated to yield CO2
and an acetyl group attached in an ester linkage to a thiol
on a large molecule, known as coenzyme A, or CoA. (See
Fig. 2.) Acetyl CoA condenses with a four-carbon dicarboxylic
acid to form the tricarboxylic acid citrate. Free
CoA is also a product (Fig. 6). A total of four oxidation–
reduction reactions, two of which are oxidative decarboxylations,
take place, which results in the generation of the
three remaining NADH molecules and one molecule of
. The citric acid cycle is a true cycle. For each
two-carbon acetyl moiety oxidized in the cycle, two CO2
molecules are produced and the four-carbon dicarboxylic
acid with which acetyl CoA condenses is regenerated.
|Figure 6 A view of the oxidation of pyruvate. The oxidation of pyruvate generates three CO2, four NADH, and one
FADH2. The oxidation of NADH and FADH2 by the mitochondrial electron transport chain is exergonic and provides
most of the energy for ATP synthesis.
The mitochondrial inner membrane (Fig. 7) contains
proteins that act in concert to catalyze NADH and FADH2
oxidation by molecular oxygen. [See reactions (2) and (3)
above.] These reactions are carried out in many small steps
by proteins that are integral to the membrane and that undergo
oxidation–reduction. These proteins make up what
is called the mitochondrial electron transport chain. Components
of the chain include iron proteins (cytochromes
and iron–sulfur proteins), flavoproteins (proteins that contain
flavin), copper, and quinone binding proteins.
1960s that electron transport through the mitochondrial
chain is obligatorily linked to the movement of protons
across the inner membrane of the mitochondrion. In this
way, part of the energy liberated by oxidative electron
transfer is conserved in the form of the proton electrochemical
potential. This potential, ΔµH+
, is the sum of
contributions from the activity gradient and that of the
|⇒ Equation 
||ΔµH+ = RT ln ([H+]a/[H+]b) + FΔφ,
is the gas constant; T
, the absolute temperature; a
, the aqueous spaces bounded by the membrane; F
Faraday’s constant; and Δφ, the membrane potential. As
Mitchell suggested, the mitochondrial inner membrane is
poorly permeated by charged molecules, including protons.
The membrane thus provides an insulating layer
between the two aqueous phases it separates. Thus the
transport of protons across the membrane generates an
electrochemical potential. In the case of mitochondria, the
membrane potential is the predominant component of the
electrochemical of the proton. The total ΔµH+
respiring mitochondria is on the order of −200 mV, if one
uses the convention that the inside space bounded by the
membrane is negative.
Electron transport from NADH and FADH2
provides the energy for the generation of the electrochemical
potential of the proton. The flow of protons down this
potential is exergonic and is the immediate source of energy
for ATP synthesis. The proton-linked synthesis of
ATP is catalyzed by a complex enzyme called ATP synthase.
Remarkably similar enzymes are located in the coupling
membranes of bacteria, mitochondria, and chloroplasts,
the intracellular sites of photosynthesis in higher
plants. Even though the reaction that they catalyze seems
relatively straightforward (see Fig. 2), the ATP synthases
contain a minimum of 8 different proteins and a total of
about 20 polypeptide chains.
ATP is formed in the aqueous space bounded by the mitochondrial
inner membrane. This space is known as the
matrix (see Fig. 7). Most of the ATP generated within mitochondria
is exported to the cytoplasm where it is used to
drive energy-dependent reactions. The ADP and Pi
in the cytoplasm must then be taken up by the mitochondria.
The inner membrane contains specific proteins that
mediate the export of ATP and the import of ADP and
. One transporter catalyzes counterexchange transport
of ATP out of the matrix with ADP in the cytoplasm into
the matrix (Fig. 8). At physiological pH, ATP bears four
negative charges, and ADP, three. Thus, the one-to-one
exchange transport of ATP with ADP creates a membrane
potential that is opposite in sign of that created by electrontransport-
driven proton translocation. ATP/ADP transport
costs energy and the direction of transport is poised by
the proton membrane potential. In addition, phosphate
uptake into mitochondria is coupled to the electrochemical
proton potential. The phosphate translocator (see Fig. 8)
catalyzes the counterexchange transport of H2
hydroxide anion (OH−
). The outward movement of OH−
causes acidification of the matrix, whereas the direction
of proton transport driven by electron transport is out of
the mitochondrial matrix and results in an increase in the
pH of the matrix.
|Figure 7 Diagrams of the structures of mitochondria and chloroplasts. The inner membrane of mitochondria and
the thylakoid membrane of chloroplasts contain the electron transport chains and ATP synthases. Note that the
orientation of the inner membrane is opposite that of the thylakoid membrane.
|Figure 8 ATP, ADP, and Pi transport in mitochondria. ATP is
formed inside mitochondria. Most of the ATP is exported to the
cytoplasm where it is cleaved to ADP and Pi. The mitochondrial
inner membrane contains specific proteins that mediate not only
ATP release coupled to ADP uptake, but also Pi uptake linked to
hydroxide ion (OH−) release.
In the total oxidation of glucose to CO2
and water, six
are released and six O2
are reduced to water. For
each pyruvate oxidized, four NADH and one FADH2
generated. Since two molecules of pyruvate are derived by
means of glycolysis from one molecule of glucose, a total
of eight NADH and two FADH2
are formed by pyruvate
oxidation. Four electrons are required for the reduction
to two molecules of H2
O. Thus, pyruvate oxidation
accounts for the reduction of five of the six molecules of
in the complete oxidation of glucose. The sixth O2
reduced to water by electrons from the NADH formed by
the oxidation of triose phosphate in glycolysis.
Fermentation, or anaerobic glycolysis, yields but 2 mol
of ATP per 1 mol of glucose catabolized. In contrast, complete
oxidation of glucose to CO2
and water yields about
15 times more ATP. Thus, it is understandable why yeasts
and some bacteria consume more glucose under anaerobic
conditions than when oxygen is present.
In animals, glucose is normally completely oxidized.
During strenuous exercise, however, the demand for oxygen
by muscle tissues can outstrip its supply and the tissue
may become anaerobic. Muscle contraction requires
ATP, and rapid breakdown of glucose and its storage polymer,
glycogen, takes place under anaerobiosis. Glycolysis
would stop quickly if the NADH produced by the oxidation
of triose phosphate were not converted back to NAD+
In muscle cells under O2
-limited conditions, pyruvate is
reduced by NADH to lactic acid (see Fig. 5), a source
of muscle cramps during exercise. At rest, lactic acid is
converted back to glucose in the liver and kidneys and
returned to muscle tissues where it stored in the form of