GTP is converted to ATP by nucleoside-diphosphate
kinase:
GTP + ADP ^ GDP + ATP
Succinyl-CoA can also be synthesized from propionyl-
CoA by way of methylmalonyl-CoA, which is formed in
the oxidation of branched-chain amino acids (e.g., valine,
isoleucine) and in the terminal stage of oxidation of odd-
chain-length fatty acids (Chapter 18). Succinyl-CoA is
utilized in the activation of acetoacetate (Chapter 18) and
the formation of <$-aminolevulinate, a precursor of pro-
phyrin (Chapter 29).
D eh yd ro g en a tio n o f S u ccin a te to F u m a ra te
Succinate is dehydrogenated to the trans-unsaturated
compound fumarate by succinate dehydrogenase, an FAD
enzyme:
C H 2C O C T
H C C O C T
|
+ E — F A D -------►
II
+ E — FA D H
2
C H
2
C O O “
“ O O C C H
S u c c in a te
F u m a ra te
This reaction is the only dehydrogenation reaction of the
TCA cycle in which NAD+ does not mediate the trans-
port of reducing equivalents. The FAD is covalently linked
to a histidyl residue of the enzyme, which is an inte-
gral protein of the inner mitochondrial membrane and
contains iron-sulfur centers that undergo redox changes
(Fe3++ e_ ^ Fe2+). Reducing equivalents from FADH
2
enter the electron transport chain at the coenzyme Q level,
bypassing one of the sites of oxidative phosphorylation
(Chapter 14). The enzyme is stereospecific for the trans
hydrogen atoms of the methylene carbons of the substrate,
so that only the trans isomer is produced (the cis isomer is
maleate).
Succinate dehydrogenase is competitively inhibited
by malonate, the next lower homologue of succinate
(Chapter
6
).
H yd ra tio n o f F u m a ra te to M a la te
In this reaction, water is added across the double bond
of fumarate by fumarate hydratase (fumarase) to yield the
hydroxy compound L-malate:
H C — C O O “
H O — C H — C O O “
II
+ H
2
0 * ±
|
“ O O C — C H
C H 2— C O O H
F u m a ra te
i-M a la te
The enzyme has absolute specificity for the double bond
of the tran.v-unsaturated acid and for the formation of
244
L-hydroxy acid. The reaction is freely reversible (near-
equilibrium).
D eh yd ro g en a tio n o f M a la te to O x a lo a ceta te
In the last reaction of the cycle, L-malate is oxidized to
oxaloacetate by malate dehydrogenase, an NAD+-linked
enzyme:
H O — C H C O O "
|
+NAD+?±
C H
2
— C O O "
i-M a la te
0
II
+
C — C O O “
+ NADH + H
I
C H
2
— C O O "
O x a lo a c e ta te
Although the equilibrium of this reaction favors malate
formation,
in vivo
the reaction proceeds toward the for-
mation of oxaloacetate, since the latter is rapidly removed
by the citrate synthase reaction to initiate the next round
of the cycle.
S tereo ch em ica l A sp ects o f th e TCA C ycle
Aconitate dehydratase, succinate dehydrogenase, and
fumarase yield stereospecific products. Labeling experi-
ments with !4C in methyl and carboxyl carbons of acetyl-
CoA or in all of the carbons of oxaloacetate yield the
following results in terms of the intermediates or product
formed.
When only labeled acetyl-CoA is used, the label does
not appear in CO
2
during the first revolution of the cy-
cle, and upon completion of the first revolution, two of
the four carbon atoms of oxaloacetate are labeled. Thus,
during the first revolution, neither carbon atom of acetyl-
CoA is oxidized to CO
2
and the carbon atoms of the CO
2
produced are derived from oxaloacetate. This finding is
due to discrimination of the two -CH
2
COO" groups of
citrate by aconitate dehydratase, so that citrate is treated
as a chiral molecule at the asymmetrical active center of
the enzyme. However, during the succeeding revolutions
of the cycle, the label is randomized because succinate, a
symmetrical compound, is treated as such by the enzyme
without discriminating between its two carboxyl groups.
If only labeled oxaloacetate is used, half of the label is re-
tained in the oxaloacetate at the end of the first revolution
of the cycle.
A m p h ib o lic A sp ects o f th e T C A C ycle
At each turn of the TCA cycle, oxaloacetate is re-
generated and can combine with another acetyl-CoA
chapter 13
Carbohydrate Metabolism I: Glycolysis and TCA Cycle
