TCA cycle

tricarboxylic acid (TCA) cycle/Krebs cycle/citric acid cycle

BC3.3
TCA cycle
Central metabolic hub of the cell
• gateway to the aerobic metabolism of any molecule that can be
transformed into an acetyl group or dicarboxylic acid
• important source of precursors for the storage forms of fuels and also
for amino acids, nucleotide bases, cholesterol, and porphyrin
• accounts for more than two-thirds of the adenosine triphosphate
(ATP) generated from fuel oxidation.
• The TCA cycle requires a large number of vitamins and minerals to
function. These include niacin (NAD), riboflavin (flavin adenine
dinucleotide [FAD] and flavin mononucleotide [FMN]), pantothenic
acid (coenzyme A), thiamine, Mg2+, Mn2+,Ca2+, Fe2+, and phosphate.
The Formation of Acetyl Coenzyme A from
Pyruvate
• Under aerobic conditions, the pyruvate is transported into
mitochondria in exchange for OH- by the pyruvate carrier, an
antiporter.
• In the mitochondrial matrix, pyruvate is oxidatively decarboxylated by
the pyruvate dehydrogenase complex to form acetyl CoA. This is an
irreversible reaction. High transfer-potential electrons in the form of
NADH are captured.
 Pyruvate dehydrogenase complex (PDC)

Electron micrograph
image of PDH complexes
The conversion of pyruvate into acetyl CoA
consists of three steps: decarboxylation, oxidation,
and transfer of the resultant acetyl group to CoA.
Pyruvate Dehydrogenase Complex
(1) Pyruvate combines with TPP and is then
decarboxylated by Pyruvate dehydrogenase
component (E1) to form the hydroxyethyl TPP.
(2) The dihydrolipoyl arm of E2 moves into the
active site of E1.
(3) the hydroxyethyl group attached to TPP is
oxidized to form an acetyl group and
concomitantly transferred to lipoamide.
(4) Dihydrolipoyl transacetylase (E2) catalyzes
the transfer of the acetyl moiety to CoA to form
the product acetyl CoA. The dihydrolipoamide
arm then swings to the active site of E3.
(5,6) By the action of dihydrolipoyl
dehydrogenase (E3) the oxidized form of
lipoamide is regenerated through the transfer of
the protons and electrons to FAD and then to
NAD+
The Pyruvate Dehydrogenase Complex Is Regulated
Allosterically and by Reversible Phosphorylation
• High concentrations of reaction products of the complex inhibit the
reaction: acetyl CoA inhibits the transacetylase component (E2), whereas
NADH inhibits the dihydrolipoyl dehydrogenase (E3).
• Pyruvate dehydrogenase complex activity is controlled principally through
phosphorylation by pyruvate dehydrogenase kinase, which inhibits the
enzyme and dephosphorylation by pyruvate dehydrogenase phosphatase,
which activates it.
• Adrenergic agonists stimulate pyruvate dehydrogenase by triggering a rise
in the cytosolic Ca2+ level, which in turn elevates the mitochondrial Ca2+
level. The rise in mitochondrial Ca2+ activates the pyruvate dehydrogenase
complex by stimulating the phosphatase.
• Insulin also actives pyruvate dehydrogenase by stimulating the
dephosphorylation of the complex.
REGULATION OF PDC
The Citric Acid Cycle Has
Eight Steps
1. Formation of Citrate by the enzyme Citrate
Synthase

The CoA liberated in this reaction is

recycled to participate in the
oxidative decarboxylation of another
molecule of pyruvate by the PDH
complex.
 2. Formation of Isocitrate via cis-Aconitate by the
enzyme aconitase
Aconitase contains an iron-sulfur center, which acts both in the binding of the
substrate at the active site and in the catalytic addition or removal of H2O.
3. Oxidation of Isocitrate to α-Ketoglutarate and
CO2 by the enzyme isocitrate dehydrogenase
• first of four oxidation-reduction reactions in the citric acid cycle.
• oxidative decarboxylation
• Mn2+ is present in the active site of the enzyme.
• There are two different forms of isocitrate dehydrogenase in all cells, one
requiring NAD as electron acceptor and the other requiring NADP. NAD-
dependent enzyme occurs in the mitochondrial matrix and serves in the
citric acid cycle. The main function of the NADP-dependent enzyme, found
in both the mitochondrial matrix and the cytosol, may be the generation of
NADPH, which is essential for reductive anabolic reactions.
• The rate of formation of α-ketoglutarate is important in determining the
overall rate of the cycle
3. Oxidation of Isocitrate to α-Ketoglutarate and
CO2 by the enzyme isocitrate dehydrogenase
4. Oxidation of -Ketoglutarate to Succinyl-CoA
and CO2 by α-ketoglutarate dehydrogenase
• second oxidative decarboxylation reaction

• This reaction is virtually identical to the pyruvate dehydrogenase

reaction discussed above, and the α-ketoglutarate dehydrogenase
complex closely resembles the PDH complex in both structure and
function.
5. Conversion of Succinyl-CoA to Succinate by
succinyl CoA synthetase (succinate thiokinase).
• substrate-level phosphorylation
• Succinyl CoA is an energy-rich thioester compound. The cleavage of
the thioester bond is coupled to the phosphorylation of GDP.
6. Oxidation of Succinate to Fumarate by
succinate dehydrogenase
• succinate dehydrogenase is tightly bound to the inner mitochondrial
membrane.
• contains three different iron-sulfur clusters and one molecule of covalently
bound FAD
• FAD is nearly always the electron acceptor in oxidations that remove two
hydrogen atoms from a substrate.
• succinate dehydrogenase is directly associated with the electron-transport chain,
the link between the citric acid cycle and ATP formation.
7. Hydration of Fumarate to Malate by
fumarase
8. Oxidation of Malate to Oxaloacetate by L-
malate dehydrogenase
TCA cycle
SUMMARY OF THE TCA CYCLE
ENERGETICS
REGULATION OF TCA
CYCLE
• The Pyruvate Dehydrogenase
Complex Is Regulated Allosterically
and by Reversible Phosphorylation
(discussed in the previous slides).
• The rate of the citric acid cycle is
precisely adjusted to meet an
animal cell's needs for ATP.
• The primary control points are the
allosteric enzymes isocitrate
dehydrogenase and α-
ketoglutarate dehydrogenase.
Anaplerotic Reactions Replenish Citric Acid
Cycle Intermediates
• As intermediates of the citric acid cycle are removed to serve as
biosynthetic precursors, they are replenished by anaplerotic reactions.
• The most important anaplerotic reaction in mammalian liver and
kidney is the reversible carboxylation of pyruvate by CO2 to form
oxaloacetate, catalyzed by pyruvate carboxylase. When the citric acid
cycle is deficient in oxaloacetate or any other intermediates, pyruvate is
carboxylated to produce more oxaloacetate.
• Pyruvate carboxylase is a regulatory enzyme and is virtually inactive in
the absence of acetyl-CoA, its positive allosteric modulator.
Major
anaplerotic
pathways of
the
tricarboxylic
acid (TCA)
cycle.
ROLE OF TCA CYCLE IN
ANABOLISM
Clinical correlate: The Disruption of Pyruvate
Metabolism Is the Cause of Beriberi and Poisoning
by Mercury and Arsenic
Which biochemical processes might be affected by a deficiency of
thiamine? Thiamine pyrophosphate is the prosthetic group of three
important enzymes: pyruvate dehydrogenase, α-ketoglutarate
dehydrogenase, and transketolase.
Symptoms similar to those of beriberi arise if an organism is exposed to
mercury or arsenite. Both elements have a high affinity for neighboring
sulfhydryls, such as those in the reduced dihydrolipoyl groups of the
dihydrolipoyl dehydrogenase component of the pyruvate
dehydrogenase complex. The binding of mercury or arsenite to the
dihydrolipoyl groups inhibits the complex and leads to central nervous
system pathologies.