BIO462

THE CITRIC ACID CYCLE

Learning Outcome
Upon completion of this chapter, students should
be able to:
i. explain the steps of the citric acid cycle
ii. differentiate between citric acid cycle and
glyoxylate cycle
iii. relate citric acid cycle as an energy source

Week 8
Lecture Outline

1) The Central Role of he Citric Acid Cycle in

Metabolism
2) The Overall Pathway of the Citric Acid Cycle
3) How Pyruvate is Converted to Acetyl-CoA
4) The Individual Reactions of the Citric Acid Cycle
5) Energetics and Control of the Citric Acid Cycle
6) The Glyoxylate Cycle: A Related Pathway
7) The Citric Acid Cycle in Catabolism
8) The Citric Acid Cycle in Anabolism
9) The Link to Oxygen
The Central Role of the Citric Acid Cycle
• Three processes play central roles in aerobic
metabolism
• The citric acid cycle
• Electron transport
• Oxidative phosphorylation
• Metabolism consists of:
• Catabolism: the oxidative breakdown of nutrients
• Anabolism: the reductive synthesis of biomolecules
• The citric acid cycle is amphibolic; that is, it plays a
role in both catabolism and anabolism. It is the
central metabolic pathway.
 Catabolism Anabolism
Polysac-
Fats charides Proteins
Catabolism Excretion
beakdown
of larger
molecules Fatty acids Monosac- Amino
Products of anabolism,
to smaller and glycerol charides Acids
including proteins and
ones nucleic acids
S mall Anabolism energy and
molecules of proteins reducing
oxidation and the agents
release of energy S ome nutrients and
Excretion Anabolism products of catabolism
The role of ATP as an energy source
The Central Relationship of the Citric Acid
Cycle to Catabolism

The central relationship of the citric acid cycle to catabolism. Amino acids, fatty
acids, and glucose can all produce acetyl-CoA in stage 1 of catabolism.

In stage 2, acetyl-CoA enters the citric acid cycle. Stages 1 and 2 produce reduced
electron carriers (shown here as e–).

In stage 3, the electrons enter the electron transport chain, which then produces
ATP.
Where does the Citric Acid Cycle Take
Place?
• In eukaryotes, cycle takes place in the mitochondrial matrix
Features of Cycle

An overview of the citric acid cycle.

Note the names of the enzymes.

The loss of CO2 is indicated, as is the

phosphorylation of GDP to GTP.

The production of NADH and FADH2 is

also indicated.

Note
NAD – electron acceptor.
- reduced to NADH
FAD – intermediate electron acceptor.
- flavine adenine dinucleotide
- derived from riboflavin (vit B2)
- takes up 2 e- and 2 H+ to produce FADH
oxidative
decarboxylation
• In the citric acid cycle and the pyruvate
dehydrogenase reactions, one molecule of
pyruvate is oxidized to 3 molecules of CO2 as a
result of oxidative phosphorylation.

• The oxidations are accompanied by reductions.

• 4 NAD+ are reduced to NADH
• 1 FAD is reduced to FADH2
• 1 GDP is phosphorylated to GTP
How Pyruvate is Converted to Acetyl-CoA
• Pyruvate dehydrogenase complex is responsible for the
conversion of pyruvate to CO2 and the acetyl portion of
acetyl-CoA

• Five enzymes in complex: pyruvate dehydrogenase,

dihydrolipoyl transacetylase, dihydrolipoyl dehydrogenase,
pyruvate dehydrogenase kinase, pyruvate dehydrogenase
phosphatase
Pyruvate is Converted to Acetyl-CoA
(Cont’d)
• First, pyruvate loses CO2 and hydroxyethylTPP (HETPP) is
formed
• In the second step, the active form of lipoic acid is bound to
the enzyme, dihydrolipoyl transacetylase, by an amide bond
to the -amino group of a lysine
• The hydroxyethyl group (HE) is oxidized and transferred to a
sulfur atom of the reduced form of lipoamide
• Lipoamide is reduced to dihydrolipoamide
• In step 3, the acetyl group is transferred to the sulfhydryl group
of coenzyme A
• Next, dihydrolipoamide is oxidized to lipoamide
The Mechanism of the Pyruvate
Dehydrogenase Complex
The mechanism of the
pyruvate dehydrogenase reaction.

• Decarboxylation of pyruvate occurs with formation of

hydroxyethyl-TPP (Step 1).

• Transfer of the two-carbon unit to lipoic acid in Step 2

is followed by formation of acetyl-CoA in Step 3.
Lipoic acid is reoxidized in Step 4 of the reaction.
Summary
• The two-carbon unit needed at the start of the citric
acid cycle is obtained by converting pyruvate to
acetyl-CoA

• This conversion requires the three primary enzymes

of the pyruvate dehydrogenase complex, as well as,
the cofactors TPP, FAD, NAD+, and lipoic acid

• The overall reaction of the pyruvate dehydrogenase

complex is the conversion of pyruvate, NAD+, and
CoA-SH to acetyl-CoA, NADH + H+, and CO2
Individual Reactions of the Citric Acid Cycle
• In step 1, there is a condensation of acetyl-CoA with
oxaloacetate to form citrate
• G°’ = -32.8 kJ•mol-1, therefore, the reaction is exergonic

• Reaction is catalyzed by citrate synthase, an allosteric

enzyme that is inhibited by NADH, ATP, and succinyl-CoA
Individual Reactions of the Citric Acid Cycle
(Cont’d)
• In step 2, citrate is isomerized to isocitrate. The
reaction is catalyzed by aconitase

• Citrate is achiral; it has no stereocenter

• Isocitrate is chiral; it has 2 stereocenters and 4

stereoisomers are possible

• There is only one of the 4 stereoisomers of isocitrate

formed in the cycle
The Reactions of the Citric Acid Cycle
Individual Reactions of the Citric Acid Cycle
(Cont’d)
Individual Reactions of the Citric Acid Cycle
(Cont’d)
• In step 3, there is an
oxidation of isocitrate
followed by decarboxylation
to form -ketoglutarate and
CO2

• The reaction is catalyzed by

isocitrate dehydrogenase,
an allosteric enzyme, which
is inhibited by ATP and
NADH, and activated by
ADP and NAD+
Individual Reactions of the Citric Acid Cycle
(Cont’d)
• In step 4, there is an oxidative decarboxylation of -
ketoglutarate to succinyl-CoA

• This reaction is catalyzed by the -ketoglutarate

dehydrogenase complex, which is, like pyruvate
dehydrogenase, a multienzyme complex and requires
coenzyme A, thiamine pyrophosphate, lipoic acid, FAD, and
NAD+
Individual Reactions of the Citric Acid Cycle
(Cont’d)
• Next in step 5, the thioester bond of succinyl-CoA if
hydrolyzed in the formation of succinate
• The two CH2-COO- groups of succinate are equivalent
• This is the first energy-yielding step of the cycle
• The overall reaction is slightly exergonic
Individual Reactions of the Citric Acid Cycle
(Cont’d)
• Next, there is an oxidation of succinate to fumarate

• Then, the hydration of fumarate to L-malate occurs

Individual Reactions of the Citric Acid Cycle
(Cont’d)
• Then step 8 (regeneration of oxaloacetate),
• malate is oxidized to Oxaloacetate
Oxidation of Pyruvate Forms CO2 and ATP
Summary
• In the citric acid cycle and the pyruvate
dehydrogenase reaction, one molecule of pyruvate is
oxidized to three molecules of CO2 as a result of
oxidative decarboxylation

• The oxidations are accompanied by reductions

involving NAD+ to NADH, FAD to FADH2

• GDP is phosphorylated to GTP

Energetics and Control of the Citric Acid Cycle
• There are 3 points of control within the cycle:
• Citrate synthase: inhibited by ATP, NADH, and
succinyl CoA; also product inhibition by citrate
• Isocitrate dehydrogenase: activated by ADP and
NAD+, inhibited by ATP and NADH
• -ketoglutarate dehydrogenase complex: inhibited
by ATP, NADH, and succinyl CoA; activated by ADP
and NAD+
• There is one control point outside the cycle
• Pyruvate dehydrogenase: inhibited by ATP and
NADH; also product inhibition by acetyl-CoA
Energetics and Control of the Citric Acid Cycle
(Cont’d)
Control of the Citric Acid Cycle (Cont’d)
Control of the Citric Acid Cycle (Cont’d)
The Glyoxylate Cycle: A Related Pathway
• In plants and some bacteria, there may be a
modification of the citric acid cycle to produce four-
carbon dicarboxylic acids and eventually glucose
• The glyoxylate cycle bypasses the two oxidative
decarboxylations of the citric acid cycle
• Instead, it routes isocitrate via glyoxylate to malate
• Key enzymes in this cycle are isocitrate lyase and
malate synthase
Unique Reactions of the Glyoxylate Cycle
The Glyoxylate Cycle (Cont’d)

This pathway results in the net

conversion of two acetyl-CoA
to oxaloacetate.
(All the reactions are shown in purple).

The unique reactions of the

glyoxylate cycle are shown
with a light green highlight in
the center of the circle.
The Glyoxylate Cycle (Cont’d)
• The glyoxylate cycle takes place:
• In plants: in glyoxysomes, specialized organelles
devoted to this cycle
• In yeast and algae: in the cytoplasm

• Helps plants grow in the dark:

• Seeds are rich in lipids, which contain fatty acids
• During germination, plants use the acetyl-CoA
produced in fatty acid oxidation to produce
oxaloacetate and other intermediates for carbohydrate
synthesis
• Once plants begin photosynthesis and can fix CO2,
glyoxysomes disappear
The Citric Acid Cycle in Catabolism
• The catabolism of proteins, carbohydrates, and fatty
acids all feed into the citric acid cycle at one or more
points

Note:

PEP is phosphoenolpyruvate;
α-KG is α-ketoglutarate;
TA is transamination;
→→→ is a multistep pathway.
Summary
• All metabolic pathways are related, and all of them
operate simultaneously

• In catabolic pathways, nutrients, many of which are

macromolecules, are broken down to smaller
molecules, such as sugars, fatty acids, and amino
acids

• Small molecules are processed further, and the end

products of catabolism frequently enter the citric acid
cycle, which plays a key role in metabolism
The Citric Acid Cycle in Anabolism
• The citric acid cycle is the source of starting materials
for the biosynthesis of other compounds

• If a component of the citric acid cycle is taken out for

biosynthesis, it must be replaced

• oxaloacetate, for example, is replaced by the

carboxylation of pyruvate

• A reaction that replenishes a citric acid cycle

intermediate is called an anaplerotic reaction
The Citric Acid Cycle in Anabolism (Cont’d)
The Citric Acid Cycle in Anabolism (Cont’d)
Lipid Anabolism
• Lipid anabolism begins with acetyl-CoA and takes
place in the cytosol
• acetyl-CoA is produced mainly in mitochondria from
catabolism of fatty acids and carbohydrates
• an indirect transfer mechanism exists involving citrate

Citrate + CoA-SH + ATP -----> Acetyl-CoA + Oxaloacetate + ADP + Pi

• the oxaloacetate thus formed provides a means for

the production of the NADPH needed for biosynthesis
Lipid Anabolism (Cont’d)
• The the net effect of these two reactions is replacement of
NADH by NADPH
• While there is some NADPH produced by this means, its
principal source is the pentose phosphate pathway
• The anabolic reactions that produce amino acids and many
other biomolecules begin with CA cycle molecules that are
transported into the cytosol
Summary of Anabolism in the Citric Acid
Cycle
Summary
• The citric acid cycle plays a central role in anabolic
pathways as well as in catabolism

• Pathways that give rise to sugars, fatty acids, and

amino acids all originate with components of the citric
acid cycle
The Link To Oxygen
• The citric acid cycle is considered part of the aerobic
metabolic process because of its link to the electron
transport chain and oxidative phosphorylation

• NADH and FADH2, two important cofactors

generated by the citric acid cycle, ultimately pass
their electrons to oxygen