Chapter 16

The Citric Acid Cycle: CAC

Kreb’s Cycle
Tricarboxylic Acid Cycle: TCA
 Overall Picture

Acetyl-CoA production
The area blocked off all occurs in the
takes place in the mitochondria.
Mitochondrion. So, first
pyruvate has to get Acetyl-CoA enters the
transported from the CAC.
cytoplasm into the
mitochondrion.
In this Figure, only
Glycolysis is in the
Cytoplasm.
 The Citric Acid Cycle
Key topics: To Know
– Also called Tricarboxylic Acid Cycle (TCA) or
Krebs Cycle. Three names for the same thing.
– Cellular respiration and intermediates for
biosynthesis.
– Conversion of pyruvate to activated acetate
– Reactions of the citric acid cycle
– Anaplerotic reactions to regenerate the acceptor
– Regulation of the citric acid cycle
– Conversion of acetate to carbohydrate precursors
in the glyoxylate cycle
Discovered CAC in Pigeon Flight Muscle
 Cellular Respiration

• Process in which cells consume O2 and produce CO2

• Provides more energy (ATP) from glucose than Glycolysis
• Also captures energy stored in lipids and amino acids
• Evolutionary origin: developed about 2.5 billion years ago
• Used by animals, plants, and many microorganisms
• Occurs in three major stages:
- acetyl CoA production (This chapter)
- acetyl CoA oxidation (This chapter)
- electron transfer and oxidative phosphorylation
(Next Chapter)
 Pyruvate Dehydrogenase

Model TEM
 Pyr DH is a Complex Enzyme

There is a lot of stuff going on here. Overall a nicely exothermic enzyme. We

already know about NAD+, TPP. FAD. So lets look at lipoic acid. PyrDH is really
three enzymes as a module
One Unit of Pyr DH
Lipoic Acid is linked to a Lys (K)

You don’t have to know the structure in detail,

the important point is the disulfide (oxidized
state) and –SH (reduced state) and that it can
form a thioester (just like acetyl-S-CoA
 Remember HSCoA?

It is down here
Pyr DH is a Cool Enzyme
The citric acid cycle has eight stages:
1. Production of citrate from oxaloacetate and acetyl CoA (catalyzed
by citrate synthase).
2. Isomerization of citrate to isocitrate (catalyzed by aconitase).
3. Oxidation of isocitrate to -ketoglutarate (catalyzed by isocitrate
dehydrogenase; the reaction requires NAD+)
4. Oxidation of -ketoglutarate to succinyl CoA (catalyzed by the -
ketoglutarate dehydrogenase complex; the reaction requires
NAD+).
5. Conversion of succinyl CoA to succinate [catalyzed by succinyl CoA
synthetase; the reaction requires inorganic phosphate and GDP (or
ADP)].
6. Oxidation of succinate to fumarate (catalyzed by succinate
dehydrogenase; the reaction involves FAD).
7. Hydration of fumarate to malate (catalyzed by fumarase).
8. Oxidation of malate to oxaloacetate (catalyzed by malate
dehydrogenase; the reaction requires NAD+).
 Citrate Synthase
Convention to write
incoming Acetyl on
Top

EOC Problem 32, further on the

thermodynamics of Citrate
Synthase.
Aconitase, the Ferris Wheel
The Aconitase Iron Sulfur Complex
 Aconitase has More than One Role

Mitochondrial aconitase: Citric Acid Cycle

Cytosolic aconitase: 2 roles:
1. citrate  isocitrate
2. iron response regulator
 Aconitase binding iron/RNA

To become an iron
response regulator,
aconitase changes it
shape (due to lack of iron)
so it can bind RNA.
 Isocitrate DH

ΔGo’ = -21 kJ/mole

Mn++ cofactor

EOC Problem 8 is all about IsocitDH.

αKG DH is Just Like Pyr DH

TPP,
lipoate
FAD
Succinyl CoA Synthetase : Substrate Level Phosphorylation

One GTP = One ATP Nucleoside diphosphate kinase:

GTP + ADP  GDP + ATP ΔGo’ = 0

Succinate DH = Old Yellow
Malonate was One of the First Competitive
Inhibitors Known
Fumarase: the addition of water in two parts
Don’t Confuse Malate and Maleate
Malate DH is Endothermic
CAC Energetics
 Watch Where the Label Goes

EOC Problem 18: Labeled glucose carbons

and where they go in CAC.
Citrate is Prochiral
 The Acetyl Portion does not get oxidized to CO2
Until the Second Round

And it gets randomized at Succinate

 Energi Yield
For each turn of the cycle, 12 ATP
molecules are produced …….
one directly from the cycle and
11 from the re-oxidation of the three NADH
and one FADH2 molecules produced
by the cycle by oxidative
phosphorylation
Energetics of Glycolysis and CAC in ATPs
CAC in Anaerobic Not-Respiratory Organisms

It’s a 2 input
FORK
This is Why
Amino acids, purines and pyrimidines, fatty acids and glucose
are all synthesized by pathways that use citric acid
intermediates as precursors.
Anaplerotic Reactions
 Regulation of CAC

The regulation of CAC (all

allosteric) is at the drivers of the
cycle (exothermic reactions).
Both positive (green) and
negative (red) regulation:
molecules that signal low energy
level signal the cycle to go
faster; molecules that signal
energy levels high signal the
cycle to slow down.
• The citric acid cycle is regulated at the steps catalyzed by
citrate synthase, isocitrate dehydrogenase and -
ketoglutarate dehydrogenase via feedback inhibition by
ATP, citrate, NADH and succinyl CoA, and stimulation of
isocitrate dehydrogenase by ADP.
• Pyruvate dehydrogenase, which converts pyruvate to
acetyl CoA to enter the cycle, is inhibited by acetyl CoA
and NADH.
• In addition, this enzyme is inactivated by phosphorylation,
a reaction catalyzed by pyruvate dehydrogenase kinase.
• A high ratio of NADH/NAD+, acetyl CoA/CoA or ATP/ADP
stimulates phosphorylation of pyruvate dehydrogenase and
so inactivates this enzyme.
• Pyruvate inhibits the kinase.
• Removal of the phosphate group (dephosphorylation) by a
phosphatase reactivates pyruvate dehydrogenase.
Pathway Proteins Form Functional Units but
It’s Concentration Dependent
 Pathways are Protein Modules

Flagella
LPS
Outer Membrane
Peptidoglycan
ATPase
Cytoplasmic
Membrane
Glycolysis
RNA
 In Animals CAC can not be used for
Gluconeogensis from Ac-SCoA

D, N, L, K,
M, T, I

Porphrins:
heme
(cytochromes, E, Q, P, R
hemoglobin),
chlorophyll
 In Bacteria and Plants, Not Vertebrates

Overall:
2 Ac-SCoA  Succinate
Succinate  OAA

NADH and FADH2

Oxaloacetate
CAC
Glyoxylate Cycle in Plants in a Membrane Body
 Linkage to
Gluconeogenesis
in Plants
Regulation
Linkage
 Things to Know and Do Before Class
1. Pyruvate DH…all three parts and cofactors.
2. Chemistry of each step in Citric Acid Cycle.
3. Overall CAC thermodynamics (which steps are at Eq and
which are drivers.
4. Prochiral nature of citrate.
5. Amphibolic nature of CAC and why fermenters need
almost all of CAC.
6. Importance of anaplerotic reactions and how they work.
7. Glyoxylate Cycle (mammals lack) but plants, some
invertebrates and bacteria have it. What does it do?
8. EOC Problems 1-9, 16, 18, 19, 30-32.