Study Unit 10:

Cellular Respiration- Harvesting energy from

carbohydrates and other fuel
molecules

Dr. Rebamang Mosa

Department of Biochemistry, Genetics and Microbiology
Agricultural Science Building, Office Number: 3-5
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E-mail: rebamang.mosa@up.ac.za
 Lecture 1: Learning Outcomes (LO)

1. Recognize how energy from the oxidation of food (e.g. glucose) is coupled to
ATP synthesis (the main purpose of cellular respiration) and write down the
reaction for the overall oxidation of glucose to carbon dioxide and water.

2. Distinguish between substrate-level phosphorylation and oxidative

phosphorylation in cellular ATP formation.

3. Recognize the importance of oxidation-reduction reactions (redox reactions)

in cellular respiration.

4. Name the cellular compartment where each of the four stages (glycolysis,
pyruvate oxidation, citric acid cycle and oxidative phosphorylation) of cellular
respiration occurs and describe the purpose of each stage.

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LO 1
Cellular Respiration
C6H12O6 + 6O2 → 6CO2 + 6H2O + energy (heat and ATP)
LO 1-4
Cellular Respiration:
Generating ATP to Fuel Cellular Work

(Cytoplasm) (Mitochondria) (Mitochondria) (Mitochondria)

Substrate-level phosphorylation Oxidative phosphorylation

LO 2
Formation of ATP by substrate level phosphorylation
Cells make ATP by two fundamentally different
mechanisms: substrate-level and oxidative
phosphorylation
1. Substrate level phosphorylation: ATP is formed
by a direct transfer of a phosphate group from
a “high energy” compound to ADP
ADP + X-Pi ATP + X
2. Oxidative phosphorylation: ATP is synthesized
by the ATP synthase (enzyme) using energy
from proton (H+) gradient. The proton gradient
results from the movement of high energy
electrons down the electron transport chain
(ETC).
ATP synthase
ADP + Pi ATP
High energy compound (X)-a compound with a functional group, often a phosphate group, whose
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free energy of transfer to another compound proceeds with a large negative ΔG.
LO 3 Oxidation Reactions of Cellular Respiration

• In cellular respiration, the

energy stored in glucose is
harnessed in electron
carriers as glucose is
oxidized into carbon
dioxide (See Fig 7.2a).
• NAD+ and FAD are
important electron carriers
in cellular respiration.

NAD+ + 2e- + 2H+ → NADH + H+ Oxidation is loss of electrons.

FAD + 2e- + 2H+ → FADH2 Reduction is gain of electrons.

NAD- Nicotinamide adenine dinucleotide

FAD- Flavin adenine dinucleotide
LO 3
Reduction Reactions of Cellular Respiration

• Oxygen is the final electron

acceptor in cellular
respiration.
• When oxygen is reduced, it
forms water.
• The original electron donor
in cellular respiration is
glucose, but electrons move
from one molecule to the
next during cellular
respiration via reduction
reactions.

ΔG = -686 kcal/mol)
LO 1-4 Free Energy Changes

• The figure shows three of the

four stages of cellular
respiration.
• Notice the production of
important electron carriers,
NADH and FADH2. The
reduced forms of these
carriers have high potential
energy and will be used in
the electron transport chain
to generate ATP for the cell-
oxidative phosphorylation
 Lecture 2: Learning Outcomes (LO)

5. Write down an overview of the 10 reactions of the glycolytic pathway, and list
the net products of the pathway.

6. Recognize the energy inputs and outputs of the glycolytic pathway and the
reactions where substrate-level phosphorylation occurs.

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LO 5&6
Glycolysis- glucose lysis or degradation

• Occurs in nearly all living cells

• Stepwise degradation of glucose

(and other monosaccharides)

• Basically anaerobic process- its

principal steps occur with no
requirement for O2
LO 5&6 Glycolysis

Glycolysis has phases:

1. the preparatory phase
(investment phase), where
energy (ATP) is consumed.
2. the cleavage phase, where
glucose (6C) is split into two
3C phosphorylated
molecules.
3. the oxidation and payoff-
phase, where NADH and
ATP are two of the products.
LO 5&6
Phase 1:
Energy-Consuming Reactions

ATP is hydrolyzed, and its

phosphate groups are attached
to glucose, “priming” the 6C
glucose to be cleaved into two
3C molecules
LO 5&6
Phase 2:
Splitting Glucose

6C phosphorylated keto form of glucose (Fructose) is converted

into two 3C phosphorylated molecules (glyceraldehyde-3-
phosphate, G3P)
LO 5&6
Phase 3:
Energy-Producing Reactions
G3P is oxidized to produce 2NADH; 4 ATPs are produced
via substrate phosphorylation
LO 5&6
At the End of Glycolysis
At the end of glycolysis:
• there is a net gain of
two ATP: phase 3
produced four ATP,
but two were
consumed in phase 1.
• Phase 3 also
produces two NADH.
These will be used in
the last reaction of
cellular respiration.
• Pyruvate is the end
product of glycolysis-
further oxidized in the
mitochondrion
LO 5&6
Reaction 1: Phosphorylation of glucose to glucose 6-phosphate

Nelson and Cox, 2017

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LO 5&6
Reaction 6: Oxidation and phosphorylation of glyceraldehyde 3-phosphate

• Dehydrogenation and phosphorylation of glyceraldehyde 3-phosphate

catalyzed by glyceraldehyde 3-phosphate dehydrogenase.
• The inorganic phosphate is from the cytosolic pool.
• High energy electrons are transferred from glyceraldehyde 3-phosphate
(Oxidized) to NAD+ to become NADH + H+ (Reduced).
• Phosphate group is added to C1 of glyceraldehyde 3-phosphate to produce
1,3-bisphosphoglycerate (high energy compound).

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Nelson and Cox, 2017
LO 5&6
Reaction 7: Formation of ATP by substrate level phosphorylation

• Transfer of a phosphate group from 1,3-bisphosphoglycerate (1,3-BPG) to

ADP to yield ATP and 3-phosphoglycerate, a reaction catalyzed by
phosphoglycerate kinase.

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Nelson and Cox, 2017
LO 6
Reaction 10: Formation of ATP by substrate level phosphorylation

• Transfer of a phosphate group from PEP to ADP to yield ATP and

pyruvate, a reaction catalyzed by pyruvate kinase

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LO 5&6
Important to note about the glycolytic pathway

a) Glucose is phosphorylated (1), isomerized (2), degraded to 2

x 3C molecules (4), oxidized to pyruvate (6 →10)
b) 2 ATP consuming steps (1, 3)
c) 2 ATP yielding steps (7, 10)
d) 3 irreversible steps (1, 3, 10)

Net products:
Glucose + 2NAD+ + 2ADP + 2Pi → 2Pyruvate + 2NADH + 2H+ + 2 ATP + 2H2O

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 Practice questions

1. Assume D-glucose is the only substrate for glycolysis and the enzyme catalyzing
reaction six of glycolysis is inhibited. What would happen to the concentration of
the glycolytic intermediates before and after the affected reaction?
2. The first step of glycolysis involves phosphorylation of D-glucose to glucose 6-
phosphate. Explain the importance of this phosphorylation

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 Lecture 3: Learning Outcomes (LO)

7. Write down the pyruvate oxidation reaction (as a link between glycolysis
and the citric acid cycle) and list the net products of the reaction.

8. Write down an overview of the 8 reactions of the citric acid cycle and list
the net products of the cycle.

9. Write down the oxidation reactions of the citric cycle and recognize the
release of the reduced electron carriers (NADH and FADH2) and loss of
CO2 in the process.

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LO 7-9
Mitochondria
Stages 2-4 of cellular
respiration take place in
the mitochondria:

Stage 1: glycolysis
(cytoplasm)
Stage 2: pyruvate oxidation
(mitochondria)
Stage 3: citric acid cycle
(mitochondria)
Stage 4: oxidative
phosphorylation
(mitochondria)
LO 7
Pyruvate Oxidation

• The pyruvate produced by

glycolysis still contains chemical
potential energy in its bonds.
• In the presence of oxygen,
pyruvate can be oxidized to
produce carbon dioxide (CO2)
and NADH and ultimately
acetyl-CoA.
• These reactions occur in the
mitochondrial matrix.
LO 7 Acetyl-CoA Synthesis
At the end of glycolysis, there are two molecules of pyruvate. Thus,
at the end of pyruvate oxidation, there are two molecules of CO2,
two molecules of NADH, and two molecules of acetyl-CoA for each
glucose molecule.

2 Pyruvate + 2NAD+ → 2CO2 + 2 Acetyl-CoA + 2NADH + H+

 LO 7-9
Citric Acid Cycle (CAC)
• The CAC is considered a cycle because
oxaloacetate is the first reactant in the
process and is regenerated at the end of
the process.

• The CAC completes the oxidation of glucose

into CO2. ATP and additional energy
management molecules (NADH and FADH2)
are also products of the citric acid cycle.
LO 7-9
CAC completely oxidizes acetyl-CoA

• The oxidation of acetyl-CoA

produces the carbon dioxide
we exhale.

• Chemical potential energy that

was stored in acetyl-CoA is then
stored in NADH and FADH2.

• The production of GTP is the

result of substrate-level
phosphorylation, similar to
reactions 7 & 10 of glycolysis.
LO 7-9
Reaction 1: Formation of citrate

• Condensation of acetyl-CoA with oxaloacetate form citrate, a

reaction catalyzed by citrate synthase.
• Activity of the cycle is dependent on the [oxaloacetate]-
limiting intermediate.

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LO 9 Reaction 3: Oxidation of isocitrate to α-ketoglutarate
and CO2

• Isocitrate dehydrogenase catalyzes the oxidative

decarboxylation of isocitrate to α-ketoglutarate.
• First reaction in the cycle where CO2 is released and NADH
produced.

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LO 9
Reaction 4: Oxidation of α-ketoglutarate to succinyl-CoA
and CO2

• α-ketoglutarate dehydrogenase complex catalyzes the

oxidative decarboxylation of α-ketoglutarate to succinyl-CoA
• Second reaction where CO2 is released and second NADH is
produced

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LO 7-9
Reaction 5: Conversion of succinyl-CoA to succinate
• Conversion of succinyl-CoA to succinate is catalyzed by succinyl-
CoA synthetase
• The linkage between the succinyl group and CoA is a high-
energy bond (thioester)
• The reaction couples hydrolysis of the thioester linkage to
formation of GTP (Guanine triphosphate)
• GTP can transfer a phosphate to ADP converting it into ATP-
substrate level phosphorylation

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LO 9
Reaction 6: Oxidation of succinate to fumarate
• Succinate dehydrogenase catalyzes oxidation of succinate to
fumarate
• Succinate dehydrogenase:
– integral protein of the mitochondrial inner membrane in
eukaryotes
– Complex II in the electron transport chain
– contains covalently bound FAD (coenzyme)
• Third oxidation reaction and FADH2 is produced

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LO 9
Reaction 9: Oxidation of malate to oxaloacetate

• NAD-linked malate dehydrogenase catalyzes the oxidation of

malate to oxaloacetate
– Regenerates oxaloacetate for citrate synthase
• Final oxidation reaction, and third reaction where NADH is
produced

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LO 7-9
Net products of the citric acid cycle

• The energy of oxidations in the cycle is efficiently conserved

• Energy released by
oxidation is conserved
in the production of:
– 3 NADH
– 1 FADH2
– 1 ATP

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Fig 16-14: Products of one turn of the citric acid cycle.
 Practice questions

1. Is it possible to get a net synthesis of oxaloacetate by adding acetyl-CoA to

an extract of mitochondria that contains only the enzymes and cofactors of
the citric acid cycle? Explain your answer.
2. Why is the citric acid cycle considered part of aerobic metabolism, even
though molecular oxygen (O2) does not appear in any of its reactions?

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 Lecture 4: Learning Outcomes (LO)

10. Understand that oxidative phosphorylation consists of an electron transport

chain and ATP synthase. Consider the electron transfer system as a solid-
state biological electric circuit powered by NADH and FADH2.

11. Understand how energy released from electron transport is coupled to ATP
synthesis by explaining the electrochemical proton gradient process
(chemiosmosis) that drives the ATP-synthase motor in the mitochondrion.

12. Calculate the net ATP yield from the complete oxidation of n (a given
number of) glucose molecules to CO2 and H2O.

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LO 10-12 Oxidative Phosphorylation
• High energy electron carriers (NADH and FADH2) produced at
any stage in respiration converge at the electron transport
chain (ETC)- NADH and FADH2 are not oxidized directly by O2,
but via several oxidation-reduction reactions catalyzed by
electron carriers which make up the respiratory chain (ETC).
• Free energy is released as electrons are transferred along the
ETC from an electron donor (NADH and FADH2) to an electron
acceptor (O2)
• ETC is coupled to oxidative phosphorylation- the redox
reactions of the ETC convert the energy in the electrons to
potential energy used to synthesize ATP

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LO 10-12 Where do NADH and FADH2 go next in cellular
respiration?

NADH and FADH2 produced through redox reactions in the four

stages of cellular respiration will transfer electrons to other carriers
in the electron transport chain. Oxygen acts as the final electron
acceptor.
LO 10-12
Electron Transport Chain

Electrons are transferred from energy storage molecules to proteins

in the ETC in a series of redox reactions that helps build a proton
gradient that stores potential energy to be used for ATP production.
LO 10-12 Electron Transport Chain: The Proton Gradient
e- of NADH enter the ETC at complex I
NADH + H+ → NAD+ + 2e- + 2H+

e- of FADH2 enter the ETC at complex II (Succinate dehydrogenase to Q)

FADH2 → FAD + 2e- + 2H+

Protons flow down their concentration gradient and power ATP

synthase by converting one form of energy into energy in bonds of ATP.
LO 10-12
Electron Transport Chain: ATP Synthase

Protons flow through ATP synthase, and the potential energy in the
proton gradient is converted to the potential energy found in ATP.
LO 11
ATP Synthase

• Potential energy stored in the

proton gradient is converted
to mechanical (kinetic) energy
as protons move down their
concentration gradient
through ATP synthase.
• The change in conformation
of the subunit in the
mitochondrial matrix helps
catalyze the reaction,
converting ADP and Pi to ATP.
LO 10-12 The Flow of Energy in Cellular Respiration

• The energy of the electron carriers is transformed into energy stored

in a proton electrochemical gradient.
• ATP synthase then converts the energy of the proton gradient to
rotational energy, which drives the synthesis of ATP.
LO 12 Theoretical ATP Yield
Electron Number of H+ Number of H+ Number of ATP produced
carrier transported across needed for ATP
the inner membrane synthesize
NADH 10 4 10/4 = 2.5 ATP/NADH mol
FADH2 6 4 6/4 = 1.5 ATP/FADH2 mol

Electron flow through the complexes is highly exergonic and powers the transport of
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protons across the inner mitochondrial membrane
 Lecture 5: Learning Outcomes (LO)

13. Describe the glucose fermentation process (to lactic acid or ethanol) and its
biological significance.

14. Recognize that the breakdown products from the degradation of

carbohydrates, proteins and lipids converge on the citric acid cycle to
produce ATP

15. Explain how cellular respiration is regulated by energy levels of the cell.

16. Compare and contrast respiration and photosynthesis. Focus special on ATP,
water, CO2, and high-energy electron carriers NADH or NADPH in the two
processes.

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LO 13
Fermentation: What happens when oxygen is not available?

In the absence of oxygen, pyruvate produced from glycolysis can be

reduced to lactic acid or ethanol (alcohol).
LO 13
Lactic acid Fermentation

• Lactic acid fermentation occurs in

animal, human and some bacteria
cells.
• In the absence of oxygen, pyruvate
produced is reduced to lactic acid.
• This regenerates NAD+.
• The NAD+ can then be reduced in
glycolysis, and ATP is still
synthesized in small amounts for
use by the cell.

Glucose + 2ADP + 2Pi 2NADH → 2 lactic acid + 2ATP + 2NAD+

LO 13
Ethanol Fermentation

• Ethanol fermentation
occurs in plants and fungi.
• Pyruvate releases carbon
dioxide, and the
acetaldehyde produced is
reduced to form ethanol.
• This regenerates NAD+ so
that ATP can be generated
during glycolysis.

Glucose + 2ADP + 2Pi 2NADH → 2 ethanol + 2ATP + 2NAD+

 Cellular Respiration in the Absence of Oxygen

• Cellular respiration can occur in the absence of oxygen, but it

needs a final electron acceptor such as nitrate or sulfate.
• Respiration in the absence of oxygen, known as anaerobic
respiration, occurs today in some bacteria.
• The electron transport chain in these bacteria is located in
the plasma membrane, not in an internal membrane.
LO 14
Other Sugars Contribute to Glycolysis
LO 14 Fatty acids and amino acids are also useful sources of
energy
LO 15 Regulation of Cellular Respiration

• Pathways that generate ATP can be slowed or down-regulated

depending on the levels of free energy available in the cell.
• Indicators of energy available to the cell are concentrations of ATP,
NAD+, and NADH.
LO 16 Photosynthesis vs Respiration

Citric acid cycle

vs
Calvin cycle

Use the table in the tutorial questions

for a detailed comparison

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