Pentose Phosphate Pathway (PPP) also called Hexose Monophosphate Shunt (HMS)

Aims:
1. produce pentoses such as Ribose-5-P for nucleotide synthesis
2. Produce NADPH [Reducing Agent], important in lipid biosynthesis and detoxification process
- PPP is an alternative pathway of Glycolysis, Occurs in the Cytosol
- PPP is active In tissues that require NADPH such as RBCs, adipose tissue, Liver, testis, Ovary,
mammary glands
Oxidative phase (Irreversible)

PPP

Nonoxidative interconversion phase (Reversible)

Ketose
Irreversible Oxidative phase
▪ This phase consists of 2 irreversible reactions:

1. Glucose 6-phosphate is oxidized to 6-phosphogluconate (NADP+ is reduced to NADPH) Catalyzed by

Glucose 6-phosphate dehydrogenase (G6PD)
- This reaction is the committed/ rate limiting/ regulated step Inhibited by NADPH (High NADPH/NADP+)
- Insulin upregulates the expression (Transcription) of G6PD gene→ increase the rate of PPP in the
absorptive well-fed state (after meal).
2. The oxidative decarboxylation of 6-phosphogluconate to ribulose 5-phosphate (Ketopentose)
Catalyzed by 6-phosphogluconate dehydrogenase (6PGD), this enzyme also induced by insulin
- Produces the Ribulose-5-P, CO2 , and a second molecule of NADPH.

• In this phase one CO2 molecule and one pentose sugar-phosphate plus two NADPH are produced per
each glucose 6-P molecule
Reversible Non-oxidative phase
• Isomerization and Epimerization of Ribulose-5-P (Ru5P)
- Ru5P is converted to Ribose-5-P (R5P) by Isomerase or to Xylulose 5-phosphate (Xu5P) by epimerase
 Then, rearrangements of carbon atoms.
- three G6P molecules converted to 1 R5P and 2 Xu5P
- The conversion of 1 R5P and 2 Xu5P (3 C5) to two molecules of fructose-6P (2 C6) and one molecule
of Glyceraldehyde-3-P (1 C3) is catalyzed by two enzymes, Transaldolase and Transketolase.
• Transketolase catalyzes the transfer of two carbon units (C2 unit) and Transaldolase catalyzes the
transfer of three carbon units (C3 unit)
1. Transketolase, which has a Thiamine pyrophosphate cofactor (TPP), catalyzes the transfer of a C2
unit from Xu5P to R5P, yielding Glyceraldehyde-3-P and sedoheptulose-7-phosphate
2. Transaldolase catalyzes the transfer of a C3 unit from S7P to GAP yielding erythrose-4-phosphate
(E4P) and F6P
3. In a second Transketolase reaction, a C2 unit is transferred from a second molecule of Xu5P to E4P
to form GAP and another molecule of F6P.

Ketose + Aldose Aldose + Ketose

This phase of the PPP thus transforms two molecules of Xu5P and one of R5P to two molecules of F6P
and one molecule of GAP.
 (2 Glucose-6-P)

PPP ‫ في ال‬ATP ‫ال يتم انتاج او استهالك‬

• Transaldolase: Transfer 3C from Ketose to Aldose

• Transketolase: Transfer 2C from Ketose to Aldose
Note:
• If the cell requires NADPH and not the Pentose sugars “Like RBCs”, then it goes through the
oxidative and non-oxidative phases of the PPP
• If the cell require Pentoses and NOT NADPH; we can synthesize Pentoses from 2 Fructose-6-P and
1 Glyceraldehyde-3-P through the non-Oxidative phase (Reversible)
• If the cell requires both NADPH and Pentoses, we go through the Oxidative phase only
NADP + /NADPH ratio is the main Regulator for PPP
High NADP + /NADPH ratio Activate PPP
Low NADP + /NADPH ratio Inhibit PPP

Some Enzymes use NADH while other

Enzymes use NADPH
Normally
NAD+/NADH ratio = 1000/1; mostly NAD+ → Should
be ready for food oxidation.

NADP+/NADPH ratio = 1/10; mostly NADPH → Should

be ready for biosynthesis and detoxification reactions
 Summary

Q: All of the following sugar arrangement are part of pentose Q: in addition to pentoses , the pentose phosphate
phosphate pathway except:- pathway involves sugars of these sizes except :-
A. C5 + C5  C7 + C3 A. 3 carbons
B. C5 + C5  C6 + C4 B. 4 carbons
C. C7 + C3  C6 + C4 C. 6 carbons
D. C5 + C4  C6 + C3 D. 7 carbons
E. all of these arrangements occurs in pentose phosphate pathway E. all of these sizes are used in this pathway
What are the uses of NADPH??
1. Reductive Biosynthesis of Fatty acids and Steroids.
2. Antioxidant role
- Oxidizing agents such as Reactive Oxygen Species (ROS)
a. H2O2 Hydrogen Peroxide
b. O2∙⁻ Superoxide (Free radical)
c. OH∙⁻ Hydroxyl radical (Free radical) ‫اخطر واحد على الخلية‬
• These oxidizing agents formed as by-products of aerobic metabolism, Infections, drugs, toxins,
alcohol, food…
• ROS → oxidize and damage DNA, Lipids, proteins → cancer, Inflammation and cell death
How can we detoxify ROS??
You Body have several enzymes to do this job

(Selenium-containing Enzyme)

Regeneration of GSH (Reduced/active form of Glutathione)

NADPH is required for the regeneration of reduced glutathione (GSH) from oxidized glutathione (GSSG)
by Glutathione Reductase

• Also Dietary Ascorbate (Vitamin C), Vitamin E and carotenoids are Antioxidant
3. NADPH is required for the action of Monooxygenases
- such as Cytochromes P450 Monooxygenases [Cyt P450]
- Cyt P450 is a family of Enzymes (100 Enzymes) found in:
a. Mitochondria of Testis, Ovaries, liver, adrenal gland important for the synthesis and modification of
steroids such as sex hormones, Bile, Vitamin D
b. Endoplasmic Reticulum mainly of liver [microsomal System] important for Detoxification of
Xenobiotics (alcohol, Drugs, Toxins.. ).
• This family use NADPH as source of electrons
Electrons transferred from NADPH to FAD then to FMN in the
Cytochrome reductase then to Heme of Cyt P450 which use O2;
one O is added to the Substrate (become OH) and the other is
given the 2 electrons forming H2O

Substrate
4. NADPH is important for WBCs such as Macrophages to kill microbes during infections
The bacteria/microbe bind to antibodies usually IgG (Label), IgG bind to
an antibody receptor on the membrane of phagocytes.
 After binding, the bacterium with the IgG and its receptor are ingested,
forming a phagosome.
 The phagosome merges with a lysosome to form a phagolysosome in
which the destruction of the microbe occurs using:
- ROS: Reactive Oxygen Species Superoxide dismutase
- RNOS: Reactive Nitrogen Species

Bind to O2∙⁻ forming

Reactive nitrogen-
oxygen species

The process of production of ROS and RNOS during infection called Respiratory burst
5. NADPH is required for the Synthesis of Nitric Oxide (NO)
• Synthesis of NO require Arginine + O2 + NADPH
Functions:
- Vasodilator (relax smooth muscle)
- Prevent platelet aggregation
- Neurotransmitter
- Formation of RNS (Tumoricidal, Bacteriocidal)
• The Enzyme that synthesize NO called NO-Synthase; 3 Types
a. eNOS (Endothelium) synthesize NO for Vasodilation and inhibition of platelet aggregation
b. nNOS (Neuronal) synthesize NO as Neurotransmitter
c. iNOS (Inducible; found in Macrophage, Monocytes, Neutrophil)
 eNOS and nNOS are Ca+2-Calmodulin Dependent and Constitutive ‫تركيزهم يبقى ثابت وقليل في الخاليا‬
 iNOS is Ca+2 Independent and Inducible by Infections and Tumors

The Relaxing effect of NO on smooth muscle

 NO activate Enzyme called Guanylyl Cyclase which convert GTP to cGMP
→ cGMP activate Protein Kinase G
→ Protein kinase G add P to Ca+2 channels preventing Ca+2 entry to the cell
→ This will ↓ Ca+2 and relaxing the muscle
Glucose-6-P Dehydrogenase (G6PD) Deficiency
- Most common Intracellular Enzyme Deficiency
- 200-400 Million Individuals are affected, Mainly in the Middle East, South East Asia, Mediterranean,
and Tropical Africa
- Recessive X-linked (more common in Males XY)
- There are about 400 types of mutations affect in the G6PD gene that cause G6PD deficiency most of
them are Point mutation (Missense)

• Patients of G6PD deficiency suffer from attacks Hemolytic anemia; (RBCs mostly affected); Why?
- PPP is the Only source of NADPH in RBCs, Other cells have other source (Malate Dehydrogenase
System “Malic Enzyme”, also other cells increase the synthesis of G6PD)
 So, if G6PD is deficient, RBCs can NOT synthesize adequate amount of NADPH
 NADPH is required for regeneration of GSH
 Low NADPH in RBCs means Low GSH, Consequently RBCs will be weak against oxidizing agents

So, if oxidative stress occurs → oxidation of lipids and Hemoglobin, S-S bonds of the plasma proteins
→ loss of flexibility → Hemolysis → Hemolytic anemia → accumulation of bilirubin → Jaundice
In Words
1. Glutathione Peroxidase consume GSH to
GSSG for detoxification of H2O2
2. Glutathione Reductase regenerate GSH
using NADPH
• G6PD (PPP) is the only source of NADPH
• If G6PD is deficient then GSH can NOT be
regenerated then the RBC can NOT
detoxify oxidizing agents and eventually
will have shorter life span→ Lyse
Oxidative Stress can result from:
Note: G6PD deficiency provide resistance against
1. Drugs
Falciparum Malaria (the parasite that cause Malaria)
- Antibiotics(sulfamethoxazol)
- Antimalarial (Primaquine)
- Antipyretics (Acetanalide)
2. Fava beans (Favism)
3. Infections which is the most common factor
4. Neonatal Jaundice
.‫بعض المصابين باكلو فول بدون ما يصير عندهم تكسر في خاليا الدم الحمراء النو نقص االنزيم ما بكون عندهم حاد‬
G6PD deficiency ‫لكن اي شخص باكل فول وبصير عندو تكسر في خاليا الدم الحمراء هاد اكيد مصاب ب‬
G6PD Variants
• Wild Variant (B): activity 100%
• Mediterranean Variant (B ̄) ‫معظم المصابين في منطقة حوض البحر االبيض المتوسط‬
- Point mutation nucleotide 563 C→T this leads to missense Ser→Phe
- (B ̄) is class II G6PD has ˂ 10% of wild type activity

• African Variants
a. (A): one point mutation nucleotide 376 A → G, this leads to missense Asn to Asp, the A variant is
Class IV G6PD with 80% activity
b. (A⁻) two point mutations:
- nucleotide 376 A → G, this leads to missense Asn to Asp.
- nucleotide 202 G → A, this leads to missense Val to Met
- The A⁻ variant is Class III G6PD with 10-20% activity
Stability of different variants of G6PD over time
B wild Type: activity decline slowly over time

B ̄ Mediterranean: activity is low even in the young RBCs

A ̄ Variant: activity is almost normal in young cells but after 2-3 weeks, the
activity decline rapidly

After Hemolytic attack the body generate new (young) RBCs

 If you test Mediterranean Patient after hemolytic attack the deficiency can be detected from the day 1
 If you test (A ̄) patient after hemolytic attack you will find that G6PD activity is almost normal so it can be
misdiagnosed
• So you should repeat the test 2-3 weeks after the attack to role out G6PG Deficiency
Sources of ROS
ROS either produced as by-products in aerobic metabolism or produced to serve some functions:
• Oxidases E.g. Peroxidase O2 + H2 → H2O2
• Oxygenases “Monooxygenase and Dioxygenases”
- Monooxygenases (Hydroxylases)
- Dioxygenases such as COX enzymes that generate prostaglandins, thromoxanes and leukotrienes
• Electron transport chain “specifically Q”
• Respiratory Burst “Infections”
• Ionizing radiation such X-ray, nuclear radiation

Generation of O2ֹ‾
by respiratory
chain mainly
(CoQ)

Binuclear center
prevents release of
free O2 radicals