PENTOSE PHOSPHATE PATHWAY,

(phosphogluconate pathway or hexose

monophosphate pathway)
and
GLUCURONIC ACID PATHWAY
Professor Nazmi Özer, PhD, MedSp
Department of Biochemistry
Faculty of Pharmacy
Girne American University
Karmia / Girne / TRNC
Spring 2024
The pentose phosphate pathway (PPP) is primarily
a catabolic pathway that utilizes the 6 carbons of
glucose to generate 5 carbon sugars and reducing
equivalents:

The primary functions of this pathway are:

1. To generate reducing equivalents, in the form of NADPH,

for reductive biosynthesis reactions within cells.

2. To provide the cell with ribose-5-phosphate (R5P) for the

synthesis of the nucleotides and nucleic acids.
Ø The pentose phosphate pathway is active
in rapidly dividing cells (bone marrow, skin
and intestinal mucosa) and tumors use
ribose 5-phosphate to make RNA, DNA,
ATP, NADH, FADH2 and coenzyme A.

Ø NADPH is needed for reductive

biosynthesis or fight with the damaging
effects of oxygen radicals (ROS).

Ø Tissues that carry out extensive fatty acid

synthesis (liver, adipose, lactating
mammary gland) or very active synthesis
of cholesterol and steroid hormones (liver,
adrenal glands, gonads) require NADPH.
Ø Erythrocytes and the cells of the lens and cornea are
directly exposed to oxygen and damaging free radicals
generated by oxygen.

Ø By maintaining a high ratio of NADPH / NADP and GSH /

GSSG, cells can prevent or reverse oxidative damage to
proteins, lipids etc.

Ø As a consequence, cells of the liver, adipose tissue,

adrenal cortex, testis and lactating mammary gland have
high levels of the PPP enzymes.
Ø In fact 30% of the glucose oxidized in the liver occurs via the PPP.

Ø Additionally, erythrocytes utilize the reactions of the PPP to generate

large amounts of NADPH used in the reduction of glutathione.

Ø The conversion of ribonucleotides to deoxyribonucleotides (through

the action of ribonucleotide reductase) requires NADPH as the
electron source therefore, any rapidly proliferating cell needs large
quantities of NADPH.
 neutrophil

Ø The highest levels of PPP enzymes (in particular glucose

6-phosphate dehydrogenase) are found in neutrophils
and macrophages.

Ø These leukocytes are the phagocytic cells of the immune

system and they utilize NADPH to generate superoxide
radicals from molecular oxygen in a reaction catalyzed by
NADPH oxidase.
 Macrophage
Superoxide anion, in turn, serves to
generate other reactive oxygen
species (ROS: H2O2 and HOCl) to kill
the phagocytized microorganisms.

Following exposure to bacteria and

other foreign substances there is a
dramatic increase in O2 consumption
by phagocytes. This phenomenon is
referred to as the respiratory burst
(oxidative).
 Favism
Malaria parasite (Plasmodium falciparum) can Africa, parts of the Middle
not grow on G6PD-deficient erythrocytes.
Vicia fava
Toxic ingredient in vicia fava
is divicine in FAVİSM
Jaundince and kidney failure
Same symptoms can occur
by the ingestion of the
antimalarial drug,
primaquine or of sulfa
antibiotics and exposure to
certain herbicides.
Frequency: 25% tropical
East and Southeast Asia.
Ø The reactions of the PPP operate exclusively in the cytoplasm.
From this perspective it is understandable that fatty acid
synthesis (as opposed to oxidation) takes place in the
cytoplasm.
Ø The pentose phosphate pathway has both an oxidative and a
non-oxidative arm.
Ø Oxidative phase produces pentose phosphates and NADPH.
Ø Nonoxidative phase recycles pentose phosphates to glucose 6-
phosphate.
 Pentose
Phosphate Pathway
(PPP)
or
Phosphogluconate
Pathway
or
Hexose
Monophosphate
Shunt
(HMS)
 OXIDATIVE PHASE - 1

Step 1 Step 2

Glu-6-P + NADP+ + H2O → 6PGluconate + NADPH + H+

 OXIDATIVE PHASE - 2

Step 3 Step 4

6-P-Gluconate + NADP+ → R-5-P + CO2 + NADPH + H+

Sum: Glu-6-P + 2NADP+ + H2O → R-5-P + CO2 + 2NADPH + H+
 NONOXIDATIVE PHASE
Ø In tissues that require primarily NADPH, the pentose
phosphates produced in the oxidative phase are recycled
into glucose 6-phosphate.
Ø The primary enzymes involved in the non-oxidative steps
of the PPP are transaldolase and transketolase.
Ø Ribulose 5-phosphate is first epimerized to xylulose 5-
phosphate.
Transketolase catalyzes the transfer of a two-carbon
fragment from a ketose donor to an aldose acceptor
1. The first reaction catalyzed by
transketolase:
Conversion of two pentose phosphates to a triose
phosphate and a seven-carbon sugar phosphate,
sedoheptulose 7-phosphate.
2. The reaction catalyzed by transaldolase:
3. The second reaction catalyzed by
transketolase
SUMMARY

C5 + C5 « C7 + C3 Transketolase
C7 + C3 « C6 + C4 Transaldolase
C4 + C5 « C6 + C3 Transketolase
3 C5 « 2 C6 + C3
Wernicke-Korsakoff syndrome
Ø Wernicke - Korsakoff syndrome is a disorder caused by
a severe deficiency of thiamine, a component of TPP.
Ø More common among people with alcoholism.
Ø A genetic defect in transketolase that lowers its affinity
for TPP.
Ø Memory loss
Ø Mental confusion
Ø Partial paralysis
Glucose 6-phosphate is shared between
glycolysis and the pentose phosphate pathway.
Ø Entry of glucose 6-phosphate either into glycolysis or into the
pentose phosphate pathway is largely determined by the
relative concentrations of NADP and NADPH.

When NADPH is forming

faster than it is being used
for biosynthesis and
glutathione reduction,
[NADPH] rises and inhibits the
first enzyme in the pentose
phosphate pathway (product
inhibition). As a result, more
glucose 6-phosphate is
available for glycolysis.
 Antioxidant Enzyme Systems - 1
Reactive Oxygen Species ( • unpaired electron)

Oxygen Superoxide ion Peroxide Hydroxyl radical Hydroxyl ion

 Antioxidant Enzyme Systems - 2
1. Superoxide dismutases (SODs) are metalloenzymes

They contain copper, zinc (Cu/Zn-SOD, cytosol) and

manganese (Mn-SOD, mitochondria).

SOD converts two molecules of superoxide ion to 1

molecule of O2 and 1 molecule of H2O2.

SOD
O2•- + O2•- H2O2 + O2

2. Catalase (CAT) is a iron containing metalloenzyme

CAT
2 H2O2 2 H2O + O2
 Antioxidant Enzyme Systems - 3
Ø Although NADPH is a major reducing power, it does not give
electrons to every reaction or molecule. Mediator molecules
are required.
Ø Most important mediator molecule is glutathione (GSH) (g-
Glu-Cys-Gly). In GSH thiol group (-SH) of cysteine can accept
or donate electrons.
Ø Some reducing and defence systems use GSH as a reducing
power.

GSH GSH
-2e- +2e-
GSSG
g-glutamylcysteinylglycine
 Antioxidant Enzyme Systems - 4
3. Glutathione peroxidase (GPx):
Glutathione peroxidase contains selenium in the
active site.
GPx
2GSH + H2O2 Þ GSSG + 2H2O
This reaction is important for organisms. Therefore
glutathione must be kept in the reduced (GSH) state.

Reduction of GSH is achived enzymatically. For

reduction, NADPH is the reducing power.
 Reducing Power Sources - 1
Main enzymes that produce NADPH are
located in cytosol.

1. Glucose-6-phosphate dehydrogenase

Glu-6-P + NADP++H+Þ6-Phosphogluconolactone + NADPH

2. 6-Phosphogluconate dehydrogenase
6-PGA + NADP+ +H+ Þ Ribulose-5-phosphate + NADPH

3. Isocitrate dehydrogenase (cytosolic)

Isocitrate + NADP+ +H+ Þ alpha-KGA + CO2 + NADPH

4. Malic enzyme
Malate + NADP+ +H+ Þ Pyruvate + CO2 + NADPH
 Reducing Power Sources - 2
In organisms, main reducing power is NADPH

X GS-X
Glutathione S-Transferase

NADP GSH H2O2

+ 2x
1. G6PD
2. 6PGD Glutathione Glutathione
3. (c) IDH
4. MALIC ENZYME Reductase Peroxidase

NADPH GSSG 2
H2O
Reducing power sources and role of glutathione in cells
 Antioxidant Systems in Erythrocytes
Oxidative stress
Drugs, Infection,
Glucose Fava beans

Glucose

G-6-P
NADP+ 2 GSH H2O2
G6PD GSH GSH
Glycolysis reductase peroxidase

6-PGA NADPH GSSG 2H2O

Lactate
PP

Lactate
 Antioxidant Systems in Leukocytes
Ø Leukocytes use series of enzymes and mechanisms in order to
kill foreign organisms. NADPH is required for these enzymes.

Ø Chronic granulomatous disease: Deficiency of enzymes

(NADPH oxidase, myeloperoxidase). Leukocytes can take up
bacteria by phagocytosis but can not kill them!!!

PHAGOSOME
Fe2+ Fe3+

T E
Fenton
NADPH Reaction OH
•

O2 oxidase O2•- SOD H2O2

B A K
HOCl

RIA
NADPH NADP
Cl- Myeloperoxidase
 Glucuronic Acid (Uronic Acid) Pathway

A uronic acid is a sugar acid

with both a carbonyl and a
carboxylic acid function. It is
best thought of as a sugar in
which the terminal carbon's
hydroxyl function has been
oxidized to a carboxylic acid.

It is a quantitatively minor
route of glucose metabolism.
It provides biosynthetic
precursors and interconverts
some less common sugars to
ones that can be utilized.
Glucuronic Acid Synthesis
PPi
Glu-6-P UDP-Glucose
UDP-Glucose 2NAD+ H2 O
Phosphoglucomutase pyrophorylase
UDP-Glucose
Glu-1-P UTP
dehydrogenase
2NADH + 2H+
UDP-Glucuronate

Glucuronides
Proteoglycans Glucuronate
Glycosaminoglycans
L-Ascorbate*
*Do not exist in primates thus, it is a vitamin for primates.
Functions of Glucuronic Acid Pathway
Ø The uronic acid pathway is utilized to synthesize UDP-
glucuronate, glucuronate and L-ascorbate.

Ø The pathway involves the oxidation of glucose-6-phosphate to

UDP-glucuronate.

Ø The oxidation is uncoupled from energy production.

Ø UDP-glucuronate is used in the synthesis of

glycosaminoglycans and proteoglycans as well as forming
complexes with bilirubin, steroids and certain drugs.

Ø The glucuronate complexes form to solubilize compounds for

excretion.

Ø The synthesis of ascorbate (vit C) does not occur in primates.

Glucuronidation
UDP-glucuronic acid reacts with alcohol (OH), carboxylic acid (COOH),
amine, amide (NH2) and thiols (SH).
Ether formation with alcohols Ester formation with acids
Glucuronyltransferase Glucuronyltransferase
Phenol + UDP-GA Þ a) Benzoic acid + UDP-GA Þ
Phenyl-glucuronide + UDP Benzoyl-glucuronide + UDP
Glucuronyltransferase
b) Bilirubine + (UDP-GA)2 Þ
Bilirubine diglucuronide + UDP

UDP-Glucuronate
UDP-glucuronate
transferase
(microsomal)

Phenylglucuronide
Bile, Urine Glucuronates Bilirubine
 Physiological Significance of
Glucuronide Formation
Ø UDP-glucuronyltransferase catalyzes the
conjugation of endogenous and exogenous
substances to form glucuronides.
Ø Conjugation with glucuronic acid produces a water
soluble, strongly acidic compound.
Ø Important in drug detoxification, steroid excretion
and bilirubin metabolism.

Ø Diseases related to glucuronic acid metabolism:

- Physiological jaundice of the newborn
- Gilbert Disease
- Crigler-Najjar Syndrome
 Physiological jaundice of newborn
Jaundice in newborn babies is common and usually harmless. It causes
yellowing of the skin, the whites of the eyes and dark yellow urine (a
newborn baby's urine should be colourless). The medical term for jaundice
in babies is neonatal jaundice.

The symptoms of newborn jaundice usually develop 2 days after the birth
and tend to get better without treatment by the time the baby is about 2
weeks old. breastfeeding can improve the mother's milk supply and, in
turn, improve caloric intake and hydration of the infant, thus reducing the
elevated bilirubin

The baby's liver will take only a few days to

process bilirubin properly. Phototherapy is
the most common treatment. Phototherapy
transforms the bilirubin in the baby's skin
into a less harmful chemical. Your baby
may be undressed and placed in a warm
incubator under blue lights.
Clinical Significance of Glucuronate - 1
Ø In adults, a significant number of erythrocytes die each
day. This turnover releases significant amounts of the
iron-free portion of heme, porphyrin, which is
subsequently degraded.

Ø The primary sites of porphyrin degradation are found in

the reticuloendothelial cells of the liver, spleen and bone
marrow.

Ø The breakdown of porphyrin yields bilirubin, a product

that is non-polar and therefore, insoluble.
Ø In the liver, to which is transported in the plasma bound
to albumin, bilirubin is solubilized by conjugation to
glucuronate.
Clinical Significance of Glucuronate - 2
Ø The soluble conjugated bilirubin diglucuronide is then
secreted into the bile.

Ø An inability to conjugate bilirubin, for instance in hepatic

disease or when the level of bilirubin production
exceeds the capacity of the liver, is a contributory cause
of jaundice.

Ø The conjugation of glucuronate to certain non-polar

drugs is important for their solubilization in the liver.

Ø Glucuronate-conjugated drugs are more easily cleared

from the blood by the kidneys for excretion in the urine.
Crigler-Najjar Syndrome
Ø Crigler-Najjar syndrome is a rare autosomal recessive
inherited disorder characterized by the absence or decreased
activity of UDP-glucuronosyltransferase-1A1 (UGT1A1), an
enzyme required for the conjugation of bilirubin in the liver. It
is one of the major causes of congenital non-hemolytic
jaundice.

Ø In Crigler-Najjar syndrome type I and II mutations lead to the

exchange of amino acids, changes of the reading frame or to
stop codons.

Ø Severe hyperbilirubinemia has the potential to cause

irreversible brain damage. Hence, prompt diagnosis and
treatment is of utmost importance.
 Gilbert Syndrome
Ø Gilbert's syndrome is a mild disorder that typically doesn't need
medical treatment. People with the disorder lead normal, healthy
lives. There is no evidence to suggest that the condition is harmful
or leads to more serious diseases

Ø In Gilbert syndrome reduced glucuronidation of bilirubin in liver is

observed which leads to unconjugated hyperbilirubinemia (up to
95% of bilirubin unconjugated) and recurrent episodes of jaundice.

Ø Gilbert syndrome is due to a genetic variant in the UGT1A1 gene

(two bases are inserted into the promoter of the gene) which
results in decreased activity of the bilirubin uridine diphosphate
glucuronosyltransferase enzyme. It is typically inherited in an
autosomal recessive pattern and occasionally in an autosomal
dominant pattern depending on the type of variant.