Nucleotides: Chemistry and

Metabolism
Dr. Ankit Gupta
 Objectives
1. Nucleosides vs Nucleotides and their functions
2. Purine/Pyrimidine Biosynthesis
3. De novo biosynthesis pathway
4. Salvage Pathway
5. Regulation of Purine/Pyrimidine nucleotide synthesis
6. Inhibitors of Purine/Pyrimidine nucleotide synthesis
Nucleoside vs Nucleotide
 Pentose sugars

D-Ribose and 2’-Deoxyribose

*Lacks a 2’-OH group

Nitrogenous Bases
 Naming Conventions
 Nucleosides:
 Purine nucleosides end in “-sine”
(Adenosine, Guanosine)

 Pyrimidine nucleosides end in “-dine”

(Thymidine, Cytidine, Uridine)

 Nucleotides:
 S t a r t with the nucleoside name from above
and add “mono-”, “di-”, or “tri-phosphate”
(Adenosine Monophosphate, Cytidine
Triphosphate, Deoxythymidine Diphosphate)
 Purines and Pyrimidines are heterocyclic
compounds
 Smaller pyrimidines molecule has longer name and larger purine has shorter
name.
 Six-atom rings are numbered in opposite direction.
 Purines and pyrimidines with –NH2 group are weak bases.
 The planer character of purines and pyrimidines facilitate their close association, or
stacking that stabilizes dsDNA.
Uncommon naturally occurring purines
and pyrimidines
 Nucleosides are N-glycosides
 Purines or pyrimidines bases with Sugar through an N-glycosidic linkage.
 Purines are linked to C-1 carbon of sugar through N-9 atom.
 Pyrimidines are linked to C-1 carbon of sugar through N-1 atom.
 Due to no freedom of rotation about β-N-glycosidic bond of nucleosides or
nucleotides, therefore both exist as noninterconvertible Syn or Anti conformers.
 Both Syn and Anti conformers exists in nature, the anti conformers predominates.

Ribonucleosides: Syn conformers

 Nucleosides

Ribonucleosides: Anti conformers

 Nucleotides are phosphorylated
nucleosides
 Nucleosides with one or more phosphorylated group esterified to –OH group (3’ or
5’) of sugar.
 Nucleotides
 RNA is a polymer of ribonucleotides, whereas DNA is of deoxyribonucleotides.
 Both DNA and RNA contain Adenine, Guanine and Cytosine.
 RNA consists Uracil whereas, DNA consists Thymine.
 Nucleotides absorb UV light through conjugated double bonds of purine and
pyrimidine derivatives.
 At pH=7.0, all common nucleotides absorb light at wavelength close to 260nm,
therefore conc. of nucleic acid is expressed in terms of absorbance at 260nm.
 Nucleosides triphosphates are important energy carriers e.g. ATP, GTP.
 Non-hydrolyzable nucleoside triphosphate analogs serve as research tools.

Hydrolyzable

Unhydrolyzable

Unhydrolyzable
 Nucleotides serves diverse physiologic
functions
 Apart from precursors of nucleic acids, nucleotides e.g. ATP serves a role of
principle biologic transducer of free energy and secondary messenger cAMP.
 GTP serves as a role of an energy source in protein synthesis and cGMP as a
secondary messenger in response to NO during smooth muscle relaxation.
 UDP-sugar derivatives participate in sugar epimerization and in glycogen
biosynthesis, glycoproteins and proteoglycans.
 UDP-glucoronic acid forms urinary conjugate of bilirubin and many drugs e.g.
aspirin.
 CTP participates in biosynthesis of phosphoglycerides, sphingomyelin, and other
substituted sphingosines.
Synthetic nucleotide analogs are
used in chemotherapy
Nucleotide metabolism
 Nucleotide
Biosynthesis

Who don’t synthesize??

 Nucleotide biosynthesis
 All forms of life synthesize purines and pyrimidines except parasitic
protozoa.

 These have been synthesized in vivo at rates consistent with

physiologic need.

 Study of nucleotide biosynthesis was first employed at birds and

isotopic precursors of uric acid fed to pigeons established the source of
each atom of purine.
 Nucleotide Biosynthesis

De novo Salvage
synthesis pathway
Where nucleotides are Used to recover bases and
synthesize new from nucleosides formed during
simple precursor molecules degradation of RNA and DNA
De novo synthesis

Salvage pathway
• De novo synthesis: Major pathway
 Synthesis of purine nucleotides from various small
molecules derived as intermediates of many
metabolic pathways in the body.
• Salvage pathway: Minor pathway
 Synthesis of purine nucleotides from intermediates
in the degradative pathway for nucleotides.
 Purine nucleotide biosynthesis
 The three processes contribute in purine nucleotide biosynthesis in
order of decreasing importance:

1. Synthesis from amphibolic (a biochemical pathway that involves both

catabolism and anabolism) intermediates known as de novo synthesis.

2. Phosphoribosylation of purines
3. Phosphorylation of purine nucleosides
The assembly of purine ring is from
various sources
 Purine nucleotide: de novo synthesis
 This pathway includes two stages:
- Stage I: Synthesis of parent nucleotide “IMP”
- Stage II: Synthesis of AMP and GMP from IMP

 The end product of stage I (11 steps) is Inosine monophosphate (IMP).

 IMP is basically a nucleotide of base, hypoxanthine and branch point from which
two monophosphates, AMP and GMP are formed in stage II.
- Conversion of IMP to AMP
- Conversion of IMP to GMP
 Stage I: Synthesis of IMP
 Step 1: Activation of ribose-5-phosphate
 Transfer of two phosphoryl groups from ATP to C-1 of ribose-5-phosphate
(product of pentose phosphate pathway) leads to formation of
phosphoribosyl pyrophosphate (PRPP), catalyzed by PRPP synthetase.

 Step 2: 5-Phosphoribosylamine synthesis

 It is a committed and rate limiting step in purine biosynthesis pathway and
involves conversion of PRPP to 5-phosphoribosylamine using glutamine.
This step is catalyzed by PRPP amidotransferase.

 Steps 3-11: Purine ring is constructed in a step wise manner using

contributions from various precursors (THF, Gln, Asp, CO2, Glycine).
Purine biosynthesis from ribose-5-phosphate and ATP
 Stage II: Conversion of IMP to AMP and GMP
 Conversion of IMP to AMP: Addition of aspartate to IMP followed by removal of
fumarate generates AMP.
 Conversion of IMP to GMP: Oxidation of IMP followed by addition of amino group
from glutamine generates GMP.
Aspartate Fumarate
 Formation of nucleotide di- and tri-phosphates
 Conversion of nucleoside monophosphates (NMPs) to di- and tri-phosphates occurs
by various kinases and utilize ATP as a substrate.

(a) The nucleoside monophosphate kinase convert NMPs to corresponding di-

phosphates.
Adenylate kinase
AMP ADP
ATP ADP
Guanylate kinase
GMP GDP
ATP ADP

(b) Conversion of nucleoside diphosphates to corresponding triphosphates is catalyzed

by a single enzyme k/a nucleoside diphosphate kinase. This enzyme have
broad specificity.

Nucleoside diphosphate kinase

Nucleoside diphosphate Nucleoside triphosphate
ATP ADP
Salvage pathway
 Salvage reactions convert purine &
ribonucleosides to mononucleotides
 Salvage reactions require far less energy than
de novo biosynthesis.

 First salvage rxn: Phosphoryl transfer from

PRPP catalyzed by APRT and HGPRT converts
adenine, hypoxanthine and guanine to their
mononucleotides (Pu-RP):
Pu + PR-PP  Pu-RP + PPi

 Second salvage rxn: Phosphoryl transfer from

ATP to a purine ribonuceloside (Pu-R) e.g.
conversion of adenosine to AMP by adenosine
kinase.
Pu-R + ATP  PuR-P + ADP
Free purine bases resulted from
nucleotide degradation are
recycled to resynthesize
nucleotides in Salvage Pathway.
 Purine Salvage
 Adenine phosphoribosyl transferase (APRT)
Adenine + PRPP AMP + PPi

 Hypoxanthine-Guanine phosphoribosyl
transferase (HGPRT)
Hypoxanthine + PRPP IMP + PPi
Guanine + PRPP GMP + PPi

 Free purines can be salvaged (recycled) to resynthesize

nucleotides which requires PRPP-dependent salvage enzymes,
APRT and HGPRT.
 Interestingly, PRPP is the key element of Salvage pathway and
the starting point of the de novo synthesis pathway.
 Significance of Salvage pathway over
de novo synthesis

 Simpler and more cost-effective way:

- reducing the requirement for the energetically expensive de novo synthesis.

 Preventing wastage of raw materials:

- Important for tissues e.g. brain and erythrocytes where de novo pathway is
poorly developed or absent due to deficiency or absence of PRPP
amidotransferase.

 Diverse in character and distribution:

- de novo synthesis is virtually identical in all cells.
Regulation of purine
nucleotide
synthesis
 Regulation of de novo synthesis of
purine nucleotides
A. First level of regulation: Synthesis of IMP from R-5-P
 The principal regulatory enzymes are: PRPP
synthetase and PRPP amidotransferase.
 PRPP synthetase is inhibited by AMP, ADP, GMP,
GDP.
 Rate of PRPP synthesis depends on availability of
Rib-5-phosphate.
 PRPP amidotransferase, a committed and rate-
limiting enzyme, is a dimeric protein to which GMP
(potent inhibitor) binds at different inhibitory sites.
 PRPP amidotransferase is apparently inactive in
dimeric form and increase levels of PRPP stimulates
its dissociation into two subunits so that it acquires
catalytic activity.
 The activity of PRPP amidotransferase is sterically
stimulated by PRPP.
B. Second level of regulation: Synthesis of AMP and GMP from IMP

 The IMP to AMP/GMP conversion is

feedback inhibited by AMP and GMP.
 AMP and GMP are competitive inhibitors
of IMP and balance their own production
as required nearly in equal volume.
 Conversion of IMP to AMPS requires
GTP.
 Conversion of XMP to GMP requires
ATP.
 This cross-regulation between the
pathways of IMP metabolism balance the
synthesis of both ATP and GTP during
their deficiency.
 Regulation of Salvage pathway
 HGPRT is competitively inhibited by
IMP and GMP.

 APRT is competitively inhibited by

AMP.
 Inhibitors of Purine Synthesis

 Para-aminobenzoic acid (PABA) analogues

- De novo synthesis requires THF, a derivative of folic
acid.
- Reduced THF levels inhibits de novo pathway.
- Sulphonamides (PABA analogue) have no
deleterious effect on humans because of lacking
capability to synthesize folic acid from PABA.

 Antifolates
- Methotrexate, aminopterin and trimethoprim
competitively bind to DHFR with 1000-fold more
avidity than DHF.
- Decreased THF availability blocks synthesis of dTMP
from dUMP by thymidylate synthase.
Pyrimidine Biosynthesis
 Objectives
1. Purine Biosynthesis
2. De novo synthesis of pyrimidine nucleotide
3. Salvage Pathway
4. Regulation of Pyrimidine nucleotide synthesis
 De novo synthesis of pyrimidines
nucleotides
 Here PRPP participates only in assembly of pyrimidine ring unlike purine
biosynthesis where PRPP serves as a scaffold for assembly of purine ring.

 Purine biosynthesis is energetically more costly than biosynthesis of

pyrimidines.
 Steps in de novo pathway
 Step 1: Carbamoyl phosphate synthetase II
(CPS II) catalyzes the formation carbamoyl
phosphate.
 Step 2-4: CAP condenses with aspartate and
processed to form orotic acid, the first
pyrimidine which is produced.
 Step 5: Orotic acid is converted to orotidine
monophosphate (OMP), a parent pyrimidine
nucleotide.
 Step 6: OMP is decarboxylated to uridine
monophosphate (UMP), catalyzed by OMP
decarboxylase.
 Step 7-12: UMP is eventually converted to
 First three enzyme activities
other nucleotides.
reside on multifunctional
 All enzymes of this pathway are polypeptides k/a CAD (single
cytosolic except dihydrooorotate polypeptide named for first
dehydrogenase (step 4), which is letters of enzymes).
mitochondrial.
It is a polypeptide made up of four different
domains which make for a multi enzyme unit:
Glutaminase (GLN),
carbamoyl phosphate synthetase (CPS II),
Dihydroorotase (DHO) and aspartate
transcarbamoylase (ATC).
 Salvage pathway of pyrimidines
nucleotide synthesis

 Orotate phosphoribosyl transferase (OPRT) is an enzyme of de novo

pathway but also salvages other pyrimidine bases.

Orotate OMP
OPRT
Uracil + PRPP UMP + PPi
Cytosine CMP
 Reduction of NDPs to dNDPs

 Reduction of 2’-OH of purine and

pyrimidine ribonucleotides are catalyzed by
ribonucleotide reductase.

 Reduction requires:
- reduced thioredoxin
- thioredoxin reductase
- NADPH.

 Reduced thioredoxin is produced by

NADPH-dependent reduction of oxidized
thioredoxin.

 NDPs to dNDPs conversion has complex

regulatory control over production of
dNTPs for DNA synthesis.
 Regulation of pyrimidine nucleotide
biosynthesis
 CPS II is:
- main regulatory enzyme which exists
in a polyprotein k/a CAD
- inhibited by UTP.
- activated by ATP and PRPP.

 OMP decarboxylase (step 6) is inhibited by

UMP and CMP.

 Purine and pyrimidine nucleotide biosynthesis

are cross-regulated.

 PRPP synthetase, a precursor for both

processes is feedback inhibited by both
purine and pyrimidine nucleotides.
Regulation of conversion of purine and
pyrimidine NDPs to NTPs
 Methotrexate blocks reduction of
Dihydrofolate
 The reaction catalyzed by thymidylate synthase, only requires tetrahydrofolate
(THF) derivative during pyrimidine nucleotide biosynthesis.
 During this reaction, the methylene group of N5, N10-methylene-tetrahydrofolate is
reduced to 5-position of pyrimidine ring, and THF is oxidized to dehydrofolate.
 Dihydrofolate must be reduced back to tetrahydrofolate for further pyrimidine
biosynthesis.
 This reduction is catalyzed by dihydrofolate reductase which is inhibited by
methotrexate.
 Dividing cells (generate TMP and dihydrofolate) are specially sensitive to inhibitor
of dHFR e.g. anticancer drug methotrexate.
Purine nucleotide
degradation
 Degradation of Purine nucleotides
 5´-nucleotidases: convert a nucleotide to respective nucleoside.
 Purine nucleoside phosphorylase: liberate free base from respective nucleoside.
 Adenosine deaminase: converts adenosine to Inosine.
 Xanthine oxidase: converts Hypoxanthine and Xanthine to Uric Acid.
 Xanthine is a common product in both adenine and guanine catabolic pathways
which is finally converted to uric acid.
 The end product of purine degradation in humans is Uric Acid.
 This process occurs mainly in liver.
 Humans catabolize purines to
uric acid
 Ribose sugar is recycled in form of (R-1-P to R-5-P)
and incorporated into PRPP.

 In mammals other than higher primates, uricase

converts uric acid to water-soluble product allantoin.

 Since, humans lack uricase , the end product of purine

catabolism in human is uric acid.
Disorders of Purine
metabolism
 Disorders of purine metabolism
1. Gout

2. Lesch-Nyhan syndrome

3. von Gierke Disease: Glucose-6-phosphatase deficiency leads purine

overproduction and hyperuricemia due to enhanced generation of PRPP precursor
Rib-5-phosphate.

4. Hypouricemia: associated with a deficiency of xanthine oxidase leading to

increased excretion of hypoxanthine and xanthine. Patients exhibit xanthinuria.

5. Adenosine deaminase deficiency: leads to accumulation of adenosine and

deoxy-adenosine which is toxic to lymphocytes and suppress immune function also
k/a Severe Combined Immunodeficiency Disease (SCID).

6. Purine nucleoside phosphorylase deficiency: Accumulation of inosine and

guanosine occurs in plasma followed by urinary excretion of these compounds.
Gout
 Gout
 Gout is characterized by hyperuricemia and recurrent acute arthritis.

 Over 90% of the hyperuricemia patients are males; suffering women are usually
postmenopausal.

 Not all patients with hyperuricemia develop gout (asymptomatic).

 Uric acid is barely soluble in plasma therefore even a moderate rise in its
concentration leads to precipitation of uric acid crystals.

 Subcutaneous deposition of sodium urate crystals (25% of cases) and its

decreased renal excretion (75% of cases) are most underlying cause of Gout.
 Types of Gout
Gout

Primary Secondary
- 90% due to inborn error of - Due to various disease causing
metabolism e.g. defect in increased synthesis and
enzymes: decreased excretion of UA:
- ↑ PRPP synthetase - Increased Dietary intake
- HGPRT deficiency - Chronic renal failure
- Glu-6-Phosphatase deficiency - Leukemia and multiple
myeloma
- 10% idiopathic - Starvation
 Primary Gout
 Increased uric acid generation due to synthesis and degradation of purine
nucleotides.

1. Increased activity of PRPP synthetase:

- X-linked trait
- Enhanced purine biosynthesis

2. Glu-6-Phosphatase deficiency:
- k/a von Gierke disease
- Intracellular accumulation of G-6-P leading to R-5-P formation through
HMP shunt pathway.

3. HGPRT deficiency:
- X-linked trait leads to Lesch-Nyhan syndrome.
- Increased purine synthesis by 3xfold
- Recycling of guanine and hypoxanthine is impaired; so metabolized to UA.
- PRPP accumulation which is channeled into de novo synthesis.
- Decreased levels of AMP and GMP leading to decreased feedback
inhibition of PRPP amidotransferase which speeds up purine synthesis
 Low solubility: the offending factor in Gout

 The low solubility and consequent precipitation

of UA leads to-
- Gouty arthritis
- UA nephropathy
 When serum urate concentration rises; its
sodium salt readily forms in body.
 Monosodium urate (MSU) is predominant form
of UA at neutral physiological pH.
 Focal depositions of these precipitates are
MSU: Needle shaped crystals
called tophi.

Low pH
 Clinical manifestations of Gout

 Gouty arthritis:
- Accumulation of tophi in joints triggers inflammatory response of gouty
arthritis.
- Big toe is mostly involved
- Severe pain triggered by alcohol
- UA crystals engulfed by leucocytes and release of lysosomal enzymes
during rupture those degrade articular tissue and induce inflammatory
response.

 Nephropathy:
- MSU or UA crystals may precipitate in kidneys as renal stones.
- Prolonged kidney damage results in its impaired functioning k/a UA
nephropathy.
Treatment of Gout
 Treatment of Gout
 Principal of Gout treatment is: decreasing UA production, increasing its excretion
and reducing inflammation.

 Decreasing urate production:

- Xanthine Oxidase inhibitors e.g. Allopurinol, Fabuxostat
- accompanied by accumulation of xanthine and hypoxanthine which are
more soluble than UA so readily excreted through urine.

 Increased renal excretion of urate with uricosuric drugs e.g. probenecid and
salicylates.

 Anti-inflammatory drugs useful in acute gouty arthritis e.g. Colchicine.

Allopurinol: Suicide inhibitor of Xanthine Oxidase
 It is a hypoxanthine analogue (structurally similar) with interchanged N-7 and
C-8 positions.

 Xanthine oxidase hydroxylates allopurinol to yield alloxanthine, which remains

tightly bound to the enzymes and thereby inactivating it.
 Alcohol consumption and Gout
 Lactic acid is an organic anion, and is a
counter ion for uric acid at the URAT1
antiporter on the apical surface of the
collecting duct; thus the more lactate
secreted into the urine, more uric
acid reabsorbed into the blood
Causes
1. PRPP Amido transferase
2. PRPP Synthetase
3. Deficiency of enzymes of salvage pathway
4. Glucose-6-Phosphatase deficiency
Gout vs Pseudogout
 Lesch-Nyhan Syndrome
 Caused due to defect in an enzyme involved in
purine salvage; HGPRT.
 Leads to increased levels of hypoxanthine and
guanine resulting into uric acid production through
their degradation.
 Resulted in loss of GMP-mediated negative
feedback inhibition leads to PRPP accumulation
thereby increased production of purines and their
subsequent degradation.
 It is X-linked disorder caused by mutations those
abolish HGPRT activity through deletions, frame-
shift mutations, base substitutions etc.
 Symptoms: similar to gout, neurological,
aggressiveness
 First neuropsychiatric abnormality that was
attributed to a single enzyme; HGPRT.
Pyrimidine nucleotide
degradation
Pyrimidine catabolites are water
soluble
 Unlike low soluble products of purine catabolism,
catabolic products of pyrimidines are highly water-
soluble e.g.
- CO2
- NH3
- β-alanine
- β-aminoisobutyrate

 β-alanine is converted to succinyl-CoA and

ultimately to glucose.
 β-aminoisobutyrate is further metabolized to
certain intermediates of FA metabolism.
Excretion of β-aminoisobutyrate increases during
leukemia and X-Ray exposure due to DNA
destruction.
 Many anti-cancer drugs inhibit pyrimidine
synthesis
 Several anticancer drugs inhibit both purine and pyrimidine biosynthesis thereby
inhibit DNA synthesis and effectively used in chemotherapy.

 Glutamine antagonists inhibits steps in purine and pyrimidine synthesis where

glutamine donates a nitrogen e.g. Azaserine

 5-Fluorouracil is an analogue of uracil which is converted to fluoro-UMP that

inhibits thymidylate synthase activity.

5-Fluorouracil
Disorders of Pyrimidine
metabolism
 Disorders of pyrimidine catabolism
 β-hydroxybutyric aciduria: disorder linked to
deficiency of dihydropyrimidine dehydrogenase.

 Combined uraciluria-thyminuria: disorder linked to

impaired synthesis of β-alanine and β-
aminoisobutyrate.

 Hyperuricemia – related to overproduction of PRPP,

leading to overproduction of pyrimidine nucleotides and
therefore, increased excretion of β-alanine.
 Orotic Aciduria
 An autosomal recessive disorder,
characterized by increased excretion of
orotic acid (OA) in urine.

 Symptoms: retarded growth, severe

anemia

 Orotic aciduria results from deficiency of

either or both of the last two enzymes:
OPRT and OMP decarboxylase.

 Intake of diet rich in uridine (or cytidine) te

i ne ha S)
is an effective treatment for orotic aciduria. r id osp MP
U h (U
n op se
o a
 Uridine provides enough UMP through m nth
sy
phosphorylation, and also inhibits CPS II
so as to decrease the rate of OA
synthesis.
 Types of Orotic Aciduria
 Enzyme activities of both OPRT and OMP decarboxylase are present on a
single protein as domains (bifunctional enzyme).

 Mutation in the phosphoribosyl transferase impairs its association with the

other (decarboxylase) domain, resulting in loss of activity of both enzymes
(UMPS), resulting into type 1 orotic aciduria.

 In type 2 orotic aciduria, mutation in the decarboxylase occurs, but it does

not affect its aggregation with phosphoribosyl transferase.

 In this case, only decarboxylase is inactive, whereas activity of

phosphoribosyl transferase is not affected.)
 Clinical manifestations of Orotic Aciduria
 OA is not inherently toxic, but excessive urinary level of this substance (more
than 1 g/day; normal, 1.4 mg/day) leads to its crystallization and consequent
obstruction to urine flow.

 Moreover, block in pyrimidine synthesis occurs in orotic aciduria, which leads to

shortage of pyrimidine nucleotides for incorporation into DNA and RNA.

 Insufficient genetic material decreases generation of new cells.

 The rapidly proliferating cells such as RBCs are most prominently affected,
which results in anaemia.

 The erythrocyte precursors in the bone marrow become both abundant and
unusually large.

 The diagnosis is confirmed by assay of the above enzymes in erythrocytes, or

fibroblasts.
 Drugs inducing Orotic Aciduria
 Allopurinol, an alternate substrate for OPRT competes with orotic acid.

 The resulting nucleotide product also inhibits orotidylate decarboxylase resulting

into orotic aciduria and orotidinuria.

Competes with

Allopurinol
 Drugs inducing Orotic Aciduria
 6-Azauridine, following conversion to 6-azauridylate, also competitively
inhibits orotidylate decarboxylase thereby enhancing excretion of orotic acid
and orotidine (OMP).
 Rey’s Syndrome
 Excessive production of orotic acid may also occur in deficiency of ornithine
transcarbamoylase, a urea cycle enzyme.
 It leads to accumulation of carbamoyl phosphate, which is diverted for enhanced
synthesis of orotic acid.
 Increased urinary excretion of this compound follows, hence this defect is
considered as a secondary orotic aciduria, also termed Rey’s syndrome.
Metabolic disorders of purines and
pyrimidine metabolism
 Pseudouridine (ψ) is excreted unchanged
 ψ is an isomer of the nucleoside uridine in which uracil is attached to pentose
sugar via a C-C bond instead of a N-C glycosidic bond.
 This C-C bonding provides it more rotational freedom and conformational
flexibility.
 ψ is the most abundant RNA post-transcriptional modification in cellular RNA.
 No human enzyme catalyzes hydrolysis or phosphorolysis of ψ derived from
degradation of RNA.
 This unusual nucleotide is excreted unchanged in urine of normal subjects.
 This is first isolated from human urine.
Question (1): A 42-year old male patient undergoing radiation therapy for prostate cancer develops severe
pain in metatarsal phalangeal joint of his right big toe. Monosodium urate crystals are detected by polarized
light microscopy in fluid obtained from this joint by arthrocentesis.
•This patient’s pain is directly caused by the overproduction of the end product of which of the following metabolic
pathway?
•Name the end product and linked disease?
•Which enzyme is directly linked for the overproduction of this end product?
•What treatments are available for the relief from the joint pain?
•Which enzyme human lacks for production of soluble end product?

Question (2): A 1-year old patient is lethargic, weak and anemic. Her height and weight are low for her age.
Her urine contains an elevated level of orotic acid. Activity of uridine monophosphate synthase is low.
•Name the disease based on symptoms?
•Which metabolic pathway is linked to this disease?
•Administration of which nucleotide alleviate her symptoms?
•Name the enzymes whose deficiency leads to this disease?
•Name the drugs enhancing excretion of orotic acid?
•Which enzymes are targeted by such drugs?