purine and pyrimidine

1. Metabolism of nucleotide

2. Structure of purine
Bases + sugar +
phosphate
Bases;---- nucleotides
Purine (DNA &
RNA)
adenine
guanine

4. Nucleosides

5. Digestion & absorption of nucleic acid
dietary nucleic acid
DNA RNA
Polynucleotidase
mononucleotide mononucleotide
Pi Phosphatases Pi
nucleoside Absorbed nucleoside

6. Nucleoside Absorbed nucleoside
H₂O H₂O
Phosphorylase
Bases + deoxyribose Bases + ribose
oxidation
Excreted Excreted
( uric acid) (CO₂ & ammonia)

8. • Purines & Pyrimidines are dietary nonessential
components .
• Body can synthesize Purines & Pyrimidine
nucleotides via de novo.
• Built up as nucleotides.
• Purines are first synthesized as nucleotide
inosonic acid.
• Purine ring is built on ribose -5-P.

9. Biosynthesis of Purine nucleotides
• Two parent Purine nucleotides of nucleic acids
are :-
- Adenosine monophosphate (AMP)
- Guanosine monophosphate (GMP)
• Synthesized by two pathways :-
- De novo pathway
- Salvage pathway

10. Source of carbon & nitrogen in Purine
ring

11. De novo synthesis of Purine
• Precursors for the De novo synthesis of Purine
- Glycine provides C-4 ,C-5 , N-7
- Aspartate provides N-1
- Glutamate provides N-3 & N-9
- Tetrahydrofolate C-2 & C-8
- Carbon dioxide C-6
- PRPP( phosphoribosyl pyrophosphate)
- ATP

12. Formation & importance of PRPP
glucose -6-P
P.P pathway
Ribose -5-P
Purine PRPP Pyrimidine
NAD NADP

13. Ribose-5-phosphate Phosphoribosyl
pyrophosphate (PRPP)
PRPP synthetase
ATP AMP

14. Major steps of De Novo synthesis
• Location :- liver
• Precursor :- ribose -5-P
• Major enzyme :- synthases, transferases,
carboxylases & hydroxylases
• Importance of PRPP
- PRPP required for both synthesis of Pyrimidine
& Purine.
- Intermediate in the Purine salvage pathway &
synthesis of NAD & NADP.

15. De novo synthesis of purine
D- ribose -5-P
ATP PRPP synthase
AMP Mg+
PRPP
glutamine PRPP-glutamyl
glutamate amidotransferase
5-phosphoribosylamine(PRA)

16. 5-phosphoribosylamine(PRA)
glycine phosphoribosyl
glycinamide synthetase
glycinamide ribosyl-5-phosphate(GAR)
N-10-formylTHF formyltransferase
THF
formylglycinamide ribosyl-5- phosphate(FGAR)

17. Step 3 contd.

18. Step 4

19. Step 5

20. Step 6

25. •Summary of steps of
purine synthesis

27. formylglycinamide ribosyl-5- phosphate
glutamine synthase
glutamate
formylglycinamidine ribosyl-5- phosphate
ATP synthase
(ring close) ADP
5-Aminoimidazole ribosyl-5-phosphate(AIR-P)

28. 5-Aminoimidazole ribosyl-5-phosphate
Co₂ carboxylase
Aminoimidazole carboxylate ribosyl-5-P
Aspartate synthase
H₂O
Aminoimidazole succinyl carboxyamide ribosyl-
5-phosphate

29. Aminoimidazole succinyl carboxyamide R-5-P
adenylsuccinase fumarate
Aminoimidazole carboxyamide R-5-P
formyl-THF formyl transferase
THF
formiminoimidazole carboxyamide R-5-P

30. formiminoimidazole carboxyamide R-5-P
H₂O IMP cyclohydrolase
inosine monophosphate (IMP)

33. Conversion of IMP to AMP,GMP
inosine monophosphate (IMP)
Adenosuccinate IMP dehydrogenase
synthase
Adenylosuccinate xanthosine monophosphate
adenylosuccinase glutamine guanosine-
fumarate glutamate P synthase
AMP GMP

34. Inhibitors of de novo synthesis
• Inhibitors :- Purine synthesis
• Sulfonamides :- structural analogs of para-
aminobenzoic acid (PABA).
- Inhibit the synthesis of folic acid .
• Methotrexate :- analogs of folic acid.
- Control cancer.
• 6-mercaptopurine :- inhibits the synthesis of
AMP & GMP.

35. Salvage pathway
• Salvage pathway :- Pathway involved in the
conversion of Purines , Purines ribonucleoside &
Purine deoxy ribonucleosides to
mononucleotides .
• It consist of single reaction.
• Free Purine & Pyrimidine bases are formed in
cells during the metabolic degradation of nucleic
acid are salvaged & used for resynthesis of
nucleotides.
• Advantage :- required less energy and metabolic
intermediate. Eg:- brain , RBCs and neutrophils.

36. • Two mechanism for salvage reactions:-
• Phosphoribosylation of purine bases.
( conversion of free purines bases to nucleotides
which are produced by degradation process.)
• Phosphorylation of Purine nucleosides into
nucleotides .

37. Phosphoribosylation of purine bases.
Adenine
PRPP adenine phosphoribosyl
PPi transferase ( APRTase)
Adenosine monophosphate (AMP)

38. • Hypoxanthine Guanine
PRPP Hypoxanthine –Guanine PRPP
PPi phosphoribosyl PPi
transferase(HGPRTase)
-
IMP AMP, GMP GMP
• Defect in the enzyme HGPRT causes lesch-
Nyhan syndrome.

39. Phosphorylation of Purine nucleosides
ATP kinase
Purine nucleoside Purine nucleotide
• Synthesis of deoxyribonucleotides
Ribonucleotide
reduced thioredoxin reductase
oxidized thioredoxin -
deoxyribonucleotide dATP, dGTP

40. Salvage pathway of Purine
• Significance :- salvage pathway provides
Purine nucleotides for tissues, incapable of
their biosynthesis by de novo pathway.
- eg:- human brain has low level of PRPP amido-
transferase . It depends in part on exogenous
purines.
- Erythrocytes & polymorphonuclear leukocytes

41. Regulation of de novo synthesis
• Regulated by :-
 Concentration of PRPP :- increased PRPP
levels stimulates the purine synthesis.
- PRPP depends on (i). availability of ribose-5-P
(ii). On the activity of PRPP synthase.
 feedback regulation

42. • Ist step of de novo synthesis is catalyzed by
allosteric enzyme PRPP synthase inhibited by
AMP, GMP & IMP.
• GMP & AMP inhibit HGPRTase by feedback
mechanism.
• Cross regulation between the pathways of IMP
metabolism serves to decrease synthesis of
one purine nucleotide when there is
deficiency of the other nucleotide .

43. Questionnaire ??
• Q1. Diagrammatically represent the sources of
purine ring ?
• Q2.What is the importance of PRPP ?
• Q3.Explain the pathway of De novo synthesis
of purine ?
• Q4. Importance salvage pathway of purine

44. Catabolism of purine
• End product of purine :- uric acid
• Mammals oxidized uric acid further to allantoin .
Some animals further degraded allantoic acid to
produce urea or even ammonia.
• Significance of uric acid :-
- Final excretory product in humans.
- Role as potent antioxidant.
- Very effective scavenger of free radicals .

45. • AMP GMP
H₂O nucleotidase
Pi
Adenosine guanosine
H₂O Adenosine deaminase Pi
NH₃ phosphorylase
r-1-P
Inosine guanine

46. • Inosine guanine
Pi phosphorylase guanase
R-1-P
Hypoxanthine xanthine oxidase xanthine
H₂O+O₂ H₂O₂ XO
Ammonia allantoin uricase uric acid
(mammals) (human beings)

47. Disorders of purine catabolism
• Catabolism of purine to give uric acid.
• Uric acid , at physiological pH ionized &
present in plasma as sodium urate.
• Elevated levels of serum urate is known as
hyperuricaemia .
• Normal range :- 4 to 7 mg/dl.

48. Factors affecting of uric acid
• De novo synthesis of purine .
• Metabolism of DNA , RNA & other purine
containing molecule such as ATP.
• Breakdown of nucleic acid.

49. Excretion of uric acid
• Excreted in two ways:-
- Kidney :- majority of uric acid.
- Gut :- smaller amount of urate excreted ,
where it is broken down by bacteria is called
uricolysis.

51. Gout
• Metabolic disease.
• Impaired excretion or overproduction of uric
acid.
• Uric acid crystals precipitate into joints (Gouty
Arthritis), kidneys, ureters (stones)
 Gout is associated with hyperuricaemia but
hyperuricaemia is not associated with gout.

52.  History :- over eating & alcohol consumption.
Prevalence :- 3 per 1000 persons, mostly
affecting males.
 Classification :- into two types :-
- Primary gout
- Secondary gout

53. Primary gout
• Inborn error of metabolism caused by
defective enzymes of purine biosynthesis.
( increase synthesis of purine nucleotides)
 Loss of feedback regulation
 ↑ PRPP synthase ↑ purines
↑ PRPP glutamyl amidotransferase
 Deficiency of HGPRTase:- ↑PRPP ↑ de -
novo synthesis, salvage pathway

54.  glucose -6-phosphatase deficiency :- type- I
Von gierke’s disease
- Glucose -6-P glucose
HMP shunt ribose -5-P PRPP ↑purine
Elevation of glutathione reductase

56. Sign & Symptoms
• Patients often show deposition of urate as
tophi ( clusters of urate crystals) in soft tissue
that affects the joints & leads to painful
arthritis.
• The kidneys are also affected , due to excess
urate deposition in the tubules & leads to
renal failure.

58. Secondary gout
• Due to variety of diseases that cause an
defective elimination of uric acid.
• Overproduction of urate due to increased
destruction of cells.
- Myeloproliferative disorders eg;- leukemia ,
polycythemia
- Cytotoxic drug therapy , renal disease
- Psoriasis

59. Suicide inhibition
• Irreversible inhibition.
• Original inhibitor is converted to a more
potent form by the same enzyme .
• Allopurinol is oxidized to alloxanthine by
xanthine oxidase. Alloxanthine is more
effective inhibitor of xanthine oxidase.
• Inhibition of xanthine oxidase leads to the
accumulation of hypoxanthine & xanthine .

60. Treatment of gout
• Treated by nutritional therapy & drug
therapy:-
- Avoid nucleotides rich diets.
- Consumption of plenty of water.
- Allopurinol drug ;- analogs of hypoxanthine
which inhibits xanthine oxidase , reduces the
uric acid .
- NSAIDS :- non steroid anti-inflammatory drugs
eg;- phenylbutazone & corticosteroids.

61. xanthine oxidase
Allopurinol alloxanthine
- - -
xanthine oxidase
xanthine uric acid
more soluble than uric acid easily excreted

62. Pseudogout
• Clinical manifestation similar to gout.
• Caused by deposition of calcium
pyrophosphate crystals in the joints.
• Serum uric acid conc. is normal .

63. Lesch- Nyhan syndrome
• X- linked metabolic disorder, affect only males.
• Enzyme deficiency :- HGPRTase
 in the absence of HGPRTase , salvage
pathway is inoperative & Purines cannot be
reconverted to nucleotides , instead they are
degraded to uric acid.
Lack of HGPRTase also cause an
overproduction of PRPP , purine synthesis
& uric acid.

64. Symptoms
• Hyperuricaemia
• Gout
• Urinary tract stones
• Neurological symptoms of mental retardation.
• Spasticity
• Self mutilation
 treatment :- allopurinol

65. Xanthinuria
• Enzyme deficiency :- xanthine oxidase
 genetic defect :-
 severe liver damage
 ( due to hypouricaemia & excretion of
xanthine & hypoxanthine )
 Renal damage

66. • Defect in adenosine deaminase (ADA)
–immunodeficiency Diseases Associated
with Purine Degradation.
• Defect in both B-cells and T-cells (Disease of
Lymphocytes)
• in the cells of patients with ADA deficiency ,
deoxy-adenosine & adenosine abundant ,
because they do not degrade to deoxyinosine .
SCID- severe combined
immunodeficiency

67. • Immune dysfunction is due to high levels of
ATP.
• Autosomal recessive disorders with symptoms
of recurrent & chronic infection. SCID is
usually fatal, often 18 months of age.

68. Therapies for SCID
• Can be diagnosed in infants through a simple
blood test (white cell count)
• Bone marrow transplant for infants
– Familial donor
• Continued administration of adenosine
deaminase (ADA-PEG)
• Gene therapy- repair defective gene in T-cells
or blood stem cells

69. Questionnaires
• Note on catabolism of purine ?
• Explain gout
• Note on lesch –nyhan syndrome
• Note on SCID

70. Multiple choice question
• 1. An amino group donated by glutamine is attached at
C-1 of PRPP, this results in
a) 5-phosphoribosylamine
b) 4-phosphoribosylamine
c) 3-phosphoribosylamine
d) 2-phosphoribosylamine
2. The first intermediate with a complete purine ring is
a) Inosinate
b) Formate
c) Aspartate
d) Glycine

71. • 3. Salvage pathway is used in the synthesis of
___________
a) Amino acid
b) Carbohydrate
c) Nucleotide
d) Fatty acid
• 4. All deoxyribonucleotide is synthesized by the
ribonucleotides.
a) True
b) False
• 5.Name the precursor of RNA?
a) Glutamine
b) Cytidine
c) Orotidylate
d) Uridylate

72. • 6. Which of the following is not the precursor of a purine ring?
a) Glutamine
b) Lysine
c) Glycine
d) Aspartate
7. During purine synthesis the activity of amidotransferase
enzyme is inhibited by the antitumor agent, mark the correct
one.
a) Aminopterin
b) Methotrexate
c) Texol
d) Azaserine
8. What is the final product of purine degradation in mammals?
a) Guanine
b) Inosine
c) Uric acid
d) Hypoxanthine

73. 9. Which of the following disorder is caused due to
the high serum level of urate?
a) Gout
b) Galectosemia
c) Cystic fibrosis
d) Maple syrup urine disease
10.Name the genetic disorder which is caused by the
deficiency of enzyme HGPRT?
a) SCID
b) Lesch-Nyhan syndrome
c) Cystic fibrosis
d) Down syndrome

74. 11.Name the deficiency in which T and B lymphocyte do not develop
properly?
a) XLA
b) CVID
c) SCID
d) Multiple myeloma
12. Three amino acids that donate amine groups for the purine
biosynthesis are
• a) Glycine, glutamine, aspartate
• b) Glycine, beta-alanine, aspartate
• c) Glycine, alanine, aspartate
• d) Lysine, glutamine, aspartate
• e) Lysine, glutamate, asparagine
• f) Lysine, glycine, asparagine
•
•

75. 13.Gout is characterized by elevated uric acid
concentrations in blood and urine due to a variety of
metabolic abnormalities that lead to the
overproduction of purine nucleotides. Allopurinol is
used in the treatment of gout because this drug, and its
metabolic product, alloxanthine, act as inhibitors of:
• a) Xanthine Oxidase
• b) PRPP synthetase
• c) Adenyl succinate synthase
• d) Hypoxhantine guanine phosphoribosyl transferase
• e) Nucleotides

76. Answer key
• Ans. 1-------a
• Ans.2-------a
• Ans.3-------c
• Ans.4-------a
• Ans.5-------c
• Ans.6-------b
• Ans.7-------d
• Ans.8-------c
• Ans.9-------a
• Ans.10------b
• Ans.11------c
• Ans.12------a
• Ans. 13-----a

77. De novo synthesis of Pyrimidine
• Uridine Monophosphate (UMP)
• Cytidine monophosphate (CMP)
• Thymidine monophosphate (TMP)
• Unlike purine , six membered ring is made first
& then attached to ribose phosphate (PRPP).

78. • Synthesized from:
– Glutamine provides N₃
– CO2 - C₂
– Aspartic acid – C₄, C₅, C₆ & N₁
– Requires ATP
Precursors of Pyrimidines

79. • Pyrimidine rings are synthesized independent
of the ribose and transferred to the PRPP
(ribose).
• Generated as UMP (uridine 5’-
monophosphate) first.

80. Co₂ + glutamine + ATP
UTP - carbamoyl phosphate synthase-II
PRPP + (CPS-II)
carbamoyl phosphate (CP)
aspartate transcarbamoylase
carbamoyl aspartate (CA)

81. carbamoyl aspartate (CA)
DIHYDROOROTASE
Dihydroorotate (DHO)
dehydrogenase
Orotate

82. Orotate
phosphoribosyl transferase
PRPP
orotidine monophosphate
CO₂ decarboxylase
uridine monophosphate (UMP)
kinase
UDP

83. UDP reductase
UTP dUDP
CTP synthase Pi
glutamine
CTP dUMP
thymidylate synt.
dTMP

84. Regulation of deoxyribonucleotide
formation

85. Catabolism of Pyrimidines
cytosine
NH₃
Uracil Thymine
Dihydrouracil Dihydrothymidine
alanine
CO₂ + NH₃

86. Disorders of Pyrimidine synthesis
• Orotic aciduria :-
- Enzyme deficiency :- orotate phosphoribosyl
transferase & decarboxylase
- Symptoms:- retarded growth, severe anemia,
excretion of orotic acid in the urine.
• Clinical Features
• Retarded growth
• Severe megaloblastic anaemia
• Crystals excreted in urine

87. Disorders of pyrimidine metabolism