The document provides an overview of nucleic acid metabolism and the synthesis of purines and pyrimidines. It discusses how nucleotides serve as building blocks for nucleic acids and how they are synthesized through both de novo and salvage pathways. The key steps in purine synthesis include the production of IMP from PRPP and glutamine, followed by conversion to AMP and GMP. Purines can also be salvaged from nucleic acid breakdown. Deoxyribonucleotides are synthesized from ribonucleotides by the enzyme ribonucleotide reductase. Defects in nucleotide synthesis can cause diseases.

1. 1
“Nucleic Acid Metabolism & Control,
Purine & Pyrimidine Synthesis”
PRESENTED BY:
Nahal Jehangir (18006231-011)
Course Title:
Biochemistry II

2. CONTENTS
 OVERVIEW
 WHAT ARE NUCLEOTIDES & NUCLEOSIDES
 NUCLEIC ACID DIGESTION & METABOLISM
 PURINE NUCLEOTIDE SYNTHESIS
 SYNTHESIS OF DEOXYRIBONUCLEOTIDES
 PURINE CATABOLISM
 PYRIMIDINE SYNTHESIS
 PYRIMIDINE CATABOLISM
2

3. OVERVIEW
 Nucleic acid metabolism is the process by which nucleic acids (DNA and RNA) are
synthesized and degraded.
 Nucleic acids are the polymers of nucleotides.
 Nucleotide synthesis is an anabolic mechanism generally involving the chemical reaction
of phosphate, pentose sugar, and a nitrogenous base.
 Destruction of nucleic acid is a catabolic reaction.
 Additionally, parts of the nucleotides or nucleobases can be salvaged to recreate new
nucleotides.
 Both synthesis and degradation reactions require enzymes to facilitate the event. Defects
or deficiencies in these enzymes can lead to a variety of diseases.
3

4.  Nucleotides serve as building blocks for RNA and DNA.
 Nucleotides can serve as sources of energy (i.e., ATP), physiological signaling mediators (i.e.,
adenosine in control of coronary blood flow), secondary messengers (cAMP and cGMP), and allosteric
enzyme effectors.
 Without them ,Neither RNA nor DNA can be produce and therefore protein cannot be synthesized.
 Nucleotides also serve as carriers of activated intermediates in the synthesis of some carbohydrates ,
lipids and conjugated proteins and also are structural components of several essential coenzymes like
NADH & NADPH.
 Nucleotide such as cAMP & cGMP serve as second messenger in many pathways.
 Also ,nucleotides play an important role as “energy currency” in the cell.
4

5. NUCLEOTIDE
 Nucleotides are the monomers (building blocks) of nucleic acids (RNA / DNA).
 Energy rich compounds that drive metabolic process in cell (ATP).
 Serve as chemical signals, key links in cellular systems that respond to hormones and other
extracellular stimuli.
 Each nucleotide is formed by 3 units:
I. NITROGENOUS BASE
II. SUGAR
III. PHOSPHATE
5

6. 6

7. Nitrogen Bases
 2 types of nitrogenous base
 PURINES & PYRIMIDINES DERIVATIVES
 PURINES: ADENINE AND GUANINE (ATP/
GTP)
 PYRIMIDINE: URACIL,THYMINE AND
CYTOSINE (UTP/TTP/CTP)
 Nitrogen base is attached to 1’ carbon of
sugar.
7

8. Sugars
 5 carbon keto sugar or pentose
 Deoxyribose sugar (DNA)
 Ribose sugar (RNA)
 Both sugar are present in furanose form and beta
configuration
 Pentose sugar form esters with phosphoric acid and is
called phosphodiester bond.
 Phosphate is attached to 5’ carbon of pentose sugar.
8

9. Phosphate Group
 Molecular formula H3PO4
 Contains 3 monovalent hydroxyl group and a divalent oxygen
atom
 All linked to pentavalent phosphorous atom
 The phosphate group attached to the 5' carbon of the sugar on
one nucleotide forms an ester bond with the free hydroxyl on the
3' carbon of the next nucleotide.
 Monophosphate, diphosphate & triphosphate
 AMP / ADP / ATP
9

10. 10

11. NUCLEOSIDE
 When ribose or 2-deoxyribose is combined with purine
or pyramidine base Nucleoside is formed.
 Nucleoside is a nucleotide without phoshphate group
attached
 While a nucleoside is a nucleobase linked to a sugar, a
nucleotide is composed of a nucleoside and one or more
phosphate groups. Thus, nucleosides can
be phosphorylated by specific kinases in the cell on the
sugar's primary alcohol group (-CH2-OH) to
produce nucleotides.
11

12. NAMING CONVENTIONS
 Nucleosides:
Purine nucleosides end in “-sine” (Adenosine, Guanosine)
Pyrimidine nucleosides end in “-dine” (Thymidine, Cytidine, Uridine)
 Nucleotides:
Start with the nucleoside name from above and add “mono-”, “di-”, or
“triphosphate” (Adenosine Monophosphate, Cytidine Triphosphate,
Deoxythymidine Diphosphate)
12

13. NUCLEIC ACID DIGESTION
 The nucleic acids DNA and RNA are found in most
of the foods you eat.
 Two types of pancreatic nuclease are responsible
for their digestion: deoxyribonuclease, which
digests DNA, and ribonuclease, which digests RNA.
 The nucleotides produced by this digestion are
further broken down by two intestinal brush
border enzymes (nucleosidase and phosphatase)
into pentoses, phosphates, and nitrogenous bases,
which can be absorbed through the alimentary
canal wall.
13

14. NUCLEOTIDE METABOLISM
 Nucleotides can be synthesized de novo, or from components “salvaged” from the
degradation products of nucleic acids by salvage pathways.
 2 types of pathways leads to nucleotides:
1. De novo pathway: Begins with their metabolic precursors: amino acids, ribose-5-
phosphate, and CO2. The free bases A, T, G, C, and U are not intermediates.
2. Salvage pathway: Recycle the free bases and nucleosides released from nucleic acid
breakdown. The free bases are intermediates.
 When in excess, nucleotides are degraded to products that can either be consumed by
other pathways or excreted.
 Defects in the pathways for de novo synthesis, salvage, and nucleotides degradation
result in clinical disorders, and many drugs target these pathways.
14

15. A. De Novo Synthesis of Purine Nucleotide
 The atoms of the purine ring are contributed by a number of compounds, including amino
acids (aspartate, glycine, and glutamine), CO2, and N10-formyltetrahydrofolate (Formate).
 The purine ring is constructed primarily in the liver by a series of reactions that add the
donated carbons and nitrogens to a preformed
ribose 5-phosphate.
15

16. Inosine Monophosphate (IMP)
 Inosine Monophosphate (IMP) is the precursor of both AMP &
GMP.
 It is the nucleotide containing the base hypoxanthine.
 IMP is synthesized in an eleven reaction pathway.
16

17. STEP 1: Synthesis of PRPP (5-phosphoribosyl-1-
pyrophosphate)
 5-Phosphoribosyl-1-pyrophosphate (PRPP) is an
“activated pentose” that participates in the synthesis
and salvage of purines and pyrimidines. Synthesis of
PRPP from ATP and ribose 5-phosphate is catalyzed by
PRPP synthetase .
 This enzyme is activated by inorganic phosphate and
inhibited by purine nucleotides (endproduct
inhibition).
17
Glutamine-PRPP Amindotransferase

18. 2. Synthesis of 5-phosphoribosylamine
 Synthesis of 5-phosphoribosylamine from PRPP and glutamine.
 The amide group of glutamine replaces the pyrophosphate
group attached to carbon 1 of PRPP. This is the committed step in
purine nucleotide biosynthesis.
The enzyme, glu-tamine:phosphoribosylpyrophosphate
amidotransferase, is inhibited by the purine 5 -nucleotides AMP
and [GMP], the end products of the pathway.
 The rate of the reaction is also controlled by the intracellular
concentration of PRPP. Any small change in the PRPP
concentration causes a proportional change in rate of the
reaction (acc. To Michaelis Menton eq.)
18
Amidophosphoribosyl
Transferase

19. Synthesis of IMP, the “parent” purine nucleotide
 The next nine steps in purine nucleotide biosynthesis leading to the synthesis of
inosine monophosphate ([IMP] whose base is hypoxanthine).
 Four steps in this pathway require ATP as an energy source, and two steps in the
pathway require N10-formyltetrahydrofolate as a one-carbon donor. (Hypoxanthine is
found in tRNA).
 The next 9 steps out of 11 are elaborated stepwise in next slides.
19

20. GAR Transformylase FGAM Synthetase
STEP 3: Amino group of GAR reacts
with N10-Formyl tetrahydrofolate to
form Formylglyciniamid ribonucletide
(FGAR).
STEP 4: Amide nitrogen of seconf
glutamine is added to FGAR in an ATP
dependant reaction to form
Formylglycinamide nucleotide
(FGAM).
20

21. FGAM Synthetase (Cyclase)
Air Carboxylase (N 5 -CAIR Synthetase)
STEP 5: An ATP dependant ring closing
(imidazol ring formation) to produce 5-
Aminoimidazole ribonucleotide (AIR).
STEP 6: An ATP dependant carboxylation
reaction of AIR with HCO-3 (bicarbonate) to
produce N5- Carboxyaminoimidazole
ribonucleotide (N5-CAIR).
21

22. N 5 -CAIR Mutase SAICAIR Synthetase
STEP 7: Ismerism group transfer
via mutase to form 5-Amino-4-
carboxyaminoimidazole
ribonucletide.
STEP 8: Aspartae is added and it forms
amide bond with C6 to form N-
Succinylo-5-aminoimidazole-4-
carboxamide ribonucleotide (SAICAR).
22

23. SAICAIR (adenylosuccinate) Lyase AICAIR Transformylase
STEP 9: Fumarate group is
cleaved from SAICAR to produce
AICAR.
STEP 10: Amino group of AICAR react with
N10-Formyl H4-folate to produce N-
Formylaminoimidazole- 4-carboxamide
ribonulceotide (FAICAR).
23

24. 11. Formation of IMP by IMP synthase
24
In the last reaction step, the larger
ring of FAICAR is closed
enzymatically to form IMP. This
process is cyclization to form IMP.

25. Synthesis of AMP/GMP from IMP
 The conversion of IMP to either AMP or GMP uses a
two-step, energy-requiring pathway. Note that the
synthesis of AMP requires guanosine triphosphate
(GTP) as an energy source, whereas the synthesis of
GMP requires ATP.
 Also, the first reaction in each pathway is inhibited by
the end product of that pathway. This provides a
mechanism for diverting IMP to the synthesis of the
purine present in lesser amounts. If both AMP and
GMP are present in adequate amounts, the de novo
pathway of purine synthesis is turned off at the
amidotransferase step.
25

26. Synthetic inhibitors of purine synthesis
 Some synthetic inhibitors of purine synthesis (for example, the sulfonamides), are designed
to inhibit the growth of rapidly dividing microorganisms without interfering with human cell
functions.
 Other purine synthesis inhibitors, such as structural analogs of folic acid (for example,
methotrexate), are used pharmacologically to control the spread of cancer by interfering with
the synthesis of nucleotides and, therefore, of DNA and RNA.
 Inhibitors of human purine synthesis are extremely toxic to tissues, especially to developing
structures such as in a fetus, or to cell types that normally replicate rapidly, including those of
bone marrow, skin, gastrointestinal (GI) tract, immune system, or hair follicles. As a result,
individuals taking such anticancer drugs can experience adverse effects, including anemia,
scaly skin, GI tract disturbance, immunodeficiencies, and hair loss.
26

27. Synthesis of Di- and Tri- phosphate
 Nucleoside diphosphates are synthesized from the corresponding nucleoside
monophosphates by base-specific nucleoside monophosphate kinases (These kinases do not
discriminate between ribose or deoxyribose in the substrate).
 ATP is generally the source of the transferred phosphate because it is present in higher
concentrations than the other nucleoside triphosphates.
 Adenylate kinase is particularly active in the liver and in muscle, where the turnover of
energy from ATP is high. Its function is to maintain equilibrium among the adenine
nucleotides (AMP, ADP, and ATP).
 Nucleoside diphosphates and triphosphates are interconverted by nucleoside diphosphate
kinase, an enzyme that, unlike the monophosphate kinases, has broad substrate specificity.
 AMP + ATP → 2 ADP (adenylate kinase) GDP + ATP → GTP + ADP
27

28. Purine Biosynthesis: Control
28

29. B. Salvage pathway for purines
 Purines that result from the normal turnover of cellular nucleic acids, or the
small amount that is obtained from the diet and not degraded, can be converted
to nucleoside triphosphates and used by the body. This is referred to as the
“salvage pathway” for purines.
 Most organisms can synthesize nucleotides from nucleosides or bases that are
made available in the diet or from nucleic acid breakdown.
 Salvage reactions convert free purine and pyrimidine bases into nucleotides.
29

30. 1. Salvage of purine bases to nucleotides
 Two enzymes are involved: adenine phosphoribosyltransferase (APRT) and
hypoxanthine-guanine phosphoribosyltransferase (HGPRT). Both enzymes use
PRPP as the source of the ribose 5-phosphate group.
 The release of pyrophosphate and its subsequent hydrolysis by
pyrophosphatase makes these reactions irreversible. [Note: Adenosine is the
only purine nucleoside to be salvaged. It is phosphorylated to AMP by adenosine
kinase.]
30

31. 31
PURINES AND PYRIMIDINES
BASES: RECYCLING

32. SYNTHESIS OF DEOXYRIBONUCLEOTIDES
 The nucleotides described thus far all contain ribose (ribonucleotides). The
nucleotides required for DNA synthesis, however, are 2 -deoxyribonucleotides,
which are produced from ribonucleoside diphosphates by the enzyme
ribonucleotide reductase during the Sphase of the cell cycle
 Deoxyribonucleotides are synthesized from their correpsonding ribonucleotides
by the reduction of the C2’ OH to H” .
32

33. A. Ribonucleotide Reductase
Ribonucleotide reductase (ribonucleoside diphosphate reductase) is
composed of two nonidentical dimeric subunits, R1 and R2, and is
specific for the reduction of purine nucleoside diphosphates (ADP and
GDP) and pyrimidine nucleoside diphosphates (CDP and UDP) to their
deoxy forms (dADP, dGDP, dCDP, and dUDP).
 The immediate donors of the hydrogen atoms needed for the reduction
of the 2 -hydroxyl group are two sulfhydryl groups on the enzyme itself,
which, during the reaction, form a disulfide bond.
There are four classes of ribonucleotide reductases – all use a free
radical mechanism. Fe-containing enzyme in most eukaryotes.
33

34. 34

35. 1. Regeneration of reduced enzyme:
In order for ribonucleotide reductase to continue to produce deoxyribonucleotides, the
disulfide bond created during the production of the 2 -deoxy carbon must be reduced. The
source of the reducing equivalents for this purpose is thioredoxin, a peptide coenzyme of
ribonucleotide reductase. Thioredoxin contains two cysteine residues separated by two
amino acids in the peptide chain. The two sulfhydryl groups of thioredoxin donate their
hydrogen atoms to ribonucleotide reductase, forming a disulfide bond in the process.
2. Regeneration of reduced thioredoxin:
Thioredoxin must be converted back to its reduced form in order to continue to perform its
function. The necessary reducing equivalents are provided by NADPH + H+, and the reaction
is catalyzed by thioredoxin reductase.
35
Ribonucleotide Reductase

36. B. Regulation of deoxyribonucleotide synthesis
Ribonucleotide reductase is responsible for maintaining a balanced supply of the
deoxyribonucleotides required for DNA synthesis. To achieve this, the regulation of the enzyme is
complex. In addition to the catalytic (active) site, there are allosteric sites on the enzyme involved in
regulating its activity.
1. Activity sites:
The binding of dATP to allosteric sites (known as the activity sites) on the enzyme inhibits the overall
catalytic activity of the enzyme and, therefore, prevents the reduction of any of the four nucleoside
diphosphates. In contrast, ATP bound to these sites activates the enzyme.
2.Substrate specificity sites:
The binding of nucleoside triphosphates to additional allosteric sites (known as the substrate
specificity sites) on the enzyme regulates substrate specificity. For example, deoxythymidine
triphosphate binding at the specificity sites causes a conformational change that allows reduction of
GDP to dGDP at the catalytic site.
36

37. PURINE NUCLEOTIDE CATABOLISM
Degradation of dietary nucleic acids occurs in the small intestine, where a family
of pancreatic enzymes hydrolyzes the nucleic acids to nucleotides. Inside the
intestinal mucosal cells, purine nucleotides are sequentially degraded by specific
enzymes to nucleosides and free bases, with uric acid as the end product of this
pathway.
A. Degradation of dietary nucleic acids in the small intestine
B. Formation of uric acid
37

38. A. Degradation of dietary nucleic acids in the small
intestine
 Ribonucleases and deoxyribonucleases, secreted by the pancreas, hydrolyze dietary
RNA and DNA to oligonucleotides.
 Oligonucleotides are further hydrolyzed by pancreatic phosphodiesterases, producing
a mixture of 3 - and 5 -mononucleotides.
 In the intestinal mucosal cells, a family of nucleotidases removes the phosphate groups
hydrolytically, releasing nucleosides that are further degraded by nucleosidases to free
bases plus (deoxy) ribose 1-phosphate.
 Dietary purine bases are not used to any appreciable extent for the synthesis of tissue
nucleic acids. Instead, they are generally converted to uric acid in intestinal mucosal
cells.
 Most of the uric acid enters the blood and is eventually excreted in the urine.
38

39. B. Formation of uric acid
1) An amino group is removed from AMP to produce IMP by AMP deaminase or
from adenosine to produce inosine (hypoxanthine-ribose) by adenosine
deaminase (ADA).
2) IMP and GMP are converted into their nucleoside forms (inosine and
guanosine) by the action of 5 -nucleotidase.
3) Purine nucleoside phosphorylase converts inosine and guanosine into their
respective purine bases, hypoxanthine and guanine.
4) Guanine is deaminated to form xanthine.
5) Hypoxanthine is oxidized by xanthine oxidase to xanthine, which is further
oxidized by xanthine oxidase to uric acid, the final product of human purine
degradation. Uric acid is excreted primarily in the urine.
39

40. 40

41. C. Diseases associated with Purine Degradation
1. Gout:
 Gout is a disorder initiated by high levels of uric acid in blood (hyperuricemia), as a result of either
the overproduction or underexcretion of uric acid.
 The hyperuricemia can lead to the deposition of monosodium urate (MSU) crystals in the joints
and an inflammatory response to the crystals, causing first acute and then progressing to chronic
gouty arthritis.
 Nodular masses of MSU crystals (tophi) may be deposited in the soft tissues, resulting in chronic
tophaceous gout. Formation of uric acid stones in the kidney (urolithiasis) may also be seen.
 The definitive diagnosis of gout requires aspiration and examination of synovial fluid from an
affected joint (or material from a tophus) using polarized light microscopy to confirm the presence
of needleshaped MSU crystals.
41

42. 2. Adenosine deaminase (ADA) deficiency:
 Adenosine deaminase (ADA) is expressed in a variety of tissues, but, in humans, lymphocytes have
the highest activity of this cytoplasmic enzyme.
 A deficiency of ADA results in an accumulation of adenosine, which is converted to its
ribonucleotide or deoxyribonucleotide forms by cellular kinases.
 As dATP levels rise, ribonucleotide reductase is inhibited, thereby preventing the production of all
deoxyribose-containing nucleotides. Consequently, cells cannot make DNA and divide.
 In its most severe form, this autosomal-recessive disorder causes a type of severe combined
immunodeficiency disease (SCID), involving a decrease in T cells, B cells, and natural killer cells.
 Treatments include bone marrow transplantation, enzyme replacement therapy, and gene therapy.
42

43. PYRIMIDINE SYNTHESIS
 Unlike the synthesis of the purine ring, which is constructed on a preexisting
ribose 5phosphate, the pyrimidine ring is synthesized before being attached to
ribose 5phosphate, which is donated by PRPP.
 The sources of the atoms in the pyrimidine ring are glutamine, CO2, and
aspartate.
43

44. De Novo Pyrimidine Synthesis
STEP 1: Synthesis of carbamoyl phosphate:
 The regulated step of this pathway in mammalian cells is the synthesis of
carbamoyl phosphate from glutamine and CO2, catalyzed by carbamoyl
phosphate synthetase (CPS) II.
 CPS II is inhibited by uridine triphosphate (the end product of this pathway,
which can be converted into the other pyrimidine nucleotides), and is activated
by PRPP. Note that Carbamoyl phosphate, synthesized by CPS I, is also a
precursor of urea.
 Defects in ornithine transcarbamylase of the urea cycle promote pyrimidine
synthesis due to increased availability of carbamoyl phosphate.
44

45. STEP 2: Synthesis of orotic acid
 The second step in pyrimidine synthesis is the formation of carbamoylaspartate, catalyzed by
aspartate transcarbamoylase.
 The pyrimidine ring is then closed by dihydroorotase.
 The resulting dihydroorotate is oxidized to produce orotic acid (orotate). The enzyme that
produces orotate, dihydroorotate dehydrogenase, is a flavoprotein associated with the inner
mitochondrial membrane. All other enzymes in pyrimidine biosynthesis are cytosolic.
45

46. STEP 3: Formation of a pyrimidine nucleotide
 The completed pyrimidine ring is converted to the nucleotide orotidine monophosphate
(OMP) in the second stage of pyrimidine nucleotide synthesis. PRPP is again the ribose 5-
phosphate donor.
 The enzyme orotate phosphoribosyltransferase produces OMP and releases pyrophosphate,
thereby making the reaction biologically irreversible. OMP, the parent pyrimidine
mononucleotide, is converted to uridine monophosphate (UMP) by orotidylate
decarboxylase, which removes the carboxyl group.
 Orotate phosphoribosyltransferase and orotidylate decarboxylase are also catalytic domains
of a single polypeptide chain called UMP synthase.
 UMP is sequentially phosphorylated to UDP and UTP.
46

47. 47

48. Synthesis of CTP from UTP
Cytidine triphosphate (CTP) is produced by amination of UTP by CTP
synthetase, with glutamine providing the nitrogen.
48

49. Pyrimidine Biosynthesis: Control
 In animals:
 pyrimidine biosynthesis is controlled by the activity of
carbamoyl phosphate synthetase II.
 Inhibited by UDP and UTP Activated by ATP and PRPP.
 In mammals:
 additional control at OMP decarboxylase.
 UMP & CMP are competitive inhibitors.
49

50. Degradation and salvage of Pyrimidines
 Unlike the purine ring, which is not cleaved in humans, the pyrimidine ring is
opened and degraded to highly soluble products, β-alanine (from the
degradation of CMP and UMP) and β-aminoisobutyrate (from TMP degradation),
with the production of NH3 and CO2.
 Pyrimidine bases can be salvaged to nucleosides, which are phosphorylated to
nucleotides. However, their high solubility makes pyrimidine salvage less
significant clinically than purine salvage.
50

51. 51

52. 52

53. 53

54. PYRIMIDINE CATABOLISM
54

55. Disorders associated with Pyrimidine Metabolism
Since the end product of pyrimidine catabolism are highly water soluble, overproduction of pyrimidine
catabolites is rarely associated with clinically significant abnormalities.
Orotic Aciduria:
 It is a hereditary disorder. It is results from defective in the multifunctional enzyme UMP synthase
that converts orotic acid to UMP (in pyrimidine synthesis).
 Results in the excretion of orotic acid in the urine. UMP synthase is multifunctional enzyme
containing both orotate PRTase & orotidylate decarboxylase activity.
 These are of two types; Type I in which both enzymes are deficient. Type II in this only orotidylate
carboxylase is deficient. The deficiency in UMP & other pyrimidine nucleotides results in inhibition
of DNA & RNA synthesis. This causes megaloblastic anaemia & growth retardation.
 This can be treated by feeding cytidine and uridine.
55

56. SUMMARY OF NUCLEOTIDE METABOLISM
56

57. THANKS FOR YOUR ATTENTION!
57
REFRENCES
• Data taken from Lipincott Book of Biochemistry.
• https://www.cliffsnotes.com/study-guides/biology/biochemistry-ii/purines-and-
pyrimidines/salvage-and-biosynthetic-
pathways#:~:text=Salvage%20reactions%20convert%20free%20purine,amino%20acids%20fou
nd%20in%20proteins
• https://slideplayer.com/slide/3556008/