Nucleotides, Purine, and Pyrimidine Metabolism

Nucleotide Basics

• Nucleotides consist of a phosphate group, a 5-carbon sugar (deoxyribose in DNA, ribose in RNA), and a nucleobase.
• The nucleobases are divided into two categories: purines (adenine, guanine) and pyrimidines (cytosine, thymine in DNA, uracil in RNA).
• The suffix '-osine' is used for purine nucleosides, while ‘-idine’ is used for pyrimidine nucleosides.
• Nucleotides are precursors for nucleic acids, playing a role in DNA and RNA synthesis.
• Nucleotides are important in energy metabolism through ATP, GTP, UTP, and CTP.
• Nucleotides are intracellular messengers like cAMP and cGMP.
• Nucleotides function as structure for various coenzymes like S-adenosylmethionine, NADH, NADPH, and FADH2.

Purine and Pyrimidine Metabolism

• Human tissues can synthesize purines and pyrimidines, making them nonessential in the diet.
• Purine bases oxidize to uric acid, which the body excretes through urine.
• The liver serves as the major location for purine nucleotide biosynthesis.
• Brain tissue has a low level of PRPP amidotransferase and depends on exogenous purines.
• Erythrocytes and polymorphonuclear leukocytes cannot synthesize PRPP and use exogenous purines.

Purine Nucleotide Synthesis (De Novo)

• The synthesis of purines begins with activated sugar (PRPP).
• Synthesis leads to Inosine 5'-monophosphate (IMP). which is the first fully formed purine nucleotide.
• The process of making purines involves these steps:
  • Synthesis of 5-phosphoribosyl-1-pyrophosphate (PRPP)
  • Synthesis of 5-phosphoribosylamine (PRA)
  • Synthesis of inosine monophosphate (IMP)
  • Synthesis of adenosine and guanosine monophosphate (AMP and GMP)
  • Conversion of nucleoside mono-, di-, and triphosphates.
• PRPP participates in synthesizing and salvaging purines and pyrimidines; it is an activated pentose.
• Glutamine:phosphoribosylpyrophosphate amidotransferase is inhibited by purine 5-nucleotides AMP and GMP.
• The enzyme regulates, and is significant for, purine nucleotide synthesis.

IMP Synthesis Requirements

• The formation of IMP requires the following:
  • 5 moles of ATP
  • 2 moles of glutamine
  • 1 mole of glycine
  • 1 mole of CO2
  • 1 mole of aspartate
  • 2 moles of formate

AMP and GMP Synthesis

• AMP synthesis needs guanosine triphosphate (GTP) as an energy source.
• GMP synthesis requires ATP.

Nucleoside Conversion

• Nucleoside diphosphates (NDP) are from their corresponding nucleoside monophosphates (NMP) using base-specific nucleoside monophosphate kinases.
• Nucleoside triphosphates (NTP) are from the corresponding nucleoside diphosphates using base-specific nucleoside diphosphate kinases.

Purine Deoxyribonucleotide Synthesis

• Ribonucleotide reductase is specific for reducing purine nucleoside diphosphates as well as pyrimidine nucleoside diphosphates to their deoxy forms.
• To continue producing deoxyribonucleotides, the disulfide bond created must be reduced by ribonucleotide reductase.
• Allosteric sites on enzymes regulate the activity, with the enzymes needed for deoxyribonucleotide synthesis.
• Ribonucleotide reductase helps balance deoxyribonucleotides needed for DNA synthesis.

Folate Deficiency

• Folate deficit results in defective DNA synthesis.
• Folate deficiency increases the risk of congenital defects, including neural tube defects (e.g., spina bifida).

Purine Nucleotide Salvage

• Purine nucleotide salvage involves the following enzymes:
  • Adenosine phosphoribosyltransferase (APRT): adenine + PRPP → AMP + PPi
  • Hypoxanthine-guanine phosphoribosyltransferase (HGPRT): hypoxanthine + PRPP → IMP + PPi
  • Hypoxanthine-guanine phosphoribosyltransferase (HGPRT): guanine + PRPP → GMP + PPi
• Salvage requires PRPP (Pentose Phosphate Pathway).
• Salvage decreases purine degradation and uric acid levels.

Lesch-Nyhan Syndrome (LNS)

• LNS occurs due to X-linked hereditary HGPRT gene deficiency.
• LNS leads to an accumulation of uric acid in body fluids.
• The buildup of uric acid is associated with gout, inflammatory arthritis, and kidney problems.
• HGPRT shows its highest activity in the brain and testis.
• Symptoms include poor muscle control and intellectual disabilities.
• Striking features include lip/finger biting, involuntary writhing, grimacing, and repetitive arm/leg movements.

Purine Synthesis as a Drug Target

• Sulfonamides are bacteriostatic antibiotics that inhibit rapidly dividing bacteria.
• Sulfonamides don't interfere with human cell processes.
• Other purine synthesis inhibitors like methotrexate control spread of cancer by interfering with nucleotide synthesis.
• This affects DNA and RNA production in tumor cells.

Pyrimidine Nucleotide Synthesis (De Novo)

• Orotic acid is the first fully formed pyrimidine base.
• These steps occur in pyrimidine synthesis:
  • Synthesis of carbamoyl phosphate
  • Synthesis of orotic acid
  • Formation of a pyrimidine nucleotide
  • Synthesis of cytidine triphosphate

Carbamoyl Phosphate Synthesis

• Carbamoyl phosphate synthetase (CPS) II catalyses the reaction.
• Uridine triphosphate (UTP) inhibits CPS II, while PRPP activates it.

Orotic Acid Synthesis

• Aspartate transcarbamoylase catalyzes the process.

Pyrimidine Nucleotide Formation

• PRPP serves as the ribose 5-phosphate donor.
• OMP converts to uridine monophosphate (UMP) by orotidylate decarboxylase.
• UMP is then sequentially phosphorylated to UDP and UTP.

Cytidine Triphosphate Synthesis

• Cytidine triphosphate (CTP) produces from the amination of UTP with the help of CTP synthetase.
• Glutamine provides the nitrogen for amination.

Thymidylate Synthesis

• Mechanisms to target cancer include:
  • Direct inhibition of thymidylate synthase
  • Indirect inhibition of dihydrofolate reductase

Direct Inhibition

• Thymidylate synthase: Suicide substrates (e.g., 5-fluorouracil) irreversibly inhibit the enzyme.
• Because dTTP is used only in DNA, can target rapidly proliferating cells
• This doesn't affect non-proliferative cells.

Indirect Inhibition

• Inhibitors of dihydrofolate reductase, like methotrexate, aminopterin, and trimethoprim, are folic acid analogues.
• Indirect inhibition rapidly kills replicating cells.

Catabolism of Purine Bases

• Uric acid is produced in the liver and excreted by the kidneys as the end product of purine catabolism.
• Monosodium urate is soluble in plasma at pH > 5.75.
• Uric acid exhibits limited solubility in urine (pH < 5.75).
• Acidic urine or high calcium levels can lead to coprecipitation with calcium salts, which can form stones in the kidney or bladder.

ADA Deficiency

• ADA (adenosine deaminase) transforms adenosine to inosine.
• ADA deficiency causes impaired immune response. As B and T lymphocytes are destroyed by.
• ADA deficiency is inherited, is autosomal recessive, and affects 1/2,000,000 individuals.
• It causes Primary immune deficiencies (Severe combined immunodeficiency disease-SCID).