De Novo Nucleotide Synthesis

Overview

De novo nucleotide synthesis is the biochemical pathway through which cells synthesize new nucleotides from simple precursor molecules. Nucleotides are essential building blocks of DNA and RNA, and they play vital roles in cellular processes such as DNA replication, protein synthesis, and cell signaling.

The de novo synthesis of nucleotides involves several enzymatic reactions and metabolic pathways. There are separate pathways for purine and pyrimidine nucleotide synthesis, although they share some common intermediate molecules.

The de novo synthesis of nucleotides involves several enzymatic reactions and metabolic pathways. There are separate pathways for purine and pyrimidine nucleotide synthesis, although they share some common intermediate molecules.

1. De Novo Purine Nucleotide Synthesis:
   • The pathway for purine synthesis starts with the conversion of ribose-5-phosphate, derived from the pentose phosphate pathway or glycolysis, into phosphoribosyl pyrophosphate (PRPP).
   • PRPP serves as the common precursor for all purine nucleotides. It undergoes a series of enzymatic reactions, including the addition of atoms and functional groups, to form inosine monophosphate (IMP).
   • IMP can be further converted into adenosine monophosphate (AMP) or guanosine monophosphate (GMP) through additional enzymatic steps and feedback regulation.
2. De Novo Pyrimidine Nucleotide Synthesis:
   • The pyrimidine nucleotide synthesis pathway begins with the formation of carbamoyl phosphate, derived from glutamine, carbon dioxide, and ATP.
   • Carbamoyl phosphate combines with aspartate to form carbamoyl aspartate, which undergoes subsequent enzymatic reactions to produce orotate.
   • Orotate is then converted into uridine monophosphate (UMP) through a series of enzymatic steps, which can be further phosphorylated to form other pyrimidine nucleotides, such as cytidine monophosphate (CMP) and thymidine monophosphate (TMP).

Overall, de novo nucleotide synthesis is a complex and tightly regulated process that requires the coordinated action of multiple enzymes and metabolic pathways. It ensures the availability of nucleotides for DNA and RNA synthesis and contributes to cellular homeostasis and proper functioning.

Phosphoribosyl Pyrophosphate (PRPP)

• • sugar building block formed in nucleotide synthesis
    • added to nitrogenous base to form nucleoside
  • formed from ribose-5-phosphate
    • product of pentose phosphate shunt
  • ATP + ribose-5-phosphate = PRPP + AMP
    • catalyzed by PRPP synthetase
  • once PRPP is made it can add to either a de novo or salvaged base

Pyrimidine

• Pyrimidine
  • pathway diagram
  • important enzymes
    • carbamoyl phosphate synthetase-2
      • rate limiting step
      • not the same carbamoyl phosphate as in urea cycle
    • ribonucleotide reductase
      • inhibited by hydroxyurea 
      • also reduces UDP, CDP, ADP, GDP
        • dADP and dATP negatively feedback and inhibit enzyme
      • result = ↓ dTMP, dUDP, dCDP, dADP, dGDP
        • note: good to target thymidine synthesis because it is not involved in RNA
    • thymidylate synthase
      • inhibited by 5-fluorouracil (5-FU)
      • result = ↓ dTMP
    • dihydrofolate reductase
      • inhibited by methotrexate (MTX) in eukaryotes
      • inhibited by trimethoprim (TMP) in prokaryotes
        • sulfamethoxazole (SMX) interferes with DHF synthesis in prokaryotes
        • co-trimoxazole = TMP + SMX
      • inhibited by pyrimethamine in protozoa
      • result = ↓ dTMP

Deficiency

• • orotic aciduria
    • inability to convert orotic acid to UMP
    • defect in uridine monophosphate (UMP) synthase 
    • AR
    • presentation
      • ↑ orotic acid crystals in urine
      • megaloblastic anemia
        • does not improve with administration of vitamin B₁₂ or folic acid
        • not enough thymidine to sustain normal erythropoiesis
      • failure to thrive
      • no hyperammonemia
        • distinguishes between ornithine transcarbamylase (OTC) deficiency with high [orotic acid] in urine with hyperammonemia
    • treatment
      • oral uridine administration
        • bypasses defect in de novo pyrimidine pathway 

Purine

• purine
  • pathway diagram
  • insufficient capacity in most cells
  • important enzymes
    • PRPP amidotransferase
      • rate-limiting step
      • inhibited by AMP, GMP, IMP
      • indirectly inhibited by allopurinol
      • indirectly inhibited by 6-mercaptopurine
• note: base of inosine = hypoxanthine

Studies

1. Enzymology: Investigating the specific enzymes involved in nucleotide synthesis, their catalytic mechanisms, and their regulation.
2. Metabolic regulation: Understanding the complex regulatory mechanisms that coordinate nucleotide synthesis with other metabolic pathways and cellular processes.
3. Genetics: Studying genetic mutations and polymorphisms that affect nucleotide metabolism and contribute to disease.
4. Pharmacology: Developing drugs that target nucleotide metabolism, either by inhibiting nucleotide synthesis enzymes or by exploiting metabolic vulnerabilities in cancer cells.
5. Clinical applications: Investigating the role of nucleotide metabolism in disease, such as cancer, immunodeficiency disorders, and metabolic disorders.

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