Nucleotide Metabolism

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Nucleotide Metabolism
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Bases/Nucleosides/Nucleotides
Base Base
Base Sugar Phosphate Nucleotide
Base Sugar Nucleoside
Deoxyadenosine 5-triphosphate (dATP)
Adenine
Deoxyadenosine
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Cellular Roles of Nucleotides

• Energy metabolism (ATP)
• Monomeric units of nucleic acids
• Regulation of physiological processes
• Adenosine controls coronary blood flow
• cAMP and cGMP serve as signaling molecules
• Precursor function-GTP to tetrahydrobiopternin
• Coenzyme components- 5-AMP in FAD/NAD
• Activated intermediates- UDP Glucose
• Allosteric effectors- regulate themselves and
  others

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How I hope to make this at least bearable if not
mildly interesting

• Purines and Pyrimidines
• Synthesis (de novo and salvage pathways)
• Degradation
• Relevant disease states
• Relevant clinical applications (Friday)

You are not responsible for any structures
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Purines and Pyrimidines
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Synthesis Pathways

• For both purines and pyrimidines there are two
  means of synthesis (often regulate one another)
• de novo (from bits and parts)
• salvage (recycle from pre-existing nucleotides)

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Many Steps Require an Activated Ribose Sugar
(PRPP)
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de novo Synthesis

• Committed step This is the point of no return
• Occurs early in the biosynthetic pathway
• Often regulated by final product (feedback
  inhibition)

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Purine Biosynthesis (de novo)

• Atoms derived from
• Aspartic acid
• Glycine
• Glutamine
• CO2
• Tetrahydrofolate
• Also requires
• 4 ATPs

Committed Step
Purines are synthesized on the Ribose ring
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Purine Biosynthesis (de novo)
(A bunch of steps you dont need to know)
IMP
(Inosine Monophosphate)
ATP
GTP
AMP
GMP
Feedback Inhibition
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Purine Degradation
Other species further metabolize uric acid

• Sequential removal of bits and pieces
• End product is uric acid
• Uric acid is primate-specific

Excreted in Urine
Xanthine Oxidase
Xanthine
Uric Acid
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Excess Uric Acid Causes Gout

• Primary gout (hyperuricemia)
• Inborn errors of metabolism that lead to
  overproduction of Uric Acid
• Overactive de novo synthesis pathway
• Leads to deposits of Uric Acid in the joints
• Causes acute arthritic joint inflammation

Xanthine Oxidase
Xanthine
Uric Acid
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Immunodeficiency Diseases Associated with Purine
Degradation

• Defect in adenosine deaminase
• Removes amine from adenosine
• SCID- severe combined immunodeficiency
• Bubble Boy Disease
• Defect in both B-cells and T-cells (Disease of
  Lymphocytes)
• Patients extremely susceptible to infection -
  hence the Bubble

Lymphocyte
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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

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Salvage Pathway for Purines
Hypoxanthine or Guanine

PRPP
IMP or GMP PPi
Hypoxanthineguanosylphosphoribosyl
transferase (HGPRTase)
Adenine

PRPP
AMP PPi
Adeninephosphoribosyl transferase (APRTase)
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Lesch-Nyhan Syndrome

• Absence of HGPRTase
• X-linked (Gene on X)
• Occurs primarily in males
• Characterized by
• Increased uric acid
• Spasticity
• Neurological defects
• Aggressive behavior
• Self-mutilation

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Total Aside on X-linked Diseases

• Why are X-linked diseases generally found only in
  males?
• Females have two X chromosomes - would need to
  mutate both copies to see a recessive phenotype
• Males have a single X chromosome

XY
XX
Think about Fragile X Syndrome
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Biosynthesis of Pyrimidines

• Synthesized from
• Glutamine
• CO2
• Aspartic acid
• Requires ATP

Uracil
Cytosine

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

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Regulation of Pyrimidine Biosynthesis

• Regulation occurs at first step in the pathway
  (committed step)
• 2ATP CO2 Glutamine carbamoyl phosphate

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Hereditary Orotic Aciduria

• Defect in de novo synthesis of pyrimidines
• Loss of functional UMP synthetase
• Gene located on chromosome III
• Characterized by excretion of orotic acid
• Results in severe anemia and growth retardation
• Extremely rare (15 cases worldwide)
• Treated by feeding UMP

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Why does UMP Cure Orotic Aciduria?
UMP
Carbamoyl Phosphate
Orotate
Feedback Inhibition

• Disease (-UMP)
• No UMP/excess orotate
• Disease (UMP)
• Restore depleted UMP
• Downregulate pathway via feedback inhibition
  (Less orotate)

UTP
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Biosynthesis Purine vs Pyrimidine
Purine
Pyrimidine

• Synthesized on PRPP
• Regulated by GTP/ATP
• Generates IMP
• Requires Energy

• Synthesized then added to PRPP
• Regulated by UTP
• Generates UMP/CMP
• Requires Energy

Both are very complicated multi-step process
which your kindly professor does not expect you
to know in detail
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Pyrimidine Degradation/Salvage

• Pyrimindine rings can be fully degraded to
  soluble structures (Compare to purines that make
  uric acid)
• Can also be salvaged by reactions with PRPP
• Catalyzed by Pyrimidine phosphoribosyltransferase

Degradation pathways are quite distinct for
purines and pyrimidines, but salvage pathways
are quite similar
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Wait a minuteSo far weve only made GMP, AMP,
and UMP
So how the heck are we supposed to make DNA?
We need the dNTPs according to the Know-it-All
Professor who taught us that a couple of months
ago
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Two Problems

• These are monophosphates (i.e. GMP)- we need
  triphosphates (i.e. GTP) for both DNA and RNA
  synthesis
• These are ribonucleotides- thats fine for RNA
  but we also need to make DNA

Synthesis of ribonucleotides first supports the
RNA world theory
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Specific Kinases Convert NMP to NDP
Nucleoside Monophosphates
Nucleoside Diphosphates
Monophosphate Kinases
NMP
NDP

• Monophosphate kinases are specific for the bases

Adenylate Kinase
Guanylate Kinase
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Conversion of Ribonucleotides to
Deoxyribonucleotides
BASE
BASE
Ribonucleotide Reductase
Deoxyribonucleoside
Ribonucleoside
Somehow we need to get rid of this oxygen
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Ribonucleotide Reductase

• Catalyzes conversion of NDP to dNDP
• Highly regulated enzyme
• Regulates the level of cellular dNTPs
• Activated prior to DNA synthesis
• Controlled by feedback inhibition

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dNDP to dNTP (the final step)

• Once dNDPs are generated by ribonucleotide
  reductase a general kinase can phosphorylate to
  make the dNTPs
• So far weve made GTP, ATP, and UTP (which can be
  aminated to form CTP)
• What about TTP?

Youll have to tune in tomorrow
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Plan for Tomorrow

• Brief Explanation of how dUMP is converted to
  dTMP
• Some clinically relevant treatments based on
  these pathways that are used to combat
• Cancer
• Bacterial Infections
• Viral Infections

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Take Home Concepts from Todays Lecture

• Nucleotides can be made through two pathways
• (de novo and salvage)
• Pathways are regulated by feedback inhibition
• Specific degradation pathways exist
• Molecular basis of metabolic diseases mentioned
• What steps are necessary to generate a dNTP from
  the initial NMP made