MSK TBL 2: Purine & Pyrimidine Metabolism

Nucleotides are composed of these 3 parts

1. pentose sugar

2. phosphate group

3. nitrogenous base

RNA & DNA pentose sugar differ by

presence of -OH [RNA] or -H [DNA] on 2’ carbon

Purines

adenine & guanine

2 heterocyclic rings

Pyrimidines

cytosine, thymine, uracil

1 heterocyclic ring

adenine

[NH2 at pos 6]

guanine

[NH2 at pos 2]

cytosine

[NH2 at pos 4]

uracil

[deaminated cytosine]

thymine

[methylated uracil]

hypoxanthine

[intmd in purine synth, no NH2]

xanthine

[intmd in purine synth, no NH2]

nucleoside

nitrogenous base + pentose sugar

ex. adenosine, guanosine, cytidine, thymidine, uridine

nucleotide

nitrogenous base + pentose sugar + phosphate

ex. adenose triphosphate [ATP]

purine de novo synthesis pathway requires

aspartate, glutamine, CO2, glycine, & THF

• heterocyclic rings of hypoxanthine

  • derive their nitrogen groups from

    • glutamine, glycine, & aspartate

  • derive their carbon groups from

    • CO2, glycine, & THF

pyrimidine de novo synthesis pathway requires

aspartate, glutamine, O2

IMP =

hypoxanthine + PRPP

*a purine nucleotide but not incorporated into DNA or RNA, so intmd in synth of purines

purine de novo synthesis pathway

1. convert ribose 5-phosphate [from PPP/HMP shunt of glycolysis] to PRPP molecule [ 5-phosphoribosyl-1-pyrophosphate]

2. convert PRPP → PRA [phosphoribosylamine]

3. PRA → IMP [inosine monophosphate]

4. conversion of IMP → adenosine & guanosine nucleotides

Purine salvage pathway

salvages nitrogenous bases back into purine nucleotides rather than breaking them down as uric acid [inosine → hypoxanthine → xanthine → uric acid]

salvaged by addition of nitrogenous bases to PRPP

HGPRT [hypoxanthine-guanine phosphoribosyl transferase]

enzyme that reincorporates

• Hypoxanthine + PRPP → IMP

• Guanine + PRPP → GMP

APRT [adenine phosphoribosyl transferase]

reincorporates

• Adenine + PRPP → AMP

AMP & GMP ____ cross-activate each other

positively

AKA more adenine production drives guanine production, and vice versa

Lesch-Nyhan Syndrome

• X-linked recessive disorder

• deficiency of essential salvage enzyme HGPRT

• gout attacks from high uric acid levels [b/c all purines converted to uric acid w/o enzyme]

• neurological impairments, weak muscle tone, & self-mutilating behavior

Severe Combined Immunodeficiency [SCID]

• genetic disorder of immune system

• autosomal recessive adenosine deaminase [ADA] deficiency

• insufficient deoxynucleotide triphosphates [dNTP] → dysfunction of B & T lymphocytes, which normally undergo rapid proliferation

• ADA involved in pathway converting common purine intmd → hypoxanthine & xanthine

  • deficiency → high levels of adenosine → build-up of deoxyadenosine triphosphate [dATP]

  • → dATP acts as inhibitor of ribonucleotide reductase → deficiency in dNTP synthesis

• Dysfunctional B lymphocytes → susceptible to pathogens like Pneumocystis jirovecii

• Dysfunctional T lymphocytes → susceptible to infections like Candida albicans

Cladribine

• drug used in treatment of hairy cell leukemia

• purine analog that incorporates into DNA → strand breakdown

• when drug phosphorylated/activated inside cancer cell, it resists deamination by ADA & accumulates in cancerous cells

Allopurinol, Febuxostat

• drug used in treatment of chronic gout [NOT acute gout]

• blocks xanthine oxidase [XO] → lowers serum uric acid

*also used to treat Lesch-Nyhan Syndrome

xanthine oxidase [XO]

converts hypoxanthine → xanthine → uric acid

pyrimidine de novo synthesis pathway

begins with amino acid glutamine

1. create carbamoyl phosphate synthetase II [CPS II] in cell cytoplasm

2. conversion of carbamoyl phosphate → orotic acid via addition of aspartate

3. orotic acid added to PRPP to form uridine monophosphate [UMP] via enzyme uridine monophosphate synthase [UMP synthase]

4. uridine diphosphate [UDP] convert to deoxyuridine diphosphate [dUDP] by enzyme ribonucleotide reductase

5. methylene-tetrahydrofolate [M-THF] donates 1 C [methyl group] to deoxyuridine monophosphate [dUMP] to form deoxythymidine monophosphate [dTMP]

   1. rxn mediated by thymidylate synthase

   2. from this rxn, M-THF → dihydrofolate [DHF]

CPS II

• rate-limiting enzyme in pyrimidine synthesis

• inhibited via feedback inhibition from high levels of pyrimidine uridine triphosphate

Uridine monophosphate [UMP]

serves as common precursor to both cytidine monophosphate [CMP] & thymidine monophosphate [TMP] synthesis

*much like IMP in purine synth

ribonucleotide reductase

• imp for building nucleotides for DNA replication & repair

• tightly controlled b/c dNTP levels must be balanced

  • unbalanced levels → incorporation of incorrect nucleotides into a cell’s DNA → DNA damage & cell death

  • control achieved by regulating transcription of enzyme & allosteric regulation

    • ATP is allosteric activator

    • deoxyadenosine triphosphate [dATP] is allosteric inhibitor

dihydrofolate reductase

converts DHF to THF

• from there, THF readily converted to M-THF, allowing body to continue metabolizing dUMP into thymidine monophosphate → incorporates into DNA

*Uridine monophosphate can only incorporate into RNA

pyrimidine salvage pathway

• allows for use of recovered uracil & thymidine to generate uridine & deoxythymidine & their corresponding nucleotides

• 2 types of enzymes

  • Phosphorylases

  • Kinases

Phosphorylases in pyrimidine salvage pathway

• attach ribose-1-phosphate [or deoxyribose-1-phosphate] to salvaged bases of uracil & thymine to generate nucleosides uridine & deoxythymidine

• inorganic phosphate [Pi] is displaced

Kinases in pyrimidine salvage pathway

• uridine kinase catalyzes phosphorylation of uridine → UMP, which can be converted to UTP

• thymidine kinase phosphorylates deoxythymidine → dTMP, which can be converted to dTDP →dTTP

UMP Synthase Deficiency

• defect in enzyme → orotic aciduria [autosomal recessive]

  • orotic acid builds up & is excreted in urine

  • pt no longer able to synth pyrimidines

• pts present w/ B12- & folate- resistant megaloblastic anemia

• treatment

  • administer uridine

Ornithine Transcarbamylase [OTC] Deficiency

• deficiency in urea cycle enzyme

  • carbamoyl phosphate builds up → high levels of orotic acid

  • high ammonia in blood

  • encephalopathy

Hydroxyurea

• antineoplastic [anticancer]

• inhibits ribonucleotide reductase → inhibits DNA replication

• beneficial in sickle cell disease

  • Hydroxyurea → increases NO levels → binds & activates cGMP signaling → increases fetal Hb levels

6-Mercaptopurine [6-MP], Azathioprine [prodrug of 6MP]

• decreases nucleotide synthesis → decreased proliferation of lymphocytes

• at high dosage, inhibits cell proliferation [cytostatic effect]

• at low dosage, immunosuppressive effects

Mycophenolate

• reversible inhibition of inosine monophosphate [IMP] dehydrogenase → blocks purine synthesis

Ribavarin

• antiviral

• inhibition of inosine monophosphate [IMP] dehydrogenase → blocks guanine nucleosides synthesis

Methotrexate

• anticancer [antineoplastic] & immune suppressant for autoimmune diseases

• competitive inhibition of dihydrofolate reductase via displacement of dihydrofolate → decreases THF → decreases dTMP & purine nucleotides

Trimethoprim

• antibiotic

• competitive inhibition of dihydrofolate reductase via displacement of dihydrofolate → decreases THF → decreases dTMP & purine nucleotides

Pyrimethamine

• anthelminthic [antiparasitic]

• competitive inhibition of dihydrofolate reductase via displacement of dihydrofolate → decreases THF → decreases dTMP & purine nucleotides

Leflunomide

• disease-modifying antirheumatic drug [DMARD]

• reversibly inhibits dihydrofolate dehydrogenase [an enzyme that produces orotic acid in the pyrimidine synth pathway]

Capecitabine

5FU pro-drug

5-fluorouracil [5-FU]

1. pyrimidine analog that incorporates into DNA & RNA to decrease DNA & RNA synth

2. complex formation w/ thymidylate synthase & folic acid → inhibition of thymidylate synthase → decreased dTMP prod. → decreased DNA synthesis