Purine and Pyrimidine Metabolism
nucleoside triphosphates
(ATP and GTP) provide energy for reactions that would otherwise be extremely unfavorable in the cell
Gout
Acute gouty arthritis, seen most commonly in males, results from precipitation of monosodium urate crystals in joints. The crystals, identified as negatively birefringent and needle-shaped, initiate neutrophil-mediated and acute inflammation, often first affecting the big toe. Chronic gout may manifest over time as tophi (deposits of monosodium urate) in soft tissue around joints, leading to chronic inflammation involving granulomas. Acute attacks of gout are treated with colchicine or indomethacin to reduce the inflammation. Chronic hyperuricemia, because of underexcretion, is treated with a uricosuric drug (probenecid). Overproduction of uric acid and chronic gout are treated with allopurinol. Treatment of large tumors with chemotherapeutic regimens or radiation may cause "turnor lysis syndrome" and excessive excretion of uric acid, resulting in gout. The cause of the excessive uric acid is the destruction of the cancer cell's nucleic acid into purines undergoing turnover.
Adenosine deaminase (ADA) deficiency
Adenosine deaminase (ADA) deficiency, an autosomal recessive disorder, causes a type of severe combined immunodeficiency (SCID). Lacking both B- cell and T-cell function, children are multiply infected with many organisms (Pneumocystis carinii, Candida) and do not survive without treatment. Enzyme replacement therapy and bone marrow transplantation may be used. Experimental gene therapy trials have not yet yielded completely successful cures. High levels of dATP accumulate in red cells of ADA patients and inhibit ribonucleotide reductase, thereby inhibiting the production of other essential deoxynucleotide precursors for DNA synthesis (see Figure I-18-3). Although it is believed that the impaired DNA synthesis contributes to dysfunction of T cells and B cells, it is not known why the main effects are limited to these cell types.
Nucleotides
are needed for DNA and RNA synthesis (DNA replication and transcription) and for energy transfer
A 6-month-old boy becomes progressively lethargic and pale and shows delayed motor development. Laboratory evaluation reveals normal blood urea nitrogen (BUN), low serum iron, hemoglobin 4.6 g/dL, and leukopenia. His bone marrow shows marked megaloblastosis, which did not respond to treatment with iron, folic acid, vitamin B12, or pyridoxine. His urine developed abundant white precipitate identified as orotic acid. The underlying defect causing the megaloblastic anemia in this child is most likely in which of the following pathways? A. Homocysteine metabolism B. Pyrimidine synthesis C. Urea synthesis D. Uric acid synthesis E. Heme synthesis
Answer: B. Accumulation of orotic acid indicates megaloblastic anemia arises because pyrimidines are required for DNA synthesis.
The anticancer drug 6-mercaptopurine is deactivated by the enzyme xanthine oxidase. A cancer patient being treated with 6-mercaptopurine develops hyperuricemia, and the physician decides to give the patient allopurinol. Resistance of neoplastic cells to the chemotherapeutic effect of 6-mercaptopurine would most likely involve loss or inactivation of a gene encoding A. thymidylate synthase B. hypoxanthine phosphoribosyltransferase C. purine nucleoside pyrophosphorylase D. orotic acid phosphoribosyltransferase E. adenosine deaminase
Answer: B. HPRT is required for activation of 6 mercaptopurine to its ribonucleotide and inhibition of purine synthesis. The other enzymes listed are not targets for this drug.
The anticancer drug 6-mercaptopurine is deactivated by the enzyme xanthine oxidase. A cancer patient being treated with 6-mercaptopurine develops hyperuricemia, and the physician decides to give the patient allopurinol. What effect will allopurinol have on the activity of 6-mercaptopurine? A. Enhanced deactivation of 6-mercaptopurine B. Enhanced elimination of 6-mercaptopurine as uric acid C. Enhanced retention and potentiation of activity D. Decreased inhibition of PRPP glutamylamidotransferase
Answer: C. Because allopurinol inhibits xanthine oxidase, the 6-mercaptopurine will not be deactivated as rapidly
A 12-week-old infant with a history of persistent diarrhea and candidiasis is seen for a respiratory tract infection with Pneumocystis jiroveci. A chest x-ray confirms pneumonia and reveals absence of a thymic shadow. Trace IgG is present in his serum, but IgA and IgM are absent. His red blood cells completely lack an essential enzyme in purine degradation. The product normally formed by this enzyme is A. guanine monophosphate B. hypoxanthine C. inosine D. xanthine E. xanthine monophosphate
Answer: C. The child most likely has severe combined immunodeficiency caused by adenosine deaminase deficiency. This enzyme deaminates adenosine (a nucleoside) to form inosine (another nucleoside). Hypoxanthine and xanthine are both purine bases, and the monophosphates are nucleotides.
Patients with Lesch-Nyhan syndrome have hyperuricemia, indicating an increased biosynthesis of purine nucleotides, and markedly decreased levels of hypoxanthine phosphoribosyl transferase (HPRT). The hyperuricemia can be explained on the basis of a decrease in which regulator of purine biosynthesis? A. ATP B. GDP C. Glutamine D. IMP E. PRPP
Answer: D. IMP is a feedback inhibitor of PRPP amidophosphoribosyl transferase, the first reaction in the biosynthesis of purines. IMP is formed by the HPRT reaction in the salvage of hypoxanthine.
How do cells synthesize nucleotides?
Cells synthesize nucleotides in 2 ways: de novo synthesis and salvage pathways In many cells, the capacity for de novo synthesis to supply purines and pyrimidines is insufficient, and the salvage pathway is essential for adequate nucleotide synthesis.
dihydrofolate reductase (DHFR)
Converts DHF to THF; Without DHFR, thymidylate synthesis will eventually stop
Purine catabolism and the salvage enzyme HGPRT
Excess purine nucleotides or those released from DNA and RNA by nucleases are catabolized first to nucleosides (loss of Pi) and then to free purine bases ( release of ribose or deoxyribose). Excess nucleoside monophosphates may accumulate when: • RNA is normally digested by nucleases (mRNAs and other types of RNAs are continuously turned over in normal cells). • Dying cells release DNA and RNA, which is digested by nucleases. • The concentration of free Pi decreases as it may in galactosemia, hereditary fructose intolerance, and glucose-6-phosphatase deficiency. Salvage enzymes recycle normally about 90% of these purines, and 10% are converted to uric acid and excreted in urine. When purine catabolism is increased significantly, a person is at risk for developing hyperuricemia and potentially gout.
Thiazide diuretics
Hydrochlorothiazide (HCTZ) Thiazide diuretics (hydrochlorothiazide and chlorthalidone) may cause hyperuricemia.
Hyperuricemia (gout)
Hyperuricemia may be produced by overproduction of uric acid or underexcretion of uric acid by the kidneys. Hyperuricemia may progress to acute and chronic gouty arthritis if uric acid (monosodium urate) is deposited in joints and surrounding soft tissue, where it causes inflammation. Uric acid is produced from excess endogenous purines and is also produced from dietary purines (digestion of nucleic acid in the intestine) by intestinal epithelia. Both sources of uric acid are transported in the blood to the kidneys for excretion in urine. Allopurinol inhibits xanthine oxidase and also can reduce purine synthesis by inhibiting PRPP amidotransferase, provided HGPRT is active Hyperuricemia and gout often accompany the following conditions: Lesch-Nyhan syndrome (no purine salvage) Partial deficiency of HGPRT Alcoholism (lactate and urate compete for same transport system in the kidney) Glucose 6-phosphatase deficiency Hereditary fructose intolerance (aldolase B deficiency) Galactose 1-phosphate uridyl transferase deficiency (galactosemia) Mutations in PRPP synthetase that lower Km
Lesch-Nyhan syndrome
Lesch-Nyhan syndrome is an X-linked recessive condition involving: Near-complete deficiency of HGPRT activity Intellectual disability Spastic cerebral palsy with compulsive biting of hands and lips Hyperuricemia Death often in first decade Over 100 distinct mutations of the HGPRT gene located on the X chromosome have been reported to give rise to Lesch-Nyhan syndrome. These mutations include complete deletions of the gene, point mutations that result in an increased Km for hypoxanthine and guanine for the enzyme, and mutations that cause the encoded enzyme to have a short half-life.
The parents of a 9-month-old infant were concerned that their son appeared generally weak, had difficulty moving his arms and legs, repeatedly bit his lips, and frequently seemed to be in pain. The infant was brought to the pediatrician. The parents mentioned that since the baby was born, they often noticed tiny, orange-colored particles when they changed the infant's diapers. Lab analysis of uric acid in urine was normalized to the urinary creatinine in the infant, and it was found that the amount was 3 times greater than the normal range.
Lesch-Nyhan syndrome: One of the earliest signs of Lesch-Nyhan syndrome is the appearance of orange crystals in diapers. They are needle shaped sodium urate crystals. Without the salvaging of hypoxanthine and guanine by HGPRT, the purines are shunted toward the excretion pathway. This is compounded by the lack of regulatory control of the PRPP amidotransferase in the purine synthesis pathway, resulting in the synthesis of even more purines in the body. The large amounts of urate will cause crippling, gouty arthritis and urate nephropathy. Renal failure is usually the cause of death. Treatment with allopurinol will ease the amount of urate deposits formed.
Thymidylase synthase
Methylates dUMP to dTMP; Requires THF
Orotic aciduria
Orotic aciduria is an autosomal recessive disorder caused by a defect in uridine monophosphate (UMP) synthase. This enzyme contains two activities, orotate phosphoribosyltransferase and orotidine decarboxylase. The lack of pyrimidines impairs nucleic acid synthesis needed for hematopoiesis, explaining the megaloblastic anemia in this infant. Orotic acid accumulates and spills into the urine, resulting in orotic acid crystals and orotic acid urinary obstruction. The presence of orotic acid in urine might suggest that the defect could be ornithine transcarbamylase (OTC) deficiency, but the lack of hyperammonemia rules out a defect in the urea cycle. Uridine administration relieves the symptoms by bypassing the defect in the pyrimidine pathway. Uridine is salvaged to UMP, which feedback-inhibits carbamoyl phosphate synthase-2, preventing orotic acid formation.
Purine synthesis
Purines are synthesized de novo beginning with PRPP. The most important enzyme is PRPP amidotransferase, which catalyzes the first and rate-limiting reaction of the pathway. It is inhibited by the 3 purine nucleotide end products AMP, GMP, and IMP. The drugs allopurinol (used for gout) and 6-mercaptopurine (antineoplastic) also inhibit PRPP amidotransferase. These drugs are purine analogs which must be converted to their respective nucleotides by HGPRT within cells. The amino acids glycine, aspartate, and glutamine are used in purine synthesis. Tetrahydrofolate is required for synthesis of all the purines. Inosine monophosphate (contains the purine base hypoxanthine) is the precursor for AMP and GMP
Pyrimidine synthesis
Pyrimidines are synthesized de novo in the cytoplasm from aspartate, CO2, and glutamine. Synthesis involves a cytoplasmic carbamoyl phosphate synthetase that differs from the mitochondrial enzyme with the same name used in the urea cycle. The primary end product of pyrimidine synthesis is UMP. In the conversion of UMP to dTMP, 3 important enzymes are ribonucleotide reductase, thymidylate synthase, and dihydrofolate reductase; all are targets of antineoplastic drugs.
Pyrimidine catabolism
Pyrimidines may be completely catabolized (NH4+ is produced) or recycled by pyrimidine salvage enzymes.
Excess nucleoside monophosphates may accumulate when:
RNA is normally digested by nucleases (mRNAs and other types of RNAs are continuously turned over in normal cells). Dying cells release DNA and RNA, which is digested by nucleases. The concentration of free Pi decreases as it may in galactosemia, hereditary fructose intolerance, and glucose-6-phosphatase deficiency.
Important enzymes of Pyrimidine synthesis
Ribonucleotide reductase Thymidylate synthase Dihydrofolate reductase (DHFR)
Ribonucleotide reductase
Ribonucleotide reductase is required for the formation of the deoxyribonucleotides for DNA synthesis. All 4 nucleotide substrates must be diphosphates. dADP and dATP strongly inhibit ribonucleotide reductase. Hydroxyurea, an anticancer drug, blocks DNA synthesis indirectly by inhibiting ribonucleotide reductase.
Types of orotic acidurias
Two Orotic Acidurias: Hyperammonemia (no megaloblastic anemia) -Pathway: urea cycle -Enzyme deficient: OTC Megaloblastic anemia (no hyperammonemia) - Pathway: pyrimidine synthesis - Enzyme deficient: UMP synthase
Lesch-Nyhan disease
an enzyme for purine salvage (hypoxanthine guanine phosphoribosyl pyrophosphate transferase, HPRT) is absent or deficient. People with this genetic deficiency have CNS deterioration, intellectual disability, and spastic cerebral palsy associated with compulsive self-mutilation. Cells in the basal ganglia of the brain (fine motor control) normally have very high HPRT activity. Patients also all have hyperuricemia because purines cannot be salvaged, causing gout.
ribose-5-phosphate
for nucleotide synthesis is derived from the HMP shunt and is activated by the addition of pyrophosphate from ATP, forming phosphoribosyl pyrophosphate (PRPP). using PRPP synthetase.
De novo synthesis of nucleotides
occurs predominantly in the liver, purines and pyrimidines are synthesized from smaller precursors, and PRPP is added to the pathway at some point
Salvage pathways
preformed purine and pyrimidine bases can be converted into nucleotides by salvage enzymes distinct from those of de novo synthesis. Purine and pyrimidine bases for salvage enzymes may arise from: -- Synthesis in the liver and transport to other tissues -- Digestion of endogenous nucleic acids (cell death, RNA turnover)