Purine metabolism
synthase- no high energy compound involved
synthetase – uses high energy compound
Purine biosynthesis
- Ribose-5-P (from pentose phosphate pathway) PRPP via PRPP synthetase  5-P-betaribosylamine via PRPP amidotransferase
- PRPP used in synthesis of purine and pyrimidine nucleotides by providing ribose sugar and
alpha phosphate
- PRPP amidotransferase is rate determining and committed step of purine synthesis
- remaining reaction require ATP, amino acids (gly, glu, asp), and folic acid-derived cofactor
(N10- formyltetrahydrofolate, formyl THF)
- 6 high energy P bonds cleaved in production of inosinic acid (IMP)  first purine nucleotide
(base, sugar, P) formed
- AMP and GMP from IMP (separate pathways), figure 2 on 15-3
- GMP synthesis required redox reaction and nitrogen incorporation from amide side chain of
glutamine
- AMP acquires nitrogen from aspartate with production of fumarate; aspartate regenerated
from fumarate by formation, in citric acid cycle, of oxaloacetate, which is then aminated to
aspartate (fumarate malate oxaloacetate aspartate)
- mono-P products phosphorylated to di- and triphosphates by nucleoside monophosphate
kinase and nucleoside diphosphate kinase
- end result is production of purine ribonucleotides
- purine deoxyribionucleotides catalyzed by ribonucleotide reductase
1.
a.
Regulation of purine synthesis
- adenine and guanine nucleotides both inhibit the synthesis of IMP early in pathway at both
PRPP synthetase and PRPP amidotransferase; they also feedback inhibit their own product
from IMP
- excess ATP activates pathway from IMP  GMP, while excess GTP activates pathway from
IMP  AMP to maintain proper balance of purine nucleotides
b.
Purine degradation
- starts with removal of phosphate from purine nucleotide forms, yielding nucleoside (base,
sugar)
- adenosine  inosine catalyzed by adenosine deaminase
- sugars removed by purine nucleoside phosphorylase  purine bases guanine and
hypoxanthine, and ribose-1-P
- base products  xanthine  oxidized by xanthine oxidase  uric acid; xanthine oxidase
also converts hypoxanthine  xanthine; xanthine oxidase requires molecular oxygen and
molybdenum, non-heme iron, and FAD
Uric acid secretion
- uric acid is the purine degradative product  weak acid, with pK of 5.8; ionized form more
water-soluble than protonated form
- urine at pH 4.8  uric acid is 90% protonated  can dissolve 1/10 as much urate as urine at
pH 6.8 (90% ionized)
- normal urine pH below 5.8  overproduction of uric acid can lead to formation of stones (not
as soluble)
c.
Purine salvage pathway
- besides de novo synthesis, purine nucleotides can be formed directly from purine bases via
salvage pathway
- two enzymes: hypoxanthine-guanine phosphoribosyl transferase (HGPRT) and adenine
phosphoribosyl transferase
- HGPRT adds PRPP (from PRPP synthetase) to hypoxanthine  IMP or to guanine  GMP
- prevents irreversible destruction of hypoxanthine, guanine and adenine (purine bases reutilized)
- salvage pathway saves energy because of high energy demand of de novo synthesis pathway
- important for salvage of dietary nucleotides; salvage purine bases  uric acid kept low
(prevents gout)
- HGPRT defect  very low activity; Lesch-Nyhan syndrome
d. Disorders associated with defects of enzymes in purine metabolic pathways:
- elevation of uric acid from overproduction of purines or decreased excretion of uric acid
- hyperuricemia  associated with type I glycogen storage disease (von Gierke’s); glucose-6Pase defect  increased oxidation of glucose-6-P  ribose-5-P  elevates purine production
through saturation of PRPP synthetase
- hyperuricemia  gout  excessive accumulation of uric acid in body fluids; arthritis pain in
joints  urate crystals in cartilage around joint; kidney stones
Classification and causes of gout
- primary  inherited disorder
- secondary  induced by many disorders, leukemia
- linked to defects in metabolism of purines; PRPP synthetase superactive variant associated
with increased Vmax; another variant has an increased affinity (low Km) for ribose-5-P 
leading to overproduction of PRPP
- 3rd defect associated with loss of feedback inhibition of this enzyme by purine nucleotides;
when purine nucleotides reach excessive concentration  no signal for shutting off their further
production
- defects of HGPRT  inability to salvage purine bases from degradation  overproduction of
uric acid gout
Gout treatment
- allopurinol  competitive inhibitor of xanthine oxidase; hypoxanthine and xanthine excreted
during allopurinol therapy since xanthine oxidase uses both of these purines as substrates
- allopurinol (like purine bases) can be converted to ribonucleotide form by HGPRT; treatment
uses additional PRPP amidotransferase to reduce purine biosynthesis; analogue ribotide
produced may inhibit PRPP amidotransferase
- high [hypoxanthine] results from inhibition of xanthine oxidase  causes HGPRT to
reutilizes this base and further inhibit de no purine synthesis; purine synthesis lowered during
allopurinol treatment
- avoid animal products rich in nucleic acid (organ meats)
Lesch-Nyhan syndrome
- tremendous overproduction of uric acid; severe defect in HGPRT; males (X-linked)
- aggressive behavior, mental retardation, self-mutilation
- enzyme has essential role in non-hepatic tissue where de no synthesis of purines is slow 
non-hepatic tissues depend on circulating purine bases or nucleosides from liver
- non-hepatic tissues thought to take up circulating purines and through HGPRT form
nucleotides
2. PYRIMIDINE METABOLISM
Pyrimidine biosynthesis and its regulation
a. - glutamine (nitrogen donor)  carbamoyl phosphate through carbamoyl phosphate
synthetase II; urea cycle carbamoyl synthetase I uses ammonia
- genetic defect of ornithine transcarbamoylase, in urea cycle, causes accumulation of
carbamoyl phosphate that leaks from mitochondria to cytoplasm  increase pyrimidine
synthesis; increased pyrimidines excreted into urine (diagnose urea cycle defects)
- pyrimidine biosynthesis continues with addition of aspartate via aspartate
transcarbamoylase to provide remainder of ring elements  carbamoyl aspartate
- final product of pyrimidine synthesis is UMP  can be phosphorylated to UTP  cytidine
triphosphate (CTP)
- pyrimidines and ATP/GTP required for RNA synthesis
b.
- regulation of pyrimidine biosynthesis occurs at carbamoyl phosphate synthetase II 
feedback inhibition by uridine nucleotides (UDP, UTP)
- PRPP in excess (PRPP, ATP) enzyme stimulated
- to maintain balance of purine and pyrimidine nucleotides  high amounts of purine
nucleotides activate carbamoyl phosphate synthetase II
3. Metabolism of deoxyribionucleotides
Formation of deoxynucleotides
- 2-hydroxyl group of ribonucleotides reduced to form deoxyribonucleotide; catalyzed by
ribonucleotide reductase; requires thioredoxin as a reducing source (it is oxidized);
thioredoxin must be reduced back to its active form action of thioredoxin reductase (requires
NADPH)
- feedback regulation by deoxynucleotide triphosphates, dATP and dGTP
Formation of thymidine
- thymidine nucleotides required for DNA synthesis in place of UTP
- TTP derived from TMP, which is formed from dUMP
- dUMP  TMP catalyzed by thymidylate synthase; dUMP acquires a carbon from N5,N10methyleneTHF  converted to DHF
4. Metabolism of Folic Acid
- folic acid/folate undergoes activation and conversion to various forms used in several
biochemical reaction
- folate serves as a donor of one carbon group
- ingested folate  converted to dihydrofolate  reduced to tetrahydrofolate (THF) by
dihydrofolate reductase
- THF is backbone for production of other active forms of folate; must be in this form to carry
carbon
- N5,N10-methyleneTHF gains a carbon from side chain of serine yielding glycine product;
required for TMP synthesis from dUMP
- N5,N10-methenylTHF is a precursor for formation of N10-formyl THF  required in purine
biosynthesis (IMP formation); it is also a precursor of N5-methyl THF (in processing vitamin
B12, cobalamin)  provides methyl group for formation of methyl cobalamin; B12 deficiency
 folate trapped at N5-methyl THF because reaction from N5,N10-methyleneTHF is not
reversible
- folate supplementation prevents neural tube defects; prevent vascular disease
5. Chemotherapy and inhibition of DNA formation
- cancer cells require DNA synthesis, chemotherapeutic agents inhibit DNA synthesis at level of
synthesis of thymidine nucleotides because they are used selectively in DNA synthesis
Flurodeoxyuridylate (F-dUMP) – is the product of metabolism of 5-fluorouracil (form of drug
given) via orotate phosphoribosyl transferase; competitive inhibitor of thymidylate synthase
Methotrexate – inhibits dihydrofolate reductase resulting in decreased synthesis of THF (DHF
 THF via DHF reductase normally); inhibitor ultimately decreases production of active forms
of folate used in purine and pyrimidine synthesis (DNA synthesis disrupted)
- analogues of folate given during chemotherapy to supply demand of slowly dividing normal
cells