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BIOMEDICAL IMPORTANCE
In addition to serving as precursors of nucleic acids, purine and pyrimidine nucleotides participate in metabolic functions as diverse as energy
metabolism, protein synthesis, regulation of enzyme activity, and signal transduction. When linked to vitamins or vitamin derivatives,
nucleotides form a portion of many coenzymes. As the principal donors and acceptors of phosphoryl groups in metabolism, nucleoside tri- and
diphosphates such as ATP and ADP are the principal players in the energy transductions that accompany metabolic interconversions and
oxidative phosphorylation. Linked to sugars or lipids, nucleosides constitute key biosynthetic intermediates. The sugar derivatives UDP-glucose
and UDP-galactose participate in sugar interconversions and in the biosynthesis of starch and glycogen. Similarly, nucleoside-lipid derivatives
such as CDP-acylglycerol are intermediates in lipid biosynthesis. Roles that nucleotides perform in metabolic regulation include ATP-dependent
phosphorylation of key metabolic enzymes, allosteric regulation of enzymes by ATP, AMP, and CTP, and control by ADP of the rate of oxidative
phosphorylation. The cyclic nucleotides cAMP and cGMP serve as the second messengers in hormonally regulated events, and GTP and GDP play
key roles in the cascade of events that characterize signal transduction pathways. Specifically medical applications include the use of synthetic
purine and pyrimidine analogs that contain halogens, thiols, or additional nitrogen atoms in the chemotherapy of cancer and AIDS, and as
suppressors of the immune response during organ transplantation.
CHEMISTRY OF PURINES, PYRIMIDINES, NUCLEOSIDES, & NUCLEOTIDES
Purines & Pyrimidines Are Heterocyclic Compounds
Purines and pyrimidines are nitrogen-containing heterocycles, structures that contain, in addition to carbon, other (hetero) atoms such as
nitrogen. Note that the smaller pyrimidine molecule has the longer name and the larger purine molecule the shorter name, and that their
six-atom rings are numbered in opposite directions (Figure 32–1). Purines or pyrimidines with an –NH2 group are weak bases (pKb values
10–11). The planar character of purines and pyrimidines facilitates their close association, or "stacking," that stabilizes double-stranded DNA (see
Chapter 34). The oxo and amino groups of purines and pyrimidines exhibit keto-enol and amine-imine tautomerism (Figure 32–2), although
physiologic conditions strongly favor the amino and oxo forms.
Figure 32–1.
Purine and pyrimidine. The atoms are numbered according to the international system.
Figure 32–2.
Tautomerism of the oxo and amino functional groups of purines and pyrimidines.
Nucleosides Are N-Glycosides
Nucleosides are derivatives of purines and pyrimidines that have a sugar linked to a ring nitrogen of a purine or pyrimidine. Numerals with a
prime (eg, 2' or 3') distinguish atoms of the sugar from those of the heterocycle. The sugar in ribonucleosides is D-ribose, and in
deoxyribonucleosides is 2-deoxy-D-ribose. Both sugars are linked to the heterocycle by a
-N-glycosidic bond, almost always to the N-1 of
a pyrimidine or to N-9 of a purine (Figure 32–3).
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Figure 32–3.
Ribonucleosides, drawn as the syn conformers.
Nucleotides Are Phosphorylated Nucleosides
Mononucleotides are nucleosides with a phosphoryl group esterified to a hydroxyl group of the sugar. The 3'- and 5'-nucleotides are nucleosides
with a phosphoryl group on the 3'- or 5'-hydroxyl group of the sugar, respectively. Since most nucleotides are 5'-, the prefix "5'-" usually is
omitted when naming them. UMP and dAMP thus represent nucleotides with a phosphoryl group on C-5 of the pentose. Additional phosphoryl
groups, ligated by acid anhydride bonds to the phosphoryl group of a mononucleotide, form nucleoside diphosphates and triphosphates
(Figure 32–4).
Figure 32–4.
ATP, its diphosphate, and its monophosphate.
Heterocylic N-Glycosides Exist as Syn and Anti Conformers
Steric hindrance by the heterocycle dictates that there is no freedom of rotation about the
-N-glycosidic bond of nucleosides or nucleotides.
Both therefore exist as noninterconvertible syn or anti conformers (Figure 32–5). Unlike tautomers, syn and anticonformers can only be
interconverted by cleavage and reformation of the glycosidic bond. Both syn and anti conformers occur in nature, but the anti conformers
predominate.
Figure 32–5.
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The syn and anti conformers of adenosine differ with respect to orientation about the N-glycosidic
Table 32–1 lists the major purines and pyrimidines and their nucleoside and nucleotide derivatives. Single-letter abbreviations are used to
identify adenine (A), guanine (G), cytosine (C), thymine (T), and uracil (U), whether free or present in nucleosides or nucleotides. The prefix "d"
(deoxy) indicates that the sugar is 2'-deoxy-D-ribose (for example, in dATP) (Figure 32–6).
Table 32–1. Purine Bases, Ribonucleosides, and Ribonucleotides
Purine or Pyrimidine X = H
Adenine
X = Ribose X = Ribose Phosphate
Adenosine
Adenosine monophosphate (AMP)
Guanine Guanosine
Guanosine monophosphate (GMP)
Cytosine Cytidine
Cytidine monophosphate (CMP)
Uracil
Uridine monophosphate (UMP)
Uridine
Thymine Thymidine
Thymidine monophosphate (TMP)
Figure 32–6.
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Structures of AMP, dAMP, UMP, and TMP.
Modification of Polynucleotides Can Generate Additional Structures
Small quantities of additional purines and pyrimidines occur in DNA and RNAs. Examples include 5-methylcytosine of bacterial and human DNA,
5-hydroxymethylcytosine of bacterial and viral nucleic acids, and mono- and the di-N-methylated adenine and guanine of mammalian
messenger RNAs (Figure 32–7) that function in oligonucleotide recognition and in regulating the half-lives of RNAs. Free nucleotides include
hypoxanthine, xanthine, and uric acid (Figure 32–8) that are intermediates in the catabolism of adenine and guanine (see Chapter 33).
Methylated heterocycles of plants include the xanthine derivatives caffeine of coffee, theophylline of tea, and theobromine of cocoa (Figure
32–9).
Figure 32–7.
Four uncommon but naturally occurring pyrimidines and purines.
Figure 32–8.
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Structures of hypoxanthine, xanthine, and uric acid, drawn as the oxo tautomers.
Figure 32–9.
Caffeine, a trimethylxanthine. The dimethylxanthines theobromine and theophylline are similar but lack the methyl group at N-1 and at N-7,
respectively.
Nucleotides Are Polyfunctional Acids
The phosphoryl groups of nucleosides have pKa values of about 1.0. Nucleotides therefore bear significant negative charge at physiologic pH. By
contrast, the pKa values of the secondary phosphoryl groups are about 6.2, so these can serve as proton donors or acceptors at pH values
approximately two or more units above or below neutrality.
Nucleotides Absorb Ultraviolet Light
The conjugated double bonds of purine and pyrimidine derivatives absorb ultraviolet light. The mutagenic effect of ultraviolet light is due to its
absorption by nucleotides in DNA that results in chemical modifications (see Chapter 35). While spectra are pH-dependent, at pH 7.0 all the
common nucleotides absorb light at a wavelength close to 260 nm. The concentration of nucleotides and nucleic acids thus often is expressed in
terms of "absorbance at 260 nm."
Nucleotides Serve Diverse Physiologic Functions
In addition to their roles as precursors of nucleic acids, ATP, GTP, UTP, CTP and their derivatives each serve unique physiologic functions
discussed in other chapters. selected examples include the role of ATP as the principal biologic transducer of free energy; and the second
messenger cAMP (Figure 32–10). The mean intracellular concentration of ATP, the most abundant free nucleotide in mammalian cells, is about 1
mmol/L. Since little cAMP is required, the intracellular cAMP concentration (about 1 nmol/L) is three orders of magnitude below that of ATP.
Other examples include adenosine 3'-phosphate-5'-phosphosulfate (Figure 32–11), the sulfate donor for sulfated proteoglycans (see Chapter 48)
and for sulfate conjugates of drugs; and the methyl group donor S-adenosylmethionine (Figure 32–12). GTP serves as an allosteric regulator and
as an energy source for protein synthesis, and cGMP (Figure 32–10) serves as a second messenger in response to nitric oxide (NO) during
relaxation of smooth muscle (see Chapter 49). UDP-sugar derivatives participate in sugar epimerizations and in biosynthesis of glycogen,
glucosyl disaccharides, and the oligosaccharides of glycoproteins and proteoglycans (see Chapters 47 & 48). UDP-glucuronic acid forms the
urinary glucuronide conjugates of bilirubin (see Chapter 31) and of many drugs, including aspirin. CTP participates in biosynthesis of
phosphoglycerides, sphingomyelin, and other substituted sphingosines (see Chapter 24). Finally, many coenzymes incorporate nucleotides as
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well as structures similar to purine and pyrimidine nucleotides (see Table 32–2).
Figure 32–10.
cAMP, 3',5'-cyclic AMP, and cGMP, 3', 5'-cyclic GMP.
Figure 32–11.
Adenosine 3'-phosphate-5'-phosphosulfate.
Figure 32–12.
S-Adenosylmethionine.
Table 32–2. Many Coenzymes and Related Compounds Are Derivatives of Adenosine Monophosphate
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Coenzyme
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R
R'
R″
n
H
H
0
Amino acid
H
H
2–
H
PO3
2–
1
H
PO32–
1
Active methionine
Methionine
Amino acid adenylates
Active sulfate
SO3
a
3',5'-Cyclic AMP
1
NADb
Nicotinamide
H
H
2
NADPb
Nicotinamide
PO32–
H
2
FAD
Riboflavin
H
H
Coenzyme A
Pantothenate
H
a
PO3
2
2–
2
Replaces phosphoryl group.
b
R is a B vitamin derivative.
Nucleoside Triphosphates Have High Group Transfer Potential
Acid anhydrides, unlike phosphate esters, have high group transfer potential.
G0' for the hydrolysis of each of the two terminal ( and )
phosphoryl groups of nucleoside triphosphates is about –7 kcal/mol (–30 kJ/mol). The high group transfer potential of purine and pyrimidine
nucleoside triphosphates permits them to function as group transfer reagents, most frequently of the -phosphoryl group. Cleavage of an acid
anhydride bond typically is coupled with a highly endergonic process such as covalent bond synthesis—eg, polymerization of nucleoside
triphosphates to form a nucleic acid.
SYNTHETIC NUCLEOTIDE ANALOGS ARE USED IN CHEMOTHERAPY
Synthetic analogs of purines, pyrimidines, nucleosides, and nucleotides modified in the heterocyclic ring or in the sugar moiety have numerous
applications in clinical medicine. Their toxic effects reflect either inhibition of enzymes essential for nucleic acid synthesis or their incorporation
into nucleic acids with resulting disruption of base-pairing. Oncologists employ 5-fluoro- or 5-iodouracil, 3-deoxyuridine, 6-thioguanine and
6-mercaptopurine, 5- or 6-azauridine, 5- or 6-azacytidine, and 8-azaguanine (Figure 32–13), which are incorporated into DNA prior to cell
division. The purine analog allopurinol, used in treatment of hyperuricemia and gout, inhibits purine biosynthesis and xanthine oxidase activity.
Cytarabine is used in chemotherapy of cancer, and azathioprine, which is catabolized to 6-mercaptopurine, is employed during organ
transplantation to suppress immunologic rejection (Figure 32–14).
Figure 32–13.
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Selected synthetic pyrimidine and purine analogs.
Figure 32–14.
Arabinosylcytosine (cytarabine) and azathioprine.
Nonhydrolyzable Nucleoside Triphosphate Analogs Serve as Research Tools
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Synthetic, nonhydrolyzable analogs of nucleoside triphosphates (Figure 32–15) allow investigators to distinguish the effects of nucleotides due to
phosphoryl transfer from effects mediated by occupancy of allosteric nucleotide-binding sites on regulated enzymes.
Figure 32–15.
Synthetic derivatives of nucleoside triphosphates incapable of undergoing hydrolytic release of the terminal phosphoryl group. (Pu/Py, a purine or
pyrimidine base; R, ribose or deoxyribose.) Shown are the parent (hydrolyzable) nucleoside triphosphate (top) and the unhydrolyzable -methylene
(center) and -imino derivatives (bottom).
DNA & RNA ARE POLYNUCLEOTIDES
The 5'-phosphoryl group of a mononucleotide can esterify a second hydroxyl group, forming a phosphodiester. Most commonly, this second
hydroxyl group is the 3'-OH of the pentose of a second nucleotide. This forms a dinucleotide in which the pentose moieties are linked by a
3',5'-phosphodiester bond to form the "backbone" of RNA and DNA. The formation of a dinucleotide may be represented as the elimination of
water between two mononucleotides. Biologic formation of dinucleotides does not, however, occur in this way because the reverse reaction,
hydrolysis of the phosphodiester bond, is strongly favored on thermodynamic grounds. However, despite an extremely favorable
G, in the
absence of catalysis by phosphodiesterases, hydrolysis of the phosphodiester bonds of DNA occurs only over long periods of time.
Consequently, DNA persists for considerable periods and has been detected even in fossils. RNAs are far less stable than DNA since the
2'-hydroxyl group of RNA (absent from DNA) functions as a nucleophile during hydrolysis of the 3',5'-phosphodiester bond.
Posttranslational modification of preformed polynucleotides can generate additional structures such as pseudouridine, a nucleoside in which
D-ribose is linked to C-5 of uracil by a carbon-to-carbon bond rather than by the usual
-N-glycosidic bond. The nucleotide pseudouridylic acid (
) arises by rearrangement of a UMP of a preformed tRNA. Similarly, methylation by S-adenosylmethionine of a UMP of preformed tRNA forms
TMP (thymidine monophosphate), which contains ribose rather than deoxyribose.
Polynucleotides Are Directional Macromolecules
Phosphodiester bonds link the 3'- and 5'-carbons of adjacent monomers. Each end of a nucleotide polymer thus is distinct. We therefore refer to
the "5'-end" or the "3'-end" of a polynucleotide, the 5'-end being the one with a free or phosphorylated 5'-hydroxyl.
The base sequence or primary structure of a polynucleotide can be represented as shown below. The phosphodiester bond is represented by P
or p, bases by a single letter, and pentoses by a vertical line.
Where all the phosphodiester bonds are 3'
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5', a more compact notation is possible:
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This representation indicates that the 5'-hydroxyl—but not the 3'-hydroxyl—is phosphorylated. The most compact representation shows only the
base sequence with the 5'-end on the left and the 3'-end on the right. The phosphoryl groups are assumed to be present, but not shown.
SUMMARY
Under physiologic conditions, the amino and oxo tautomers of purines, pyrimidines, and their derivatives predominate.
Nucleic acids contain, in addition to A, G, C, T, and U, traces of 5-methylcytosine, 5-hydroxymethylcytosine, pseudouridine ( ), or
N-methylated heterocycles.
Most nucleosides contain D-ribose or 2-deoxy-D-ribose linked to N-1 of a pyrimidine or to N-9 of a purine by a
whose syn conformers predominate.
-glycosidic bond
A primed numeral locates the position of the phosphate on the sugars of mononucleotides (eg, 3'-GMP, 5'-dCMP). Additional
phosphoryl groups linked to the first by acid anhydride bonds form nucleoside diphosphates and triphosphates.
Nucleoside triphosphates have high group transfer potential and participate in covalent bond syntheses. The cyclic phosphodiesters
cAMP and cGMP function as intracellular second messengers.
Mononucleotides linked by 3'
5'-phosphodiester bonds form polynucleotides, directional macromolecules with distinct 3'- and
5'-ends. For pTpGpTp or TGCATCA, the 5ʹ-end is at the left, and all phosphodiester bonds are 3'
5'.
Synthetic analogs of purine and pyrimidine bases and their derivatives serve as anticancer drugs either by inhibiting an enzyme of
nucleotide biosynthesis or by being incorporated into DNA or RNA.
REFERENCES
Adams RLP, Knowler JT, Leader DP: The Biochemistry of the Nucleic Acids, 11th ed. Chapman & Hall, 1992.
Blackburn GM, Gait MJ: Nucleic Acids in Chemistry & Biology. IRL Press, 1990.
Pacher P, Nivorozhkin A, Szabo C: Therapeutic effects of xanthine oxidase inhibitors: renaissance half a century after the discovery of allopurinol.
Pharmacol Rev 2006;58:87. [PMID: 16507884]
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