Textbook - BIOLOGICAL CHEMISTRY - Gubsky Yu.I. - 2000

Chapter IV. MOLECULAR MECHANISMS OF HEREDITY AND REALIZATION OF GENETIC INFORMATION

CHAPTER 19. NUCLEOTIDE BIOSYNTHESIS

19.3. BIOSYNTHESIS OF DEOXYRIBONUCLEOTIDES

The Biosynthesis of various classes of RNA requires purine (ATP, GTP) and pyrimidine (CTP, UTP) ribonucleotides, whereas the biosynthesis of DNA requires deoxyribonucleotides of the purine series (dATP, dGTP) and the pyrimidine series (dCTP, dTTP [TTP]).

The precursors of deoxyribonucleotides in Cells are ribonucleotides in the form of nucleoside diphosphates (NDPs) (predominantly) and nucleoside triphosphates (NTPs).

Class="center">Mechanism of The conversion of ribonucleotides to deoxyribonucleotides

The conversion of NDPs into the corresponding dNDPs is achieved by reducing the hydroxyl group at the C-2' position of ribose to form 2'-deoxyribose. The donor of reducing equivalents in this process is reduced NADP (NADPH + H⁺):

This reaction is actually a complex biochemical process involving: a low-molecular-weight (MW ~ 12 kDa) SH-containing protein, thioredoxin, and two redox Enzymes (reductases) — thioredoxin reductase and Ribonucleotide reductase — which together form the Electron Transport Chain for the reduction of NDPs to dNDPs (Fig. 19.4).

Fig. 19.4. Scheme of The transfer of reducing equivalents from (NADPH + H⁺) to ribonucleoside diphosphates:

E₁ — FADH — thioredoxin reductase;

ribonucleotide reductase;

SH

T — thioredoxin.

The process of reducing NDPs to dNDPs via protons and electrons from (NADPH + H⁺) consists of the following reactions:

(1) transfer of hydrogen from (NADPH + H⁺) to the FAD of the flavin enzyme thioredoxin reductase;

(2) transfer of hydrogen from reduced thioredoxin reductase to the SH-groups of thioredoxin;

(3) transfer of hydrogen from reduced thioredoxin to the SH-groups of ribonucleotide reductase;

(4) transfer of hydrogen from the reduced sulfhydryl groups of ribonucleotide reductase to NDPs, resulting in The formation of dNDPs.

Formation of dATP, dGTP, and dUTP

Purine deoxyribonucleotides are formed via the mechanism described above:

as well as the pyrimidine deoxyribonucleotide dUDP:

The dNDPs formed in these reactions are converted into the corresponding dNTPs (precursors for DNA biosynthesis) through Reactions Catalyzed by nucleoside diphosphate Kinases:

Biosynthesis of Thymidine NUCLEOTIDES

The biosynthesis of thymidine nucleotides, which also contain 2'-deoxyribose, begins with thymidylate (thymidine 5'-monophosphate, dTMP, TMP). The direct precursor of dTMP is deoxyuridine 5'-monophosphate (dUMP), which is formed from deoxyuridine 5'-diphosphate (dUDP) via dephosphorylation:

The conversion of dUMP to dTMP—the final step in forming the nucleotide required for DNA biosynthesis—proceeds via the methylation of dUMP According to the following reaction:

This process is catalyzed by the enzyme thymidylate synthase, with N⁵,N¹⁰-methylene-H₄-folate acting as a coenzyme, which is oxidized to dihydrofolate during the reaction. For folate to continue functioning as a coenzyme, tetrahydrofolate must be regenerated in a reaction catalyzed by Dihydrofolate Reductase:

The formation of thymidine nucleoside di- and triphosphates—dTDP (TDP), dTTP (TTP)—occurs via their phosphorylation by ATP in kinase-catalyzed reactions:

Inhibitors of dTMP Synthesis as Antitumor Agents

The biosynthesis of the four deoxyribonucleoside triphosphates (dATP, dGTP, dCTP, and dTTP) required for METABOLISM/36.html">DNA Replication is practically quiescent during the mitotic (reproductive) resting phase of The Cell (G₀) and becomes activated during the Cell Cycle stages preceding mitosis (Chapter 20). Consequently, chemical compounds that block the de novo synthesis of these dNTPs prevent genomic DNA Replication and Cell Division—a concept that forms the pharmacological basis of many antitumor drugs.

It is precisely this mechanism that underlies the inhibition of malignant tumor cell division by agents that block the Synthesis of the thymidylate dTMP (Fig. 19.5), namely:

Fig. 19.5. Mechanisms of the antitumor action of compounds blocking dTMP synthesis (E₁ — thymidylate synthase; E₂ — dihydrofolate reductase; H₄F — tetrahydrofolate; H₂F — dihydrofolate).

1) structural analogs of dUMP capable of interacting with thymidylate synthase and blocking its catalytic activity via competitive inhibition; Examples include the antimetabolite antitumor agents 5-Fluorouracil (a uracil analog) and Ftorafur (a uridine analog that also generates free 5-fluorouracil in The Human Body):

Upon entering the body, 5-fluorouracil is converted into 5-fluoro-deoxyuridine 5'-monophosphate (5-fluoro-dUMP), which is a direct structural analog of dUMP, the substrate of thymidylate synthase; the binding of 5-fluoro-dUMP to the enzyme prevents its Active Site from interacting with the true substrate (dUMP), thereby blocking the formation of dTMP;

2) pterin derivatives Aminopterin and Methotrexate, which share structural similarities with a portion of the Folic acid molecule, act as structural analogs in biochemical reactions and thus serve as Competitive Inhibitors of dihydrofolate reductase; the inhibition of this enzyme's catalytic activity prevents the regeneration of H₄-folate from H₂-folate, thereby disrupting dTMP biosynthesis.