Nucleoside reductase

Metabolic incorporation experiments indicate the presence of nucleoside reductase and thymidylate synthase activities. There is a bifunctional thymidylate synthase-dihydrofolate reductase similar to that in the kinetoplastids (91). 

The molyhdopterin cofactor, as found in different enzymes, may be present either as the nucleoside monophosphate or in the dinucleotide form. In some cases the molybdenum atom binds one single cofactor molecule, while in others, two pterin cofactors coordinate the metal. Molyhdopterin cytosine dinucleotide (MCD) is found in AORs from sulfate reducers, and molyhdopterin adenine dinucleotide and molyb-dopterin hypoxanthine dinucleotide were reported for other enzymes (205). The first structural evidence for binding of the dithiolene group of the pterin tricyclic system to molybdenum was shown for the AOR from Pyrococcus furiosus and D. gigas (199). In the latter, one molyb-dopterin cytosine dinucleotide (MCD) is used for molybdenum ligation. Two molecules of MGD are present in the formate dehydrogenase and nitrate reductase. 

Adenosine deaminase deficiency is associated with an immunodeficiency disease in which both thymus-derived lymphocytes (T cells) and bone marrow-derived lymphocytes (B cells) are sparse and dysfunctional. Purine nucleoside phosphorylase deficiency is associated with a severe deficiency of T cells but apparently normal B cell function. Immune dysfunctions appear to result from accumulation of dGTP and dATP, which inhibit ribonucleotide reductase and thereby deplete cells of DNA precursors. 

Puech F, Gosselin G, Lefebvre I, Pompon A, Aubertin A-M, Kirn A, Im-bach J-L. Intracellular delivery of nucleoside monophosphates through a reductase-mediated activation process. Antiviral Res 1993 22 155-174. 

Deoxynucleotides for DNA synthesis are made at the nucleoside diphosphate level and then have to be phosphorylated up to the triphosphate using a kinase and ATP. The reducing equivalents for the reaction come from a small protein, thioredoxin, that contains an active site with two cysteine residues. Upon reduction of the ribose to the 2 -deoxyri-bose, the thioredoxin is oxidized to the disulfide. The thioredoxin(SS) made during the reaction is recycled by reduction with NADPH by the enzyme thioredoxin reductase. 

DNA and RNA synthesis De novo formation of purine and pyrimidine nucleotide Nucleoside diphosphate reductase Thymidylate synthase Polymerase reactions Chapter 20... 

The phosphoribosyltiansferases The nucleoside kinases Nucleoside phosphokinases Nucleotide reductases Methylases and demethylases Other anabolic enzymes Catabolism... 

The synthesis of purine nucleotides (1) starts from IMP. The base it contains, hypoxanthine, is converted in two steps each into adenine or guanine. The nucleoside monophosphates AMP and CMP that are formed are then phos-phorylated by nucleoside phosphate kinases to yield the diphosphates ADP and GDP, and these are finally phosphorylated into the triphosphates ATP and CTP. The nucleoside triphosphates serve as components for RNA, or function as coenzymes (see p. 106). Conversion of the ribonucleotides into deoxyribo-nucleotides occurs at the level of the diphosphates and is catalyzed by nucleoside diphosphate reductase (B). 

The DNA component deoxythymidine triphosphate (dTTP) is synthesized from UDP in several steps. The base thymine, which only occurs in DNA (see p. 80), is formed by meth-ylation of dUMP at the nucleoside monophosphate level. Thymidylate synthase and its helper enzyme dihydrofolate reductase are important target enzymes for cytostatic drugs (see p. 402). 

In the first step, thioredoxin reductase reduces a small redox protein, thioredoxin, via enzyme-bound FAD. This involves cleavage of a disulfide bond in thioredoxin. The resulting SH groups in turn reduce a catalytically active disulfide bond in nucleoside diphosphate reductase ( ribonucleotide reductase ). The free SH groups formed in this way are the actual electron donors for the reduction of ribonucleotide diphosphates. 

ADP as a substrate in enzyme reactions, ADENYLATE KINASE (or MYOKINASE) ATP SYNTHASE CREATINE KINASE NUCLEOSIDE DIPHOSPHATE KINASE PHOSPHOGLYCERATE KINASE PYRUVATE KINASE RIBONUCLEOTIDE REDUCTASE SULFATE ADENYLYLTRANSFERASE (ADP) [ADP]/[ATP] ratio,... 

Gemcitabine is intracellularly activated by nucleoside kinases to diphosphate and triphosphate nucleosides. Gemcitabine diphosphate inhibits DNA synthesis by inhibiting ribonucleotide reductase while gemcitabine triphosphate competes with deoxycytidine triphosphate for incorporation into DNA. Gemcitabine is used for the treatment of non-small cell lung carcinoma and of adenocarcinoma of the pancreas. It has to be administred intravenously and is eliminated by metabolism with an elimination half-life of approximately 50 minutes. Its spectrum of adverse effects is comparable to that of 5-FU. 

Fluoro and 3, 3-difluoromethylene nucleosides have been prepared as inhibitors of ribonucleotide diphosphate reductase (cf. Chapter 7). ... 

Crystallographic analysis has been used quite effectively to solidify structural determinations. Several examples include dihydrofolate reductase complexes , nucleosides <1999AXC1335, 1999AXC1560>, zopi-clone <2001CC2204>, and immucillin inhibitors <2003BBR917>. 

Nucleoside monophosphates are converted to their triphosphates by enzymatic phosphorylation reactions. Ribonucleotides are converted to deoxyribonucleotides by ribonucleotide reductase, an enzyme with novel mechanistic and regulatory characteristics. The thymine nucleotides are derived from dCDP and dUMP. 

Ribonucleotide reductase ribonucleoside diphosphate reductase) is a multisubunit enzyme (two identical B1 subunits and two identical B2 subunits) that is specific for the reduction of nucleoside diphosphates (ADP, GDP, CDP, and UDP) to their deoxy-forms (dADP, dGDP, dCDP, and dUDP). The immediate donors of the hydrogen atoms needed for the reduction of the 2-hydroxyl group are two sulfhydryl groups on the enzyme itself, which, during the reaction, form a disulfide bond (Figure 22.12). 

Ribonucleotide reductases are discussed in Chapter 16. Some are iron-tyrosinate enzymes while others depend upon vitamin B12, and reduction is at the nucleoside triphosphate level. Mammalian ribonucleotide reductase, which may be similar to that of E. coli, is regarded as an appropriate target for anticancer drugs. The enzyme is regulated by a complex set of feedback mechanisms, which apparently ensure that DNA precursors are synthesized only in amounts needed for DNA synthesis.273 Because an excess of one deoxyribonucleotide can inhibit reduction of all... 

Gregus, Z. and Nemeti, B. (2002) Purine nucleoside phosphorylase as a cytosolic arsenate reductase. Toxicological Sciences, 70(1), 13-19. 

Gemcitabine is phosphorylated initially by the enzyme deoxycytidine kinase and then by other nucleoside kinases to the di- and triphosphate nucleotide forms, which then inhibit DNA synthesis. Inhibition is considered to result from two actions inhibition of ribonucleotide reductase by gemcitabine diphosphate, which reduces the level of deoxyribonucleoside triphosphates required for the synthesis of DNA and incorporation of gemcitabine triphosphate into DNA. Following incorporation of gemcitabine nucleotide, only one additional nucleotide can be added to the growing DNA strand, resulting in chain termination. 

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