Reduction of ribonucleoside

The reduction of ribonucleoside triphosphates by various dithiols which are capable of intramolecular cyclization on oxidation (dihydrolipoate, dithioerythritol, dithiothreitol) yields 2 -deoxyribonucleoside triphosphates. These reactions also require 5-deoxyadenosylcorrinoids. 

REDUCTION OF RIBONUCLEOSIDE DIPHOSPHATES FORMS DEOXYRIBONUCLEOSIDE DIPHOSPHATES... 

Figure 34-5. Reduction of ribonucleoside diphosphates to 2 -deoxyribonucleoside diphosphates.

Deoxyrihonucleotides are generally formed by reduction of ribonucleoside diphosphates. This involves a series of redox reactions in which NADP+ and FAD play a role (see Section 15.1.1), with a subsequent electron transport chain. DNA contains thymine rather than uracil, so thymidine triphosphate (dTTP) is a requirement. Methylation of dUMP to dTMP is a major route to thymine nucleotides, and is dependent upon N, A °-methylenetetrahydrofolate as the source of the methyl group (see Box 11.13). 

Deoxyribonucleotides, the precursors of DNA, are formed in E. coli by the reduction of ribonucleoside diphosphates. These conversions are catalyzed by ribonucleotide reductase. Electrons are transferred from NADPH to sulfhydryl groups at the active sites of this enzyme by thioredoxin or glutaredoxin. A tyrosyl free radical generated by an iron... 

The adenosylcobalamin-dependent RTPR catalyzes the reduction of ribonucleoside triphosphates (NTPs) to deoxyribonucleoside triphosphates (dNTPs) for DNA biosynthesis according to Equation (18). 

What is conversion of ribonucleotides to deoxyri-bonucleotides Deoxyribonucleotides for DNA synthesis are produced by the reduction of ribonucleoside diphosphates to deoxyribonucleoside diphosphates. 

As shown in Fig. 1, the enzyme catalyzes the reduction of ribonucleoside diphosphates (Fig. lA) by dithiothreitol (Fig. IB). K values for CDP and DTT are 70 x Af and 20 mAf, respectively. The requirement for a dithiol suggests that, as for other class II RNRs, such as the extensively studied enzyme from Lactobacillus leichmannii, the hydrogen donor is very likely to be a dithiol protein such as thioredoxin or glutaredoxin. However, there is still no experimental evidence that an archaeal thioredoxin operates as an electron source for RNRs. The enzyme also requires AdoCbl for which a value of 1 pAf has been obtained (Fig. 1C). Finally, the reaction has an optimal temperature of 80° (Fig. ID), with very little activity at 30°. How AdoCbl resists such a high temperature and how the enzyme controls Co-C bond homolysis required for catalysis in thermophilic AdoCbl-dependent enzymes is an intriguing question. These properties are shared by other isolated thermophilic class II RNRs (Table II). 

Fig.20 Schematic illustration of the reduction of ribonucleoside triphosphates to 2 -des-oxyribonucleoside triphosphates, as catalyzed by ribonucleotide reductases [182]...

When cytidine was the sole pyrimidine source (Table 14r-II), polynucleotide cytidylate and deoxycytidylate had identical specific activities and were incorporated without dilution. This is convincing evidence that in E. colt, reduction of ribonucleoside derivatives is able to supply all of the deoxyribose for DNA components without appreciable contribution from other sources, such as the postulated sequence involving deoxyri-boaldolase mentioned above. It will be recognized that this result does not preclude the possibility that alternate routes of deoxyribosyl biosynthesis may operate in circumstances other than the special ones of the Karlstrdm and Larsson experiment. The details of ribonucleotide reduction are discussed at length in Chapter 16. 

The deoxyribonucleotides are derived primarily, if not entirely, by reduction of ribonucleoside phosphates each of the several known ribonucleotide reductases (see Chapter 16) accepts as substrates phosphate esters of all four ribonucleosides, adenosine, guanosine, cytidine, and uridine. The ultimate fate of the uracil-containing reduction product, dUDP (or dUTP, in some microorganisms) is conversion to thymidine phosphates by way of deoxyuridylate and the thymidylate s mthetase reaction. 

The concept that the deoxyribosyl groups of DNA nucleotides are derived by reduction of ribonucleoside derivatives is based on a wide array of experimental evidence. 

The isolated reductase (above) was used in assays to direct further fractionation experiments which culminated in the isolation of the physiological reducing system this turned out to be a previously unreci ized hydrogen transport system. This system, which connects ribonucleotide reductase to the NADPH-NADP+ system, was found to be a two-component system con.sisting of a small sulfhydryl protein, thioredoxin, and a flavoprotein, thioredoxin reductase. Thioredoxin is the reductant which specifically interacts with the ribonucleotide reductase. In the presence of catalytic amounts of thioredoxin, the thioredoxin reductase will link NADPH with the reduction of ribonucleoside diphosphates as follows ... 

In the reduction of ribonucleoside triphosphates, the L. leichmannii enzyme is able to use as reductant, dihydrolipoate, dithiothreitol, or dithio-erythritol, the oxidized forms of which are cyclic disulfides it has been shown that for each mole of dihydrolipoate oxidized, a mole of ribonucleoside triphosphate is reduced. The physiological reductant for the L. leichmannii reductase is a two-component, hydrogen transport system analogous to the thioredoxin-thioredoxin reductase ssrstem of E. coli the two components of the L. leichmannii system have been purified and are generally similar to those of the E. coli system, although their molecular weights are distinctive 7). The thioredoxin-thioredoxin reductase pair from E. coli will serve the L. leichmannii reductase, although the converse is not true. 

Influence of Various Deoxyribonucleoside Triphosphates on Reduction of Ribonucleoside Triphosphates - ... 

Ribonucleotide Reductase. The ribonucleotide reductases catalyze the reduction of ribonucleoside-diphosphates (or triphosphates) to the corresponding 2 -deoxyribonucleoside-diphosphates (or triphosphates), processes of preeminent importance for the biosynthesis of DNA (see Table 2, entry 4) (65,86). A variety of metal-containing cofactors have been discovered in the ribonucleotide reductases investigated to date (eg, a binuclear iron center in the mammalian and in the E. coli ribonucleoside diphosphate reductase) and the oxidation of two protein thiols to a disulfide unit is indicated as the direct source of the two reduction equivalents. The reductase from Lactobacillus leichmanii employs coenzyme B12 as cofactor in its (normal) base-on form and acts on purine- or pyrimidine-based ribonucleoside-triphosphates. Its crystal structure reveals not only the arrangement of the bound corrinoid cofactor, but also how the enzyme is... 

Ribonucleotide reductase of Escherichia coli, which catalyzes the reduction of ribonucleoside 5 -diphosphates to 2 -deoxynucleoside 5 -diphos-phates, consists of two nonidentical subunits, proteins B1 and B2. In the presence of Mg , the two subunits form a 1 1 complex of active enzyme. When separated, neither subunit has any known biological activity. Protein B1 has a molecular weight of 160,000, contains the active dithiols, is capable of interacting with thioredoxin, and contains... 

Another reaction that depends on adenosylcobalamin is the reduction of ribonucleoside triphosphates to the corresponding 2 -deoxy compounds, the building blocks of deoxyribonucleic acids. Methylcobalamin is formed, e. g., in the methyl-ation of homocysteine to methionine with N -... 

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