Pyrimidine deoxyribonucleotides

These transcription factors activate genes that express the enzymes for purine and pyrimidine deoxyribonucleotide synthesis and DNA duplication. 

Neale, G.A.M. Mitchell, A. Finch, L.R. Enzymes of pyrimidine deoxyribonucleotide metabolism in Mycoplasma mycoides subsp. mycoides. J. Bacter-ioL, 156, 1001-1005 (1983)... 

As we have already noted, the deoxyribonucleoside phosphates occur in animal tissues in small amounts and their concentrations are increased in cells engaged in DNA synthesis. The triphosphates of the four deoxyri-bonucleosides represented in DNA have been demonstrated in cell extracts, as have the mono- and diphosphates of thymidine and deoxycytidine. The mono-, di-, and triphosphates of deoxyuridine are known as intermediary compounds in the metabolism of the pyrimidine deoxyribonucleotides. 

Metabolism of the pyrimidine deoxyribonucleotides is more complex because, in addition to transfer of phosphoryl groups, deamination and methylation reactions occur at this level. Specifically, the thymidine phosphates are derived by methylation of deoxyuridylate, and the latter may be derived from the deoxycytidine phosphates by way of deoxycytidylate deaminase. The deoxycytidine phosphates are not formed by amination of deoxyuridine phosphates, but are derived entirely from the cytidine phosphates by enzymatic reduction (Chapter 16). 

B. Formation of Pyrimidine Deoxyribonucleotides Exclusively PROM Ribonucleotides... 

That uridine or cytidine can be the exclusive precursor of the pyrimidine deoxyribonucleotide subunits of DNA was shown by Karlstrdm and Larsson ( 9) with a pyrimidine-requiring E. coli mutant, OK305. After growing the bacteria in the presence of uridine or cytidine totally labeled with C, DNA and RNA were isolated and degraded to their constituent nucleotides. The specific activities of the sugar and base portions of the nucleotides from RNA and DNA were compared (Tables 14r-I and 14-11). 

In the data of Table 14-1 it is seen that when uridine was the sole pyrimidine source, the specific activities of the sugar portions of the RNA pyrimidine nucleotides were the same as those derived from DNA. This means that the pyrimidine deoxyribonucleotides were synthesized exclusively from the same precursors as the pyrimidine ribonucleotides. There was some reduction in the isotope content of the pentose, but none in that of the base this apparently came about through exchange with cellular nonisotopic ribose phosphates by way of the reversible reaction of uridine phosphorylase. 

Figure 34-6. Regulation of the reduction of purine and pyrimidine ribonucleotides to their respective 2 -deoxyribonucleotides. Solid lines represent chemical flow. Broken lines show negative ( ) or positive ( ) feedback regulation.

Deoxyribonucleotides A purine or pyrimidine base bonded to a deoxyribose containing a... 

The two classes of nucleotide that must be synthesised are the pyrimidine and purine ribonucleotides for RNA synthesis and the deoxyribonucleotides for DNA synthesis. For the original sources of the nitrogen atoms in the bases of the pyrimidine and purine nucleotides, see Figure 20.7. The pathway for the synthesis of the pyrimidine nucleotides is... 

Regulation of the balance of the concentrations of the four deoxyribonucleotides depends on the properties of only two enzymes, the ribonucleotide reductase complex and deoxy-CMP deaminase. The balance between pyrimidine deoxynucleotides is brought about by the properties of the deoxy-CMP deaminase, which is inhibited by deoxy-TTP and stimulated by deoxy-CTP. The ribonucleotide reductase also possesses allosteric sites which bind all four deoxynucleotide triphosphates, the effect of which is to maintain approximately similar concentrations of all the triphosphates. 

The biosynthetic pathways for the pyrimidine nucleotides (2) are more complicated. The first product, UMP, is phosphorylated first to the diphosphate and then to the triphosphate, UTP. CTP synthase then converts UTP into CTP. Since pyrimidine nucleotides are also reduced to deoxyribonucleotides at the diphosphate level, CTP first has to be hydrolyzed by a phosphatase to yield CDP before dCDP and dCTP can be produced. 

DNA is a linear polymer of deoxyribonucleotides in which the sequence of purine and pyrimidine bases encodes cellular RNA and protein molecules. 

The most important pyrimidine derivatives are those upon which biological organisms depend. Cytosine 1018 and uracil 1019 are found in ribonucleic acid (RNA) in the form of their ribonucleotides, cytidine 1020 and uridine 1021, while in deoxyribonucleic acid (DNA), cytosine and thymine 1022 are found in the form of their 2 -deoxyribonucleotides, 2 -deoxycytidine 1023 and thymidine 1024. 5-Methylcytosine 1025 is also found to a small extent (c. 5%) in human DNA in the form of its 2 -deoxyriboside 1026, and 5-(hydroxymethyl)cytosine-2 -deoxyriboside 1027 has also been detected in smaller amounts <2005CBI1>. Many variants of cytosine and uracil can be found in RNA including orotic acid 1028 in the form of its ribonucleotide orotidine 1029. Other pyrimidine derivatives to have been isolated from various biological sources include 2 -deoxyuridine 1030, alloxan 1031, and toxopyrimidine (pyramine) 1032 (Figure 2). 

The sugar component in RNA is ribose, whereas in DNA it is 2-dexoyribose. In deoxyribonucleotides, the heterocyclic bases are purine bases, adenine and guanine, and pyrimidine bases, cytosine and thymine. In ribonucleotides, adenine, guanine and cytosine are present, but not thymine, which is replaced by uracil, another pyrimidine base. 

Fig. 1. Consisting of two helically intertwined strands, ihe DNA molecule is composed of dcoxyribose and phosphate. As shown here, at periodic intervals the sugar-phosphate backbones are joined together by the complementary purine and pyrimidine bases. A single base linked to a deoxyribose-pbosphate moiety constitutes a deoxyribonucleotide. Legend Solid black circles — Thymine Vertical bars = Adenine Horizontal bars = Guanine Dotted circles = Cytosine...

Purine Biosynthesis Is Regulated at Two Levels Pyrimidine Biosynthesis Is Regulated at the Level of Carbamoyl Aspartate Formation Deoxyribonucleotide Synthesis Is Regulated by Both Activators and Inhibitors... 

The pentose component of naturally occurring nucleotides is ribose or 2-deoxyribose (i.e., ribose with a hydrogen instead of a C-2 —OH). In nucleotides the purine or pyrimidine is attached to C-1 of the pentose in the /3 configuration. This means that the base is cis relative to C-5 —OH and trans relative to the C-3 —OH. The major function of deoxyribonucleotides (those that have 2-deoxyribose as the pentose) is to serve as building blocks for DNA. Although ribonucleotides similarly serve as the units for RNA synthesis, they also have a multitude of other functions in cell metabolism. In some synthetic nucleosides with therapeutic properties, other pentose components such as arabinose are present. (See fig. 12.2 for the structure of arabinose.)... 

Hydroxyurea interferes with the synthesis of both pyrimidine and purine nucleotides (see table 23.3). It interferes with the synthesis of deoxyribonucleotides by inhibiting ribonucleotide reductase of mammalian cells, an enzyme that is crucial and probably rate-limiting in the biosynthesis of DNA. It probably acts by disrupting the iron-tyrosyl radical structure at the active site of the reductase. Hydroxyurea is in clinical use as an anticancer agent. 

The manner in which the reduction of ribonucleotides to deoxyribonucleotides is regulated has been studied with reductases from relatively few species. The enzymes from E. coli and from Novikoff s rat liver tumor have a complex pattern of inhibition and activation (fig. 23.25). ATP activates the reduction of both CDP and UDP. As dTTP is formed by metabolism of both dCDP and dUDP, it activates GDP reduction, and as dGTP accumulates, it activates ADP reduction. Finally, accumulation of dATP causes inhibition of the reduction of all substrates. This regulation is reinforced by dGTP inhibition of the reduction of GDP, UDP, and CDP and by dTTP inhibition of the reduction of the pyrimidine substrates. Because evidence suggests that ribonucleotide reductase may be the rate-limiting step in deoxyribonucleotide synthesis in at least some animal cells, these allosteric effects may be important in controlling deoxyribonucleotide synthesis. 

The biosynthetic pathway to UMP starts from carbamoyl phosphate and results in the synthesis of the pyrimidine orotate, to which ribose phosphate is subsequently attached. CTP is subsequently formed from UTP. Deoxyribonucleotides are formed by reduction of ribonucleotides (diphosphates in most cells). Thy-midylate is formed from dUMP. 

The nucleotides of DNA are called deoxyribonucleotides, since they contain the sugar deoxyribose, whereas, those of RNA are called nbonucleotides since they contain nbose instead. Each nucleotide contains both a specific and a nonspecific region. The phosphate and sugar groups are the nonspecific portion of the nucleotide, while the purine and pyrimidine bases make up the specific portion. 

As much of the terminology used in molecular biology may be unfamiliar to some readers, it is appropriate to define some of the vocabulary and this is given in an appendix to this chapter. There are two types of nucleic acids, the ribonucleic acids (RNA) and the deoxyribonucleic acids (DNA). Genetic information is carried in the linear sequence of nucleotides in DNA. Each molecule of DNA contains two complementary strands of deoxyribonucleotides which contain the purine bases, adenine and guanine and the pyrimidines, cytosine and thymine. RNA is single-stranded, being composed of a linear sequence of ribonucleotides the bases are the same as in DNA with the exception that thymine is replaced by the closely related base uracil. DNA replication occurs by the polymerisation of a new complementary strand on to each of the old strands. 

The existence of metal intermediate complexes with deoxynucleotides has been elucidated by Eichhorn et al. (26). Proton NMR spectra of dAMP, dCMP, dGMP and dTMP show, especially for dAMP and dGMP, a strong reaction of Cu2+, although the interaction with the pyrimidines was markedly reduced. Further experiments employing 31P NMR spectroscopy show the broadening of the phosphate resonance of the deoxyribonucleotides of adenine and thymine (26). 

Figure 1.4 Diagrammatic representations of (a) a purine base, (b) a pyrimidine base, (c) a ribonucleotide, adenosine monophosphate (AMP) and (d) a deoxyribonucleotide, deoxyuridine monophosphate (dllMP).

Deoxyribonucleotide (Section 27.14B) ADNAbuilding block having a deoxyribose and either a purine or pyrimidine base joined together by an (V-glycosidic linkage, and a phosphate bonded to a hydroxy group of the sugar nucleus. 

Narrower specificities have been obtained with antibodies to some unusual helical structures. Poly(dG)-poly(dC) induces antibodies specific for the immunogen and unreactive with other deoxyribonucleotide polymers, such as poly(dAT) or native DNA. Double-helical polyribonucleotides with modified furanoses, such as poly(A)-poly(2 -0-methylU), induce antibodies that react with a number of polymers bearing 2 -furanose substitutions (such as methyl or ethyl groups on either the purine or pyrimidine-containing strand). Poly(G)-poly(C) induced antibodies of narrow specificity in our studies, but Lacour and co-workers obtained anti-poly(G)-poly(C) that cross-reacted with several forms of viral RNA. ... 

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