Nucleotides, deoxy

Severe combined immunodeficiency disease The enzyme adenosine deaminase degrades deoxyadenosine which is produced during DNA degradation (Chapter 10). Deficiency of the enzyme results in accumulation of deoxyadenosine which is a substrate for adenosine kinase and leads to production of deoxyadenosine and deoxyquanosine triphosphates, which reach high concentrations. This disturbs the balance of deoxy nucleotides which results in failure of DNA replication. This enzyme is normally present in lymphocytes so that a deficiency prevents proliferation of the lymphocytes, which is essential in combatting an infection. Consequently, patients are very susceptible to infections. This is one disease that is effectively treated by gene therapy. 

Radiochemicals tritiated 2 -deoxythymidine 5 -triphosphate [me/fty/-3H]dTTP, 2 -deoxyadenosine 5 -triphosphate [2,8-3H]dATP, 2 -deoxyguanosine 5 -triphos-phate [8-3H]dGTP or 2 -deoxycytidine 5 -triphosphate [5-3H]dCTP. The radiospecificities of the deoxy nucleotides usually range between 15 and 30 Ci/mmol. They can be obtained from several companies such as Amersham (Buckinghamshire, UK), ICN (Costa Mesa, CA), NEN (DuPont Ltd., Hertfordshire, UK), Moravek Biochemicals (Brea, CA), and so forth. They can be used at 2 pCi/assay. 

The operation of the anomeric effect in furanosyl derivatives is less clear. A vast amount of data has been accumulated on nucleotides and deoxy nucleotides, but the bulky substituents at C4 and Cl of the ribofuranose ring ensure that the conformational choice for these compounds is between N (VI) and S (VII), in both of which the aglycone is eclipsing one lone pair on oxygen (regarding them as equivalent) [24]. 

Table 1. Erythrocyte (deoxy)nucleotides during treatment with dCyd/THU.

Two major UMPH isozymes are also detected in Chinese hamster and rat tissues (not illustrated here). As in the mouse they differ in their substrate specificity and show consistent tissue differences in expression. However in these rodents the more anodal isozyme is designated UMPH-1 since it exhibits a narrow specificity and prefers the pyrimidine substrates UMP, CMP and dCMP the less anodal isozyme is designated UMPH-2 in hamster and rat since it displays activity with a broad range of substrates and preference for the deoxy nucleotides dUMP, dTMP and dIMP. 

Site-directed mutagenesis targets a selected enzyme (protein) for modification of its activity. Use is made of chemically synthesized deoxy-nucleotide possessing discrete changes in sequence from that present in the genome. Expression of this altered gene in a host system is then expected to yield the required modified protein (Reikofski and Tao,... 

Lim SE, Copeland WC. Differential incorporation and removal of antiviral deoxy-nucleotides by human DNA polymerase gamma. J Biol Chem 2001 276(26) 23616-23. 

As mentioned above all four deoxynucleoside triphosphates are needed for substantial synthesis to take place. As much as 50% of the deoxy-nucleotide present may be polymerized, and in the absence of any one triphosphate, DNA s3Tithesis is reduced 99.5%. If only one of the triphosphates is present, a single residue or a small number of residues may be incorporated into the chain-ends of the DNA present, without a detectable increase in size of the DNA molecule (373). 

Wolter, A, Biernat, J., and Koster, H. (1986) Polymer support oligonucleotide synthesis XX Synthesis of a henhectacosa deoxy nucleotide by use of a dimeric phosphoramidite synthon. Nucleosides Nucleotides 5, 65-77... 

Fig. 6. P-NMR spectra at 109.3 MHz of sonicated alternating purine-pyrimidine deoxy-nucleotide duplexes A, poly(dAdT)-poly(dAdT) at 24 °C B, poly(dA-br dU)-prfy(dA-br dU) at 30X C, poly(dAdU) poly(dAdU) at 3I C D, poly(dIdC)-poly(dIdC) at 39 C E, poly(dGdC) poly(dGdC)at23 C allin0.1 MNaQand0.05 mAfEDTA,pH6-7,except(B) in S toM Tris and 0.1 laM EDTA. Chemical shifts arc upiield positive from internal TMP. Reproduced with permission, fiom Cohen et al., 1981. Copyright 1981 American Chemical Society.

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