Nucleotide, adenine deamination

The isomerism existing between the pairs of nucleotides was attributed to the different locations of the phosphoryl residues in the carbohydrate part of the parent nucleoside,49 63 since, for instance, the isomeric adenylic acids are both hydrolyzed by acids to adenine, and by alkalis or kidney phosphatase to adenosine. Neither is identical with adenosine 5-phosphate since they are not deaminated by adenylic-acid deaminase,68 60 and are both more labile to acids than is muscle adenylic acid. An alternative explanation of the isomerism was put forward by Doherty.61 He was able, by a process of transglycosidation, to convert adenylic acids a" and 6 to benzyl D-riboside phosphates which were then hydrogenated to optically inactive ribitol phosphates. He concluded from this that both isomers are 3-phosphates and that the isomerism is due to different configurations at the anomeric position. This evidence is, however, open to the same criticism detailed above in connection with the work of Levene and coworkers. Further work has amply justified the original conclusion regarding the nature of the isomerism, since it has been found that, in all four cases, a and 6 isomers give rise to the same nucleoside on enzymic hydrolysis.62 62 63 It was therefore evident that the isomeric nucleotides are 2- and 3-phosphates, since they are demonstrably different from the known 5-phosphates. The decision as to which of the pair is the 2- and which the 3-phosphate proved to be a difficult one. The problem is complicated by the fact that the a and b" nucleotides are readily interconvertible.64,64... 

Several nucleotide bases undergo spontaneous loss of their exocyclic amino groups (deamination) (Fig. 8-33a). For example, under typical cellular conditions, deamination of cytosine (in DNA) to uracil occurs in about one of every 107 cytidine residues in 24 hours. This corresponds to about 100 spontaneous events per day, on average, in a mammalian cell. Deamination of adenine and guanine occurs at about l/100th this rate. 

Several of the B vitamins function as coenzymes or as precursors of coenzymes some of these have been mentioned previously. Nicotinamide adenine dinucleotide (NAD) which, in conjunction with the enzyme alcohol dehydrogenase, oxidizes ethanol to ethanal (Section 15-6C), also is the oxidant in the citric acid cycle (Section 20-10B). The precursor to NAD is the B vitamin, niacin or nicotinic acid (Section 23-2). Riboflavin (vitamin B2) is a precursor of flavin adenine nucleotide FAD, a coenzyme in redox processes rather like NAD (Section 15-6C). Another example of a coenzyme is pyri-doxal (vitamin B6), mentioned in connection with the deamination and decarboxylation of amino acids (Section 25-5C). Yet another is coenzyme A (CoASH), which is essential for metabolism and biosynthesis (Sections 18-8F, 20-10B, and 30-5A). 

A detailed discussion of the mechanism for 5 -AMP deamination is at present premature. The sigmoid relationship for substrate saturation and activation by monovalent cations and adenine nucleotides is consistent with mechanisms involving active site-effector site interaction. However, the activation brought about by this site-site interaction is a relatively... 

Fludarabine phosphate, a fluorinated deamination-resistant nucleotide analog of the antiviral agent vidarabine (9-P-D-arabinofuranosyl-adenine), is active in CLL and low-grade lymphomas. After rapid extracellular dephosphorylation to the nucleoside fludarabine, it is rephosphorylated intracellularly by deoxycytidine kinase to the active triphosphate derivative. This antimetabolite inhibits DNA polymerase, DNA primase DNA ligase, and ribonucleotide reductase, and is incorporated into DNA and RNA. The triphosphate nucleotide is an effective chain terminator when incorporated into DNA, and the incorporation of fludarabine into RNA inhibits RNA function, RNA processing, and mRNA translation. 

Figure 22.17 outlines the de novo and salvage synthetic pathways to thymine nucleotides. dUTP, an intermediate in the de novo pathways that begins with UDP, is readily recognized by DNA polymerases and can be incorporated into DNA in place of dXTP. The uracil from a dUMP residue in a DNA strand pairs with adenine (like thymine from a dXMP residue would), so there is no loss of or change in information in the DNA. However, dUMP residues can also arise from spontaneous deamination of dCMP. When this DNA is replicated, a mutation at the site will result because cytosine is meant to pair with guanine, not adenine. 

In addition to salvaging purines, most cells interconvert adenine and guanine nucleotides. Inosine monophosphate (IMP), is the common intermediate. IMP is converted into AMP by a two-step reaction catalyzed by adenylosuccinate synthetase and adenylosuccinate lyase. Guanine nucleotides are formed in a two-step reaction in which IMP is converted into xanthine monophosphate (XMP) and then aminated to GMP. Both GMP and AMP can be reconverted into IMP. Mammalian cells can also deaminate adenosine to inosine and guanine to xanthine (Fig. 6.1). 

Purine nucleosides, with the exception of adenosine, are salvaged by converting them into the base followed by phosphoribosylation. Adenosine is phosphorylated directly by adenosine kinase or deaminated by adenosine deaminase to inosine. Adenine and adenosine deaminase are present in the sporozoite and merozoite forms (64) the former is not in extracts from unsporulated oocysts (11) but the latter has apparently not been looked for. The ability to deaminate both adenine and adenosine allows this parasite to synthesize guanine nucleotides in the absence of AMP deaminase. The ratio of labeled adenine nucleotides to guanine nucleotides is about 20% higher when both adenine and adenosine are the precursors compared to the ratio obtained when hypoxanthine or inosine was used (64). This indicates that although the major route of salvage for adenine and adenosine is by conversion into hypoxanthine, there is some direct conversion of these compounds into AMP. 

Fig. 41.10. Salvage of bases. The purine bases hypoxanthine and gnanine react with PRPP to form the nucleotides inosine and gnanosine monophosphate, respectively. The enzyme that catalyzes the reaction is hypoxanthine-gnanine phosphoribosyltransferase (HGPRT). Adenine forms AMP in a reaction catalyzed by adenine phosphoribosyltransferase (APRT). Nucleotides are converted to nucleosides by 5 -nucleotidase. Free bases are generated from nncleosides by purine nucleoside phosphorylase. Deamination of the base adenine occurs with AMP and adenosine deaminase. Of the purines, only adenosine can be directly phosphorylated back to a nucleotide, by adenosine kinase.

This is small nucleolar RNA or snoRNA. These RNA species recognize their target sequence by base-pairing and then recruit specialized proteins that perform nucleotide modifications to these RNAs usually 2 O-ribose methylation, base deaminations such as adenine-to-inosine conversions, and the addition of pseudouridines. These modifications are crucial to ribosome biogenesis. 

At very low values of EC, when AMP is elevated it is deaminated via AMP deaminase to inosine monophosphate (IMP). This further displaces the adenylate kinase reaction in the direction of ATP synthesis. The IMP is dephosphorylated by nucleotide phosphatase, and the inosine is phosphorylyzed via purine nucleotide phosphorylase, releasing hypoxanthine and ribose 1-phosphate. The latter is metabolized via the pentose phosphate pathway, and most of the carbon atoms enter glycolysis. Because this course of events depletes the overall adenine nucleotide pool, and hence the scope for ATP production in the longer term, it represents a metabolic last ditch stand by the cell to extract energy even from the energy currency itself ... 

Two enzymes were selected In which proper alignment of nucleotide C3 and base moieties Is necessary for their catalytic action AMP amlnohydrolase (AMP deaminase, EC 3>3.4.6) from rabbit muscle requires substrates with an anion at C5 and catalyses deamination at C6-NH2 of adenine [42]. Vice versa, In snake venom 3 -nucleotidase (EC 3>1 3.5 hydrolysis occurs at the 5 -phosphate vdilch has to be at a defined distance from the base, as Judged by the Inactivity of 3 -phosphates [43]. 

There is little evidence regarding the cyclic operation of these reactions. One cycle may function in muscle where the extensive deamination of adenylate accompanying muscle function must be followed by its rapid resynthesis. McFall and Magasanik S3) have suggested that in cultured L cells, guanine is converted to adenine nucleotides for storage and then converted back to guanine nucleotides for incorporation into nucleic acids. 

Hypoxanthine, or 6-hydroxypurine, is also a product of nucleic acid dissimilation. It is produced by the deamination of adenine. In addition to the classical purine bases that we have just described, a group of methylated or hydroxylated purines have been discovered 6-methylaminopurine, 2-methyladenine, 2,6-dimethylaminopurine, 2,2-dimethylaminopurine, 1 -methylguanine, and 2-methylamino-6-hydroxypurine. Most of these compounds have been found in association with RNA in the form of a nucleotide linked by a 5 -phosphodiester linkage. It may be significant that these bases, present only in small amounts in mammalian tissues, are found almost exclusively in the tRNA. A small number of the bases are associated with ribosomal RNA. 

The low RBC ATP concentration is the most probable explanation for the hemolysis (1). In vitro studies of adenosine metabolism by intact patient s RBC showed as expected, a markedly decreased ATP synthesis from adenosine moreover, metabolic studies of adenylic nucleotides labelled with radioactive adenine indicate that AMP degradation (probably by hydrolysis of the phosphate ester followed by deamination of adenosine) is abnormally elevated in the patient s erythrocytes. Thus, the low RBC ATP concentration appears to be secondary to both a diminished synthesis of AMP from adenosine and an excessive catabolism of AMP. 

Insulin as well as prolactin when used separately enhanced adenine salvage into AMP (Table 1). The effect of insulin was stronger than prolactin. Insulin also resulted in increased AMP conversion to ATP, increased deamination of AMP, and to a smaller extent increased guanine nucleotide synthesis (results not presented). Prolactin also increased markedly adenine salvage into AMP, but more of the synthesized AMP was deaminated than phosphorylated. Hydrocortisone had no significant effects on this system. 

The possibility existed that adenylic acid might have been the first purine compound synthesized by the homogenate, and that the isolated inosinic acid was merely a deamination product of adenylic acid. However, labeled formate was not incorporated into the adenine nucleotides of the homogenate or into added adenylic acid. 

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