Purine ribonucleotides

Purine ribonucleotide biosynthesis Pyrimidine ribonucleotide biosynthesis Salvage of nucleosides and nucleotides Sugar-nucleotide biosynthesis and conversions Other... 

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... 

The effect of 6-mercaptopurine on the incorporation of a number of C-labelled compounds into soluble purine nucleotides and into RNA and DNA has been studied in leukemia L1210, Ehrlich ascites carcinoma, and solid sarcoma 180. At a level of 6-mercaptopurine that markedly inhibited the incorporation of formate and glycine, the utilization of adenine or 2-aminoadenine was not affected. There was no inhibition of the incorporation of 5(or 4)-aminoimidazole-4(5)-carboxamide (AIC) into adenine derivatives and no marked or consistent inhibition of its incorporation into guanine derivatives. The conversion of AIC to purines in ascites cells was not inhibited at levels of 6-mercaptopurine 8-20 times those that produced 50 per cent or greater inhibition of de novo synthesis [292]. Furthermore, AIC reverses the inhibition of growth of S180 cells (AH/5) in culture by 6-mercaptopurine [293]. These results suggest that in all these systems, in vitro and in vivo, the principal site at which 6-mercaptopurine inhibits nucleic acid biosynthesis is prior to the formation of AIC, and that the interconversion of purine ribonucleotides (see below) is not the primary site of action [292]. Presumably, this early step is the conversion of PRPP to 5-phosphoribosylamine inhibited allosterically by 6-mercaptopurine ribonucleotide (feedback inhibition is not observed in cells that cannot convert 6-mercaptopurine to its ribonucleotide [244]. 

Studies on the mechanism of action of 6-mercaptopurine are complicated by the fact that its anabolic product, thioinosinic acid, is further metabolized by oxidation to 6-thioxanthylic acid [219] and by methylation to 6-(methylthio)purine ribonucleotide [206, 296]. the effects of which could be even more important than those of thioinosinic acid itself, since the methylthio compound is about 20 times as potent as a feedback inhibitor [289]. 

Dietary purines are not an important source of uric acid. Quantitatively important amounts of purine are formed from amino acids, formate, and carbon dioxide in the body. Those purine ribonucleotides not incorporated into nucleic acids and derived from nucleic acid degradation are converted to xanthine or hypoxanthine and oxidized to uric acid (Figure 36-7). Allopurinol inhibits this last step, resulting in a fall in the plasma urate level and a decrease in the size of the urate pool. The more soluble xanthine and hypoxanthine are increased. 

The Km values for 5 -nucleotides are in the range of 0.2 mAf for purine ribonucleotides and higher (2 mAf range) for pyrimidine ribonucleotides (20). The enzyme is inhibited competitively by purine and pyrimidine bases, nucleosides, 2 - and 3 -mononucleotides, and NMN NAD and NADP display a mixed type of inhibition against 5 -AMP hydrolysis (Ki values of 1 and 7 mAf, respectively). Takei has concluded that the active sites for the two activities although residing in the same protein are not identical. It seems possible that the enzyme may be composed of two protein subunits each with a separate activity and with active centers... 

Overview of Nucleotide Metabolism Synthesis of Purine Ribonucleotides de Novo... 

The chemical structure of the nucleotides is shown in Figure 7-21. They are purine ribonucleotides with a hydroxyl group on carbon 6 of the purine ring and a phosphate ester group on the 5 -carbon of the ribose. Nucleotides with the ester group at the 2 or 3 position are tasteless. When the ester group is removed by the action of phos-phomonoesterases, the taste activity is lost. It is important to inactivate such enzymes in foods before adding 5 -nucleotide flavor enhancers. 

The committed step in purine nucleotide biosynthesis is the conversion of PRPP into phosphoribosylamine by glutamine phosphoribosyl amidotransferase. This important enzyme is feedback-inhibited by many purine ribonucleotides. It is noteworthy that AMP and GMP, the final products of the pathway, are synergistic in inhibiting the amidotransferase. 

N4. Nierlich, D. P., and Magasanik, B., Regulation of purine ribonucleotide synthesis by end product inhibition. J. Biol. Chem. 240, 358-365 (1965). 

The tumor-inhibitory activity of analogues of purine and pyrimidine bases led Shealy and several coworkers to synthesize a series of analogues of the imidazoles in the biosynthetic pathway to purine ribonucleotides and to test these analogues for antineoplastic effects. 59 Beginning... 

Two of the more important purine analogs in use clinically are 6-mercaptopurine and 6-thioguanine. These purine antagonists and glutamine antagonists such as azaserine (Table 4-6) are major antagonists in the biosynthesis of purine bases. Before understanding the mechanism of their action, it is necessary to look at the biosynthesis of inosinic acid, the purine ribonucleotide that is the precursor to both purine bases found in DNA and RNA... 

In mammals, purine ribonucleotides are synthesized de novo from amino acids, ribose, carbon dioxide and formate as well as from preformed purine bases and nucleosides through salvage pathways. The general route for de novo biosynthesis is the same in those species of mammals, birds, yeasts and bacteria that have been studied (1). Parasitic protozoans and helminths cannot synthesize purines de novo and thus rely solely on salvage pathways (2-12). 

Whether or not pyrimidine and purine ribonucleotides are reduced by a single enzyme is a matter of some controversy. The monomeric reductase of L. leichmannii and the enzyme of E. coll where substrates bind exclusively to Bas well as calf thymus and human reductase consist of only one holoenzyme species capable to reduce all the ribonucleotides. In contrast rat liver ribonucleotide reductase during purification dissociated into one protein fraction specific for CDP reduction and one reducing the three other diphosphates from which a specific UDP reductase activity could then be obtained Other reports of differential ADP and CDP reductase activities in rodent enzyme preparations are difficult to assess. In reductase assays one can easily encounter slow reduction rates for one particular ribonucleotide when not all conditions like pH, ionic strength, effector nucleotide and dithiol concentrations, and stoichiometry of subunits, are at an optimum. Therefore differential activities not necessarily indicate different enzyme species. 

Wingaarden, J.B. Ashton, D.H. (1959). The regulation of activity of phosphoribosylpyrophosphate amidotransferase by purine ribonucleotides a potential feedback control of purine biosynthesis. ]. Biol. Chem., 234, 1492-6. 

The feedback inhibition of the first specific enzyme of purine biosynthesis, amidophos-phoribosyltransferase, by the purine ribonucleotides AMP and GMP was described in a partially purified enzyme system over 20 years ago(l). The molecular basis for this inhibition, namely the aggregation of active subunits to an enzymatically inactive larger form in the presence of purine ribonucleotides, was described with the amidophosphoribosyl-transferase obtained from human placenta seven years ago(2). Phosphoribosylpyrophos-phate (PRPP) activates the mammalian enzyme by causing disaggregation of the inhibited large form to the catalytically active small form. 

Holmes, E.W., Wyngaarden, J.B., and Kelley, W.N. Human glutamine phosphoribosylpyrophosphate amidotransferase Two molecular forms interconvertible by purine ribonucleotides and phosphoribosylpyrophosphate. J. Biol. Chem., 248 6035,1973. 

Anti-Tumour Effects of Purine Ribonucleotide Analogues... 

PP-Ribose-P is the most important ribose phosphate donor for purine metabolism (see Chapters 7, 8) and participates in several important reactions of pyrimidine metabolism (Chapters 11, 12) it also transfers this group to a number of other acceptors (Chapter 5). In the course of studies of purine ribonucleotide biosynthesis, a product of the reaction of ribose-5-P and ATP was isolated and eventually identified as 5-phosphoribosyl 1-pyrophosphate. The pyrophosphate group is in the a-configuration, is quite labile, and almost certainly reacts enzymaticaUy as the magnesium complex. It has been chemically synthesized by Tener and Khorana 33). 

Purine ribonucleotides may be S nithe8ized de novo from amino acids and other small molecules (Chapter 7), or formed from preformed bases and ribonucleosides which may be derived from the diet or found in the environment of cells (Chapter 8). Ribonucleotides of adenine and guanine may be converted one into the other (Chapter 9). Uric acid is formed from purine ribonucleotides by dephosphorylation, deamination, cleavage of glycosidic bonds, and oxidation, and in turn is converted to CO and NH in some organisms (Chapter 10). 

A number of purine nucleotide analogues can mimic the effects of natural purine ribonucleotides on PP-ribose-P amidotransferase and thereby effect feedback inhibition of the entire pathway (69, 60). Such nucleotide analogues may be synthesized by cells from purine bases or nucleosides presented to them. A number of compounds with potent inhibitory properties, such as 6-mercaptopurine, 6-thioguanine, and 6-methylmercaptopurine ribonucleoside, inhibit purine biosynthesis de novo in this way, although it is not known whether this action is responsible for the growth inhibition. 

The pathway of purine ribonucleotide biosynthesis de novo is summarized below. The enzymes involved are listed in Table 7-IV. 

PURINE RIBONUCLEOTIDE SYNTHESIS FROM PURINE BASES AND RIBONUC LEO SIDES... 

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