Synthesis of Pyrimidine Nucleotides

Figure 8.29 The initial reactions of glutamine metabolism in kidney, intestine and cells of the immune system. The initial reaction in all these tissues is the same, glutamine conversion to glutamate catalysed by glutaminase the next reactions are different depending on the function of the tissue or organ. In the kidney, glutamate dehydrogenase produces ammonia to buffer protons. In the intestine, the transamination produces alanine for release and then uptake and formation of glucose in the liver. In the immune cells, transamination produces aspartate which is essential for synthesis of pyrimidine nucleotides required for DNA synthesis otherwise it is released into the blood to be removed by the enterocytes in the small intestine or by cells in the liver.

A different, simpler , pathway is involved in the synthesis of pyrimidine nucleotides. A pyrimidine base (orotate), is synthesised first. Then the ribose is added from 5-phosphoribosyl 1-pyrophosphate. The two precursors for the formation of orotate are carbamoylphosphate and aspartate, which form carbamoyl aspartate, catalysed by aspartate carbamoyltransferase. 

The fact that many agents which interrupt the synthesis of pyrimidine nucleotides from orotic acid in animals can also inhibit the growth of experimental neoplasms suggests a search for additional antimetabolites whose locus of action is in this metabolic sequence. Two in vitro biological screening systems were developed for this purpose [202—207]. From a study of systems with oxidative energy sources, 5-bromo-[208—209] (Villa), 5-chloro-[210] (Vlllb) and 5-diazo-orotic acid [211] (IX) were found to inhibit the conversion of orotic acid to the uridine nucleotides by 40—100 per cent [202]. 

FIGURE 22-36 De novo synthesis of pyrimidine nucleotides biosynthesis of UTP and CTP via orotidylate. The pyrimidine is constructed from carbamoyl phosphate and aspartate. The ribose 5-phosphate is then added to the completed pyrimidine ring by orotate phosphori-bosyltransferase. The first step in this pathway (not shown here see Fig. 18-11a) is the synthesis of carbamoyl phosphate from C02 and NH), catalyzed in eukaryotes by carbamoyl phosphate synthetase II. 

The fourth step in the de novo synthesis of pyrimidine nucleotides—the conversion of dihydroorotic acid to orotic acid—is catalyzed by dihydroorotic acid dehydrogenase. The enzyme, located on the cytosolic side of the inner membrane of mitochondria, is a target for antitumor agents. 

Dihydroorotase catalyzes the intramolecular cyclization of 7V-carbamyl-L-aspartic acid to L-dihydroorotic acid. In mammals, the activity is present in a trifunctional enzyme that catalyzes the first three steps in the de novo synthesis of pyrimidine nucleotides. 

Scheme 12-9 De novo synthesis of pyrimidine nucleotides (simplified).

The purine ring is assembled from a variety of precursors glutamine, glycine, aspartate, N formyltetrahydrofolate, and (XT. The committed step in the de novo synthesis of purine nucleotides is the formation of 5-phosphoribosyIamine from PRPP and glutamine. The purine ring is assembled on ribose phosphate, in contrast with the de novo synthesis of pyrimidine nucleotides. The addition of glycine,... 

UTP is a nucleotide that is a substrate for RNA polymerase in synthesis of RNA. It is also a precursor of CTP in de novo synthesis of pyrimidine nucleotides (Figure 22.10)... 

The activation of ATCase by ATP occurs when metabolic energy is available for DNA replication and the synthesis of pyrimidine nucleotides. Feedback inhibition by CTP prevents the overproduction of pyrimidine nucleotides and the waste of precursors. 

Shafritz, D. A. and Senior, J. R. Synthesis of pyrimidine nucleotide precursors in human and rat small intestinal mucosa. Biochim. Biophys. Acta, 141, 332-341 (1967)... 

Coleman, P. F., Suttle, D. P., and Stark, G. R., 1977, Purification from hamster cells of the multifunctional protein that initiates de novo synthesis of pyrimidine nucleotides, J. Biol. Chem. 252 6379. 

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