Purine biosynthetic pathway, de novo

Glycinamide ribonucleotide transformylase (GAR Tfase) is a folate-dependent enzyme essential to the de novo purine biosynthetic pathway. It utilizes the cofactor 10-formyl tetrahydrofohc acid (10-formyl-THF) to transfer a formyl group to the primary amine of its substrate a-glycinamide ribonucleotide. Potent, and potentially selective, inhibitors of GARTfase and de novo purine biosynthesis have been shown to be promising as antitumor drugs. 

Several steps in the de novo purine biosynthetic pathway may be inhibited by antitumor agents. Steps requiring glutamine (2 and 5) are sensitive to azaserine, a glutamine analog. Its structure is shown in Figure 10.7. Steps requir-... 

A serious genetic disorder is associated with the salvage pathways, the Lesch-Nyhan syndrome. It is believed that it is caused by a failure to control the de novo purine biosynthetic pathway. In the Lesch-Nyhan syndrome, the enzyme HGPRTase is severely depressed. Because the de novo pathway is controlled largely via feedback effects of purine nucleotides, the pathway is derepressed and excessive quantities of purine nucleotides and their degradation product, uric acid, are accumulated. This results is neurologic effects, self-mutilation, and mental retardation. 

Fig. 15-16 The de novo purine biosynthetic pathway. Rib-5-P, ribose 5-phosphate P-Rib-PP, 5-phosphoribosyl 1-pyrophosphate PRA, 5-phosphoribosylamine IO-CHO-FH4, /Vl0-formyl tetrahydrofolate GAR, glycineamide ribotide FGAR. /V-formylglycineamide ribotide FGAM, /V-formylglycineamidine ribotide AIR, 5-aminoimidazole ribotide CAIR, 4-carboxy-5-aminoimidazole ribotide SAICAR, iV-succino-5-aminoimidazole-4-carboxamide ribotide AICAR, 5-aminoimidazole-4-carboxamide ribotide FAICAR, 5-formamidoimidazole-4-carboxamide ribotide sAMP, /V-succino-AMP. Enzymes (1) amido phosphoribosyltransferase (2) GAR synthetase (3) GAR transformylase (4) FGAM synthetase (5) AIR synthetase (6) AIR carboxylase (7) SAICAR synthetase (8) adenylosuecinase (9) AICAR transformylase (10) IMP cyclohydrolase (11) sAMP synthetase (12) adenylosuecinasc (13) IMP dehydrogenase (14) GMP synthetase.

Our own work (3) and that of others (2) with E. coll have shown that the de novo purine biosynthetic pathway is regulated by both a repressor molecule (pur R gene product) and by feedback inhibition. However, Chinese hamster cells are much more sensitive to feedback inhibition by adenine than E, coli and, unlike the situation in E. coli, no repression of PRPP amidotransferase or formyglycinamide biosynthesis could be detected. If repression did occur, it would have to be by a mechanism not normally associated with the purine biosynthetic pathways or at a site late in the purine bios3mthetic pathway. Moreover, the nucleotide pools of cells treated for 2 h with with actinomycin D or cycloheximide showed a substantial increase in nucleotide levels. This Increase in nucleotide concentration is probably sufficient in itself to inhibit de novo purine biosynthesis by feedback inhibition without recourse to a repression mechanism, Snyder and Henderson (10) have also reported an effect of actinomycin D on purine metabolism in Ehrlich ascites cells. In this case, there was no large effect (11% inhibition) on de novo purine biosynthesis, Snyder and Henderson (10) proposed that this decrease was due to a 29% reduction in PRPP levels as a result of increased (1,3-fold increase in ATP and 2,8-fold Increase in GTP) nucleotide pools. These observations are consistent with our data in which a 58% decrease in PRPP level is found over a 2-h period in Chinese hamster cells grown in actinomycin D, The extent of inhibition in Chinese hamster cells is much greater than that reported for Ehrlich ascites cells and may reflect a difference between cells,... 

Thioguanine (6-TG) also inhibits several enzymes in the de novo purine nucleotide biosynthetic pathway. Various metabolic lesions result, including inhibition of purine nucleotide interconversion decrease in intracellular levels of guanine nucleotides, which leads to inhibition of glycoprotein synthesis interference with the formation of DNA and RNA and incorporation of thiopurine nucleotides into both DNA and RNA. 6-TG has a synergistic action when used together with cytarabine in the treatment of adult acute leukemia. 

The so-called salvage pathways are available in many cells to scavenge free purine and pyrimidine bases, nucleosides, and mononucleotides and to convert these to metabolically useful di- and trinucleotides. The function of these pathways is to avoid the costly (energy) and lengthy de novo purine and pyrimidine biosynthetic processes. In some cells, in fact, the salvage pathways yield a greater quantity of nucleotides than the de novo pathways. The substrates for salvage reactions may come from dietary sources or from normal nucleic acid turnover processes. 

FR901483 in suppressing the immune system results from an antimetabolite activity whereby adenylosuccinate synthetase and/or adenylosuccinate lyase are inhibited. These enzymes function as key catalysts in the de novo purine nucleotide biosynthetic pathway. Addition of adenosine or deoxyadenosine (but not deoxyguanosine, deoxycytidine, uridine or thymidine) results in elimination of the immunosuppressive activity of FR901483. Thus, FR901483 may inhibit one of the key steps for adenosine biosynthesis (Scheme 1). 

Potter has used this term to include the amount, activity, and localization of an enzyme. Little is known about the amount of any of the enzymes of the purine biosynthetic pathway, although bacterial mutants deficient in almost every one of these enzyme activities have been isolated. Mature mammalian erythrocytes also cannot synthesize purines de novo the last two enzymes of this pathway are known to be present in these cells, but PP-ribose-P amidotransferase, the first enzyme, appears to be missing (SI). 

Biosynthesis de novo is the initial regulatory level to be considered. The first complete nucleotide to be formed in the purine biosynthetic pathway is inosine-5 -phosphate (IMP), and that in the pyrimidine pathway is uridine-5 -phosphate (UMP). The two pathways are operationally separate and distinct and metabolically related only in that they share some common participants such as glutamine, CO2,... 

Phosphoribosyl pyrophosphate (PRPP) is important in both, and in these pathways the structure of ribose is retained in the product nucleotide, in contrast to its fate in the tryptophan and histidine biosynthetic pathways discussed earlier. An amino acid is an important precursor in each type of pathway glycine for purines and aspartate for pyrimidines. Glutamine again is the most important source of amino groups—in five different steps in the de novo pathways. Aspartate is also used as the source of an amino group in the purine pathways, in two steps. 

As with purines, there is indirect evidence from studies in vitro that regenerating tetrathyridia of M. corti can synthesise pyrimidines de novo (315). Furthermore, aspartate transcarbamylase, the first enzyme in the pathway, has been demonstrated in Moniezia benedini (39), while five of the six pathway enzymes have been measured in H. diminuta (326). It appears, therefore, that at least some cestodes have the capacity to synthesise pyrimidines by the biosynthetic route. Little is known of pyrimidine salvage pathways in cestodes, although the key enzyme thymidine kinase has been... 

The following sections explore nature s use of domain swapping to evolve new function. These include the formation of multifunctional proteins, tandem duplication, domain recruitment, and cicular permutation (Fig. 1). The evolution of several enzymes in the purine (Fig. 2) and pyrimidine (Fig. 3) de novo biosynthetic pathways, as well as other enzymes, are discussed as illustrative examples. 

Understand the purine and pyrmidine de novo biosynthetic pathways, with special attention to enzymes controlling pathway rates and the properties of such enzymes the positive and negative effectors steps inhibited by the various antitumor agents and their mechanisms final products of the de novo pathways and how the various nucleotides are generated from them and the biosynthesis of deoxyribonucleotides and the attendant mechanisms. 

Serine Hydroxymethyltransferase Serinehydroxymethyltrans-ferase is a pyridoxed phosphate-dependent aldolase that catalyzes the cleavage of serine to glycine and methylene-tetrahydrofolate (as shown in Figure 10.5). Serine is the major source of one-carbon substituted folates for biosynthetic reactions. At times of increeised cell proliferation, the activities of serine hydroxymethyltransferase emd the enzymes of the serine biosynthetic pathway cue increased. The other product of the reaction, glycine, is also required in increased cimounts under these conditions (for de novo synthesis of purines). 

New purine bases are produced, at the nucleotide level, by the de novo biosynthetic route which commences with 5-phospho-a-D-ribosyl pyrophosphate and proceeds via 5-phospho-/3-D-ribosylamine to produce IMP in a sequence of enzyme-controlled reactions involving aminoimidazole intermediates. IMP is further converted into AMP and GMP by separate pathways (Scheme 157). 

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