Uracil phosphoribosyltransferase

Such conversions in mammals are not very efficient, however, with the exception of, perhaps, the orotate phosphoribosyltransferase, which is a component of the pyrimidine nucleotide biosynthetic pathway (Figure 10.9). A very active uracil phosphoribosyltransferase has been isolated from microorganisms. It converts uracil to UMP using PRPP. 

UMP also can by synthesized from uracil and PRPP by uracil phosphoribosyltransferase. 

Finally, the uracil phosphoribosyltransferase of beef erythrocytes recognizes the 2,4-dioxypyrimidine moiety of xanthine and uric acid, and attaches the phosphoribosyl group to the 3-position of these purines, as it is analogous with the N-1 of uracil. Uric acid 3-ribonucleoside is found as a constituent of beef erythrocytes, and presumably arises by dephosphorylation of the nucleotide derivative (48, 49). 

The yeast orotate phosphoribosyltransferase reaction is unusual in that it is readily reversible, in contrast to the subsequently discovered purine and uracil phosphoribosyltransferase reactions (see Chapters 5 and 12). The formation of orotidylate and the pyrophosphorolysis of orotidylate readily proceed to equilibrium the equilibrium constant for the forward reaction is about 0.1 14)- The reversibility of this reaction was the basis for the earlier name of the enzyme, orotidylate pyrophosphorylase. 

A pyrimidine phosphoribosyltransferase activity with a broader specificity than the yeast enzyme has been demonstrated in animal tissues. Highly purified preparations from calf thymus (15) and beef erythrocytes (16) accepted orotate and 5-fluorouracil as substrates. Uracil phosphoribosyltransferase activity has also been demonstrated in extracts from mouse leukemia cells. Fluorouracil is a better substrate for this enzyme than uracil at pH 7.5, possibly because the acid dissociation constant for the analogue (pif 8.15) is higher than that of uracil (pK, 9.45) (17). This reasoning would suggest that the anionic form of the substrate might be the species required by the enzyme. This enzyme has been implicated in the... 

Uracil phosphoribosyltransferase has been demonstrated in the uracil-requiring protozoan, Teirahymena pyriformis, and has been partly purified, but the substrate specificity of the preparation was not investigated 17). 

In summary, a catabolic function for uridine phosphorylase is clear. However, this enzyme may also participate in the anabolism of uracil, which is the evident function of uracil phosphoribosyltransferase. The operation of these apparently alternative routes of uracil anabolism in living cells has not yet been evaluated. 

In the pyrimidines, a mammalian erythrocyte enzyme, pyrimidine phosphoribosyltransferase, converts uracil, orotic acid, and thymine into nucleotides as follows ... 

Similar salvage pathways exist for pyrimidines. Pyrimidine phosphoribosyltransferase will reconnect uracil, but not cytosine, to PRPP. 

The presenee of various nueleoside phosphorylase, phosphotransferase and nucleoside kinase activities has been described (109,110). Phosphoribosyltransferase activity was found only for uracil and orotate. 

The first reaction is catalysed by orotate phosphoribosyltransferase (orotidine 5 -phosphate pyrophosphate phosphoribosyltransferase, EC 2.4.2.10) which is readily reversible. The equilibrium constant for the forward reaction [109] is about 0.1. The reaction is specific for orotate (the enzyme usually does not accept uracil) and some synthetic analogues of orotic acid (Chapter 6). Orotate phosphoribosyltransferase activity was found in many animal tissues [110] and there are several phosphoribosyl-transferases of broad specifity which are distinct from the enzyme involved in the orotate pathway [111-113]. 

Pyrimidine ribonucleotides, like those of purines, may be synthesized de novo from amino acids and other small molecules (Chapter 11). Preformed pyrimidine bases and their ribonucleoside derivatives, derived from the diet of animals or found in the environment of cells, may be converted to ribonucleotides via nucleoside phosphorylases and nucleoside kinases. In some cells a more direct pyrimidine phosphoribosyltransferase pathway has also been recognized (Chapter 12). Ribonucleotides are catabolized by dephosphorylation, deamination, and cleavage of the glycosidic bond, to uracil. Uracil may be either oxidatively or reductively cleaved, depending on the organism involved, and can be converted to CO and NH (Chapter 13). 

Two routes are known by which the free base, uracU, can enter the ribonucleotide pool. One proceeds by the sequential actions of uridine phosphorylase and uridine-cytidine kinase (reactions 5 and 6, Fig. 12-1) this route is discussed below. The other route is by way of a single-step phosphoribosyltransferase reaction specific for uracil (reaction 4, Fig. 12-1) ... 

The orotate phosphoribosyltransferase of yeast is specific for orotate and will not accept uracil as a substrate. This enzyme occurs in most animal cells as part of the de novo pathway of pyrimidine biosynthesis, but the specificity of the animal enzyme is unknown. Animal cells have a phosphoribosyltransferase activity capable of accepting pyrimidine substrates other than orotate, but it is not clear whether this is due to a phosphoribosyltransferase distinct from that of the orotate pathway (see Chapter 11). 

More recently, Reyes (19) has shown that ceU-free extracts from cells of a transplantable rodent leukemia catalyze the PP-ribose-P-dependent synthesis of uridylate and 5-fluorouridylate the enzyme involved, apparently a phosphoribosyltransferase, was virtually absent from a line of 5-fluoro-uracil-resistant mouse leukemia L1210 cells. Because of their ability to grow without this enzyme, the resistant cells must have obtained their pyrimidine nucleotide requirements by an alternate means, possibly by the phosphorylase-kinase route (reactions 5 and 6, Fig. 12-1), or by the de novo pathway. 

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