Orotidylate decarboxylation

The pyrimidine ring is assembled first and then linked to ribose phosphate to form a pyrimidine nucleotide. PRPP is the donor of the ribose phosphate moiety. The synthesis of the pyrimidine ring starts with the formation of carbamoylaspartate from carbamoyl phosphate and aspartate, a reaction catalyzed by aspartate transcarbamoylase. Dehydration, cyclization, and oxidation yield orotate, which reacts with PRPP to give orotidylate. Decarboxylation of this pyrimidine nucleotide yields UMP. CTP is then formed by the amination of UTP. 

A common intermediate for all the nucleotides is 5-phosphoribosyl-l-diphosphate (PRPP), produced by successive ATP-dependent phosphorylations of ribose. This has an a-diphosphate leaving group that can be displaced in Sn2 reactions. Similar Sn2 reactions have been seen in glycoside synthesis (see Section 12.4) and biosynthesis (see Box 12.4), and for the synthesis of aminosugars (see Section 12.9). For pyrimidine nucleotide biosynthesis, the nucleophile is the 1-nitrogen of uracil-6-carboxylic acid, usually called orotic acid. The product is the nucleotide orotidylic acid, which is subsequently decarboxylated to the now recognizable uridylic acid (UMP). 

Finally orotidylate is decarboxylated to yield UMP, which of course contains one of the bases of RNA. Cellular kinases convert UMP to UTP. Transfer of an amido nitrogen from glutamine by CTP synthetase converts UTP to CTP this reaction uses an ATP high-energy phosphate. 

At this stage, orotate couples to ribose, in the form of 5-phosphoribosyl-l-pyrophosphate (PRPP), a form of ribose activated to accept nucleotide bases. PRPP is synthesized from ribose-5-phosphate, formed by the pentose phosphate pathway, by the addition of pyrophosphate from ATP. Orotate reacts with PRPP to form orotidylate, a pyrimidine nucleotide. This reaction is driven by the hydrolysis of pyrophosphate. The enzyme that catalyzes this addition, pyrimidine phosphoribosyltransferase, is homologous to a number of other phosphoribosyltransferases that add different groups to PRPP to form the other nucleotides. Orotidylate is then decarboxylated to form uridylate (IMP), a major pyrimidine nucleotide that is a precursor to RNA. This reaction is catalyzed by orotidylate decarboxylase. 

Orotidylate decarboxylase is one of the most proficient enzymes known. In its absence, decarboxylation is extremely slow and is estimated to take place once every 78 million years with the enzyme present, it takes place... 

In the next step of pyrimidine biosynthesis, the entire aspartate molecule adds to carbamoyl phosphate in a reaction catalyzed by aspartate transcarbamoylase. The molecule subsequently closes to produce a ring (catalyzed by dihydroorotase), which is oxidized to form orotic acid (or its anion, orotate) through the actions of dihydroorotate dehydrogenase. The enzyme orotate phosphoribosyl transferase catalyzes the transfer of ribose 5-phosphate from PRPP to orotate, producing orotidine 5 -phosphate, which is decarboxylated by orotidylic acid dehydrogenase to form... 

With in vivo experiments, Hurlbert and Potter ) first showed that in rat liver, uridine nucleotides were intermediates in the conversion of orotate to nucleic acid pyrimidines the first of the three uridine phosphates to become labeled in this process was the monophosphate, uridylate (UMP) IS). The synthesis of uridylate from orotate takes place in two steps (a) the condensation of orotate with PP-ribose-P to form orotidylate (orotidine 5 -monophosphate, or OMP), and (b) decarboxylation of orotidylate. 

The irreversible decarboxylation step, catalyzed by orotidylate decarboxylase, has been demonstrated in various animal tissues. 

Bresnick (52) has concluded that in the partially hepatectomized rat, during the first 12 hours of liver regeneration, the activity of orotate phosphoribosyltransferase determines the rate of synthesis of the uridine phosphates. The potential activity of orotidylate decarboxylase is in excess of that of orotate phosphoribosyltransferase. This probably accounts for the virtual absence of orotidylate in animal tissues. The decarboxylation step is irreversible and may be subject to feedback inhibition by uridylate, which is a competitive inhibitor of the enzyme in rat liver and yeast (19). 

CioHi3N20,iP 368.193 Formed in the biosynthetic pathway in yeast. Decarboxylation by Orotidylate decarboxylase affords Uridine 5 -phos-phate which is the route to Uridine and its derivatives de novo and consequently one of the most important processes in nucleic acid synthesis. Trihydrate (as Na salt). Moffatt, J.G. et al., J.A.C.S., 1963, 85, 1118 (synth)... 

Uridylic acid, the nucleotide found in RNA, is formed by decarboxylation of orotidylic acid in the presence of orotidine-5 -phosphate decarboxylase, an enzyme purified from yeast. This is an irreversible reaction that has been observed in bacteria, birds, and several mammalian tissues. The antimetabolite 6-azauracil blocks orotidylic acid decarboxylase. 

The final steps of pyrimidine biosynthesis novo which are catalyzed by two sequential enzymes, orotate phosphoribosyltransfer-ase (OPRT) and orotidylic decarboxylase (ODC), involve the PP-ribose P dependent conversion of orotic acid to orotidine-5 -monophosphate (OMP) followed by decarboxylation at the 7 position to form uridine 5 -monophosphate (UMP) (Fig. 1). UMP is then utilized further in the synthesis of nucleic acids and co-enzymes. Defects at this site in this metabolic pathway are important for they can result in "pyrimidine starvation" from depletion of the intracellular pool of pyrimidine nucleotides. In man the rare genetic disease, orotic aciduria, involves a deficiency of both OPRT and ODC (Type 1) (Smith, Sullivan and Huguley, 1961) or, less commonly, only ODC (Type II) (Fox, 0 Sullivan and Firken, 1969). 

The riboside of orotic acid, orotidine, was isolated subsequently from the culture medium of a uridine-requiring Neurospora mutant (350). Orotidine was readily split to orotic acid because of unusually great acid lability this offered a plausible explanation for previous failures to isolate a conjugated form of orotic acid from those natural sources that 3oelded the free acid (338, 342, 343). With the current knowledge of the individual steps in pyrimidine biosynthesis (Section VI, D, 2), it is likely that orotidine was derived from orotidylic acid (orotidine 5 -phosphate), and that a genetic block in the mutant organism prevented decarboxylation of oro-... 

The decarboxylation of orotidylic acid resulted in the formation of uridylic acid (UMP) 83) (Fig. 22). The enzyme has been purified from yeast and named orotidine 5 -phosphate decarboxylase this enzymic step... 

The details of the biosynthesis of purines and pyrimidines are discussed in Chapter 18. An interesting point of difference in the synthesis of the purine and pyrimidine ring systems is that the purine ring is synthesized as part of a nucleotide (8) whereas the parent pyrimidine, orotic acid, is synthesized as such. The element of pentose-phosphate is added to the completed pyrimidine ring to form the nucleotide orotidylic acid. Oro-tidylic acid is then decarboxylated to yield uridylic acid (da). 

Many potent pyrimidine antagonists have also been obtained by isosteric replacement in the heterocyclic ring. Oxonic acid (Fig. 12) an analogue of orotic acid, prevents the conversion of the latter to orotidylic acid (Fig. 2). A later stage of pyrimidine biosynthesis, the decarboxylation of orotidylic acid to form uridylic acid (Fig. 2) is strongly inhibited by 6-azauracil (Fig. 12 this chemical should be correctly termed 4-azauracil). The riboside of 6-azainacil acts on the same pathways as the base, but its inhibition of the orotidylate carboxylase enzyme is some 20 times more potent... 

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