BIOCHEMISTRY: A TEXTBOOK FOR MEDICAL UNIVERSITIES - E. S. Severin - 2004

SECTION 10. NUCLEOTIDE METABOLISM

V. Biosynthesis of Pyrimidine Nucleotides

Like purine NUCLEOTIDES, the pool of pyrimidine nucleotides is primarily synthesized de novo from simple precursors, with only 10 – 20% of the total amount being formed via Salvage Pathways from nitrogenous bases or nucleosides.

A. De novo synthesis of pyrimidine nucleotides

Unlike purine synthesis, in which the heterocyclic base is built directly on a ribose-5-phosphate residue, the pyrimidine ring is assembled from simple precursors—glutamine, СО₂, and aspartic acid—and is subsequently attached to ribose-5-phosphate provided by PRPP.

This process takes place in The Cell Cytosol. The Synthesis of the key pyrimidine nucleotide, UMP, involves 3 Enzymes, 2 of which are multifunctional.

Formation of dihydroorotate

In mammals, the key regulatory step in Pyrimidine Nucleotide Biosynthesis is the synthesis of carbamoyl phosphate from glutamine, СО₂, and ATP. This reaction is catalyzed by carbamoyl phosphate synthetase II (CPS II) and occurs in the cell cytosol (Fig. 10-12). In this reaction, the NH₂ group of carbamoyl phosphate is derived from the amide group of glutamine, which distinguishes it from mitochondrial carbamoyl phosphate synthesis during urea production from СО₂, NH₃, and ATP, which is catalyzed by CPS I.

Class="center">Fig. 10-12. Synthesis of carbamoyl phosphate.

Carbamoyl phosphate used for pyrimidine nucleotide formation is produced by a multifunctional enzyme that, In addition to CPS II activity, contains the catalytic domains for aspartate transcarbamoylase and dihydroorotase. This enzyme is designated as the CAD enzyme, named after the initial letters of the enzymatic activities harbored within the individual catalytic domains of this protein. The Integration of the first three Enzymes of the metabolic pathway into a single multifunctional complex ensures that nearly all carbamoyl phosphate synthesized in the initial reaction is channeled into interaction with aspartate to form carbamoylaspartate. Water is then eliminated from this intermediate to yield the cyclic product, dihydroorotate (Fig. 10-13).

Fig. 10-13. De novo biosynthesis of UMP.

Upon dissociation from the CAD enzyme, dihydroorotate undergoes dehydrogenation by a NAD-dependent dihydroorotate dehydrogenase, yielding the free pyrimidine base, orotic acid, or orotate.

Formation of UMP

In the cytosol, orotate serves as a substrate for a bifunctional enzyme, UMP synthase, which exhibits both orotate phosphoribosyltransferase and OMP decarboxylase activities. Initially, the phosphoribosyl group from PRPP is transferred to orotate to form the nucleotide orotidine-5'-monophosphate (OMP), the decarboxylation of which yields uridine-5'-monophosphate (UMP).

Thus, the six successive reactions of pyrimidine nucleotide synthesis are carried out by three enzymes encoded in The Human Genome by three distinct structural genes.

Biosynthesis of UDP, UTP, and cytidine nucleotides

Catalyzed by specific nucleoside monophosphate (NMP) and nucleoside diphosphate (NDP) Kinases, UMP is converted into UDP and UTP via The transfer of the $\gamma$-phosphate group from ATP to the appropriate substrate.

NMP kinase catalyzes the following reaction:

UMP + ATP —> UDP + ADP,

whereas NDP kinase catalyzes:

UDP + ATP —> UTP + ADP.

CTP synthetase catalyzes the amidation of UTP (Fig. 10-14), facilitating the ATP-dependent replacement of the uracil keto group with the amide group of glutamine to form cytidine-5'-triphosphate (CTP).

Fig. 10-14. Synthesis of CTP from UTP.

B. ‹Salvage› pathways of pyrimidine nucleotide synthesis

The utilization of pyrimidine bases and nucleosides in salvage reactions prevents the Catabolism of these compounds into end products involving the Cleavage of the pyrimidine ring. Certain nucleotide catabolic enzymes are involved in pyrimidine resynthesis. For instance, uridine phosphorylase can ribosylate uracil in a reversible reaction to form uridine.

Uracil + Ribose-1-phosphate —> Uridine + H₃PO₄.

The conversion of nucleosides into nucleotides is catalyzed by uridine-cytidine kinase.

A portion of CMP can be converted into UMP by the action of cytidine deaminase, thereby replenishing the pool of uridylic nucleotides.

CMP + H₂O —> UMP + NH₃.

C. Regulation of Pyrimidine nucleotide synthesis

The regulatory enzyme in pyrimidine nucleotide synthesis is the multifunctional CAD enzyme. UMP and purine nucleotides act as allosteric inhibitors, whereas PRPP activates its carbamoyl synthetase activity; conversely, The activity of the aspartate transcarbamoylase domain is inhibited by CTP but activated by ATP (Fig. 10-15).

Fig. 10-15. Regulation of pyrimidine nucleotide synthesis. The CAD enzyme catalyzes reactions 1, 2, and 3; dihydroorotate dehydrogenase catalyzes reaction 4; UMP synthase catalyzes reactions 5 and 6; NMP kinase catalyzes reaction 7; NDP kinase catalyzes reaction 8; CTP synthase catalyzes reaction 9.

This regulatory mechanism prevents the excessive synthesis not only of UMP, but also of all other pyrimidine nucleotides, ensuring the balanced production of all four major purine and pyrimidine nucleotides required for RNA Synthesis.