Orotidine 5'-phosphate decarboxylase
Encyclopedia
Orotidine 5’-phosphate decarboxylase (OMP decarboxylase) or orotidylate decarboxylase is an enzyme
involved in pyrimidine
biosynthesis
. It catalyzes the decarboxylation
of orotidine monophosphate (OMP) to form uridine monophosphate
(UMP). The function of this enzyme is essential to the de novo biosynthesis of the pyrimidine nucleotides uridine triphosphate, cytidine triphosphate
, and thymidine triphosphate
. OMP decarboxylase has been a frequent target for scientific investigation because of its demonstrated extreme catalytic efficiency and its usefulness as a selection marker for yeast
strain engineering.
residues positioned within the active site of the enzyme.
>
The exact mechanism by which OMP decarboxylase catalyzes its reaction has been a subject of rigorous scientific investigation. The driving force for the loss of the carboxyl linked to the C6 of the pyrimidine ring comes from the close proximity of an aspartate residue carboxyl group in the enzyme's active site, which destabilizes the ground state relative to the transition state of the uncatalyzed reaction. There have been multiple hypotheses about what form the transition state takes before protonation of the C6 carbon occurs to yield the final product. Many studies investigated the binding of a potent inhibitor of OMP decarboxylase, 6-hydroxy uridine monophosphate (BMP, a barbituric acid
derivative), within the active site, to identify which essential amino acid residues are directly involved with stabilization of the transition state. (See figure of enzyme bound to BMP) Several mechanisms for enzymatic decarboxylation of OMP have been proposed, including protonation at O2 to form a zwitterionic species as an intermediate , anion stabilization of O4 , or nucleophilic attack at C5 . Current consensus suggests that the mechanism proceeds through a stabilized carbanion at the C6 after loss of carbon dioxide. This mechanism was suggested from studies investigating kinetic isotope effects in conjunction with competitive inhibition and active site mutagenesis.
In this mechanism the short-lived carbanion species is stabilized by a nearby lysine residue, before it is quenched by a proton. (See schematic of catalytic mechanism)
>
, OMP decarboxylase is a single-function enzyme. However, in mammals, OMP decarboxylase is part of a single protein with two catalytic activities. This bifunctional enzyme is named UMP synthase
and it also catalyzes the preceding reaction in pyrimidine nucleotide biosynthesis, the transfer of ribose 5-phosphate
from 5-phosphoribosyl-1-pyrophosphate to orotate to form OMP. In organisms utilizing OMP decarboxylase, this reaction is catalyzed by orotate phosphoribosyltransferase
.
) leads to auxotrophy in uracil. in addition, a function OMP decarboxylase renders yeast strains sensitive to the molecule 5-fluoro orotic acid (5-FOA). The establishment of the URA3 gene as a selection
marker with both positive and negative selection strategies has made the controlled expression of OMP decarboxylase a significant laboratory tool for the investigation of yeast genetics.
Enzyme
Enzymes are proteins that catalyze chemical reactions. In enzymatic reactions, the molecules at the beginning of the process, called substrates, are converted into different molecules, called products. Almost all chemical reactions in a biological cell need enzymes in order to occur at rates...
involved in pyrimidine
Pyrimidine
Pyrimidine is a heterocyclic aromatic organic compound similar to benzene and pyridine, containing two nitrogen atoms at positions 1 and 3 of the six-member ring...
biosynthesis
Biosynthesis
Biosynthesis is an enzyme-catalyzed process in cells of living organisms by which substrates are converted to more complex products. The biosynthesis process often consists of several enzymatic steps in which the product of one step is used as substrate in the following step...
. It catalyzes the decarboxylation
Decarboxylation
Decarboxylation is a chemical reaction that releases carbon dioxide . Usually, decarboxylation refers to a reaction of carboxylic acids, removing a carbon atom from a carbon chain. The reverse process, which is the first chemical step in photosynthesis, is called carbonation, the addition of CO2 to...
of orotidine monophosphate (OMP) to form uridine monophosphate
Uridine monophosphate
Uridine monophosphate, also known as 5'-uridylic acid and abbreviated UMP, is a nucleotide that is used as a monomer in RNA. It is an ester of phosphoric acid with the nucleoside uridine...
(UMP). The function of this enzyme is essential to the de novo biosynthesis of the pyrimidine nucleotides uridine triphosphate, cytidine triphosphate
Cytidine triphosphate
Cytidine triphosphate is a pyrimidine nucleoside triphosphate.CTP is a substrate in the synthesis of RNA.CTP is a high-energy molecule equal to ATP, but its role in the organism is more specific than that of ATP....
, and thymidine triphosphate
Thymidine triphosphate
5-Methyluridine triphosphate or m5UTP is one of five nucleoside triphosphates. It is the ribonucleoside triphosphate of thymidine, but the 5-methyluridine nomenclature is used because the term thymidine triphosphate is used for the deoxyribonucleoside by convention.-References:...
. OMP decarboxylase has been a frequent target for scientific investigation because of its demonstrated extreme catalytic efficiency and its usefulness as a selection marker for yeast
Yeast
Yeasts are eukaryotic micro-organisms classified in the kingdom Fungi, with 1,500 species currently described estimated to be only 1% of all fungal species. Most reproduce asexually by mitosis, and many do so by an asymmetric division process called budding...
strain engineering.
Catalysis
OMP decarboxylase is known for being an extraordinarily efficient catalyst capable of accelerating the uncatalyzed reaction rate by a factor of 1017. To put it in perspective, the enzyme can catalyze the substrate in 18 milliseconds, in a reaction that would take 78 million years to complete without the enzyme. This extreme enzymatic efficiency is especially interesting because OMP decarboxylases uses no cofactor and contains no metal sites or prosthetic groups. The catalysis relies on a handful of charged amino acidAmino acid
Amino acids are molecules containing an amine group, a carboxylic acid group and a side-chain that varies between different amino acids. The key elements of an amino acid are carbon, hydrogen, oxygen, and nitrogen...
residues positioned within the active site of the enzyme.
The exact mechanism by which OMP decarboxylase catalyzes its reaction has been a subject of rigorous scientific investigation. The driving force for the loss of the carboxyl linked to the C6 of the pyrimidine ring comes from the close proximity of an aspartate residue carboxyl group in the enzyme's active site, which destabilizes the ground state relative to the transition state of the uncatalyzed reaction. There have been multiple hypotheses about what form the transition state takes before protonation of the C6 carbon occurs to yield the final product. Many studies investigated the binding of a potent inhibitor of OMP decarboxylase, 6-hydroxy uridine monophosphate (BMP, a barbituric acid
Barbituric acid
Barbituric acid or malonylurea or 6-hydroxyuracil is an organic compound based on a pyrimidine heterocyclic skeleton. It is an odorless powder soluble in water. Barbituric acid is the parent compound of barbiturate drugs, although barbituric acid itself is not pharmacologically active...
derivative), within the active site, to identify which essential amino acid residues are directly involved with stabilization of the transition state. (See figure of enzyme bound to BMP) Several mechanisms for enzymatic decarboxylation of OMP have been proposed, including protonation at O2 to form a zwitterionic species as an intermediate , anion stabilization of O4 , or nucleophilic attack at C5 . Current consensus suggests that the mechanism proceeds through a stabilized carbanion at the C6 after loss of carbon dioxide. This mechanism was suggested from studies investigating kinetic isotope effects in conjunction with competitive inhibition and active site mutagenesis.
In this mechanism the short-lived carbanion species is stabilized by a nearby lysine residue, before it is quenched by a proton. (See schematic of catalytic mechanism)
Unicellular vs multicellular function
In yeast and bacteriaBacteria
Bacteria are a large domain of prokaryotic microorganisms. Typically a few micrometres in length, bacteria have a wide range of shapes, ranging from spheres to rods and spirals...
, OMP decarboxylase is a single-function enzyme. However, in mammals, OMP decarboxylase is part of a single protein with two catalytic activities. This bifunctional enzyme is named UMP synthase
Uridine monophosphate synthetase
Uridine monophosphate synthetase is the enzyme that catalyses the formation of uridine monophosphate , an energy-carrying molecule in many important biosynthetic pathways...
and it also catalyzes the preceding reaction in pyrimidine nucleotide biosynthesis, the transfer of ribose 5-phosphate
Ribose 5-phosphate
Ribose 5-phosphate is both a product and an intermediate of the pentose phosphate pathway. The last step of the oxidative reactions in the pentose phosphate pathway is the production of ribulose-5-phosphate. Depending on the body's state, ribulose-5-phosphate can reversibly isomerize to...
from 5-phosphoribosyl-1-pyrophosphate to orotate to form OMP. In organisms utilizing OMP decarboxylase, this reaction is catalyzed by orotate phosphoribosyltransferase
Orotate phosphoribosyltransferase
Orotate phosphoribosyltransferase is an enzyme involved in pyrimidine biosynthesis. It catalyzes the formation of orotidine 5'-monophosphate from orotate and phosphoribosyl pyrophosphate...
.
Importance in Yeast Genetics
Mutations in the gene encoding OMP decarboxylase in yeast (URA3URA3
URA3 is gene on Chromosome V in Saccharomyces . Its systematic name is YEL021W.URA3 is a gene that encodes orotidine 5-phosphate decarboxylase , which is an enzyme that catalyzes one reaction involved in the synthesis of pyrimidine ribonucleotides in yeast RNA.Loss of ODCase activity leads to a...
) leads to auxotrophy in uracil. in addition, a function OMP decarboxylase renders yeast strains sensitive to the molecule 5-fluoro orotic acid (5-FOA). The establishment of the URA3 gene as a selection
Selection
In the context of evolution, certain traits or alleles of genes segregating within a population may be subject to selection. Under selection, individuals with advantageous or "adaptive" traits tend to be more successful than their peers reproductively—meaning they contribute more offspring to the...
marker with both positive and negative selection strategies has made the controlled expression of OMP decarboxylase a significant laboratory tool for the investigation of yeast genetics.