PTGS2
Encyclopedia
Prostaglandin-endoperoxide synthase 2, also known as cyclooxygenase-2 or simply COX-2, is an enzyme
that in humans is encoded by the PTGS2 gene
.
laboratory at Brigham Young University.
) domain; an α-helical
membrane-binding moiety; and a C-terminal catalytic domain. COX enzymes are monotopic
membrane proteins; the membrane-binding domain consists of a series of amphipathic α helices with several hydrophobic amino acids exposed to a membrane monolayer. COX-1 and COX-2 are bifunctional enzymes that carry out two consecutive chemical reactions in spatially distinct but mechanistically coupled
active sites. Both the cyclooxygenase
and the peroxidase
active sites are located in the catalytic domain, which accounts for approximately 80% of the protein. The catalytic domain is homologous to mammalian peroxidases such as myeloperoxidase
.
It has been found that human PGHS-2 functions as a conformational heterodimer having a catalytic monomer (E-cat) and an allosteric monomer (E-allo). Heme
binds only to the peroxidase
site of E-cat while substrates, as well as certain inhibitors
(e.g. celecoxib
), bind the COX site of E-cat. E-cat is regulated by E-allo in a way dependent on what ligand is bound to E-allo. Substrate
and non-substrate fatty acid (FAs) and some COX inhibitors (e.g. naproxen
) preferentially bind to the COX site of E-allo. AA
can bind to E-cat and E-allo, but the affinity of AA for E-allo is 25 times that for Ecat. Palmitic acid, an efficacious stimulator of huPGHS-2, binds only E-allo in palmitic acid/murine PGHS-2 co-crystals. Non-substrate FAs can potentiate or attenuate
COX inhibitors depending on the fatty acid
and whether the inhibitor binds E-cat or E-allo. Studies suggest that the concentration and composition of the free fatty acid pool in the environment in which PGHS-2 functions in cells, also referred to as the FA tone, is a key factor regulating the activity of PGHS-2 and its response to COX inhibitors.(Figure 1)
to prostaglandin endoperoxide H2. PGHSs are targets for NSAIDs and COX-2 specific inhibitors called coxibs. PGHS-2 is a sequence homodimer. Each monomer
of the enzyme has a peroxidase
and a COX active site
. The COX enzymes catalyze the conversion of arachidonic acid
to prostaglandins in a two steps. First, hydrogen is abstracted from carbon 13 of arachidonic acid, and then two molecules of oxygen are added by the COX-2, giving PGG2. Second, PGG2 is reduced to PGH2 in the peroxidase active site. The synthesized PGH2 is converted to prostaglandins (PGD2, PGE2, PGF2R), prostacyclin
(PGI2), or thromboxane A2
by tissue-specific isomerases.(Figure 2)
Both the peroxidase and the cyclooxygenase activities are inactivated during catalysis by mechanism-based, first-order processes, which means that PGHS-2 peroxidase or cyclooxygenase activities fall to zero within 1–2 minutes, even in the presence of sufficient substrates.
autoxidation
(Figure 3.). The 13-pro(S) -hydrogen is abstracted and dioxygen traps the pentadienyl radical at carbon 11. The 11-peroxyl radical cyclizes at carbon 9 and the carbon-centered radical generated at C-8 cyclizes at carbon 12, generating the endoperoxide. The allyl
ic radical generated is trapped by dioxygen at carbon 15 to form the 15-(S) -peroxyl radical; this radical is then reduced to PGG2 . This is supported by the following evidence: 1) a significant kinetic isotope effect
is observed for the abstraction of the 13-pro (S )-hydrogen; 2) carbon-centered radicals are trapped during catalysis
; 3) small amounts of oxidation products are formed due to the oxygen trapping of an allylic radical intermediate at positions 13 and 15. Another mechanism shown in Figure 4 in which the 13-pro (S )-hydrogen is deprotonated
and the carbanion
is oxidized to a radical
is theoretically possible. However, oxygenation of 10,10-difluoroarachidonic acid to 11-(S )-hydroxyeicosa-5,8,12,14-tetraenoic acid is not consistent with the generation of a carbanion intermediate because it would eliminate fluoride to form a conjugated diene. The absence of endoperoxide-containing products derived from 10,10-difluoroarachidonic acid has been thought to indicate the importance of a C-10 carbocation in PGG2 synthesis. However, the cationic mechanism requires that endoperoxide formation comes before the removal of the 13-pro (S )-hydrogen. This is not consistent with the results of the isotope experiments of arachidonic acid
oxygenation.
, which may contribute to gastric ulceration. Since COX-2 is generally expressed only in cells where prostaglandin
s are upregulated (e.g., during inflammation), drug-candidates that selectively inhibit COX-2 are thought to show fewer side-effects.
Non-steroidal anti-inflammatory drug
s (NSAIDs) inhibit prostaglandin
production by cyclooxygenases (COX) 1 and 2. NSAIDs selective for inhibition of COX-2 are less likely than traditional drugs to cause gastrointenstinal adverse effects, but could cause cardiovascular events, such as heart failure, myocardial infarction
, and stroke
. Studies with human pharmacology
and genetics
, genetically manipulated rodents, and other animal models and randomized trials indicate that this is due to suppression of COX-2-dependent cardioprotective prostaglandins, prostacyclin
in particular.
The expression of COX-2 is upregulated in many cancers. The overexpression of COX-2 along with increased angiogenesis and GLUT-1 expression is significantly associated with gallbladder carcinomas. Furthermore the product of COX-2, PGH2
is converted by prostaglandin E2 synthase
into PGE2, which in turn can stimulate cancer progression. Consequently inhibiting COX-2 may have benefit in the prevention and treatment of these types of cancer.
The mutant allele PTGS2 5939C carriers among the Han Chinese population have been shown to have a higher risk of gastric cancer. In addition, a connection was found between Helicobacter pylori
infection and the presence of the 5939C allele.
with Caveolin 1.
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...
that in humans is encoded by the PTGS2 gene
Gene
A gene is a molecular unit of heredity of a living organism. It is a name given to some stretches of DNA and RNA that code for a type of protein or for an RNA chain that has a function in the organism. Living beings depend on genes, as they specify all proteins and functional RNA chains...
.
History
COX-2 was discovered in 1991 by the Daniel SimmonsDaniel L. Simmons
Daniel L. Simmons is a professor of chemistry and the director of the Cancer Research Center at Brigham Young University . He was the discoverer of the COX-2 enzyme that is the target of celecoxib and other COX-2 inhibitors...
laboratory at Brigham Young University.
Structure
COX-2 exists as a homodimer, each monomer with a molecular mass of about 70 kDa. The tertiary and quaternary structures of COX-1 and COX-2 enzymes are almost identical. Each subunit has three different structural domains: a short N-terminal epidermal growth factor (EGFEGF-like domain
EGF-like domain is an evolutionary conserved protein domain, of about thirty to forty amino-acid residues long, which was found in a large number of mostly animal proteins. All these repeats are found in the extracellular domain of membrane-bound proteins or in proteins known to be secreted...
) domain; an α-helical
Alpha helix
A common motif in the secondary structure of proteins, the alpha helix is a right-handed coiled or spiral conformation, in which every backbone N-H group donates a hydrogen bond to the backbone C=O group of the amino acid four residues earlier...
membrane-binding moiety; and a C-terminal catalytic domain. COX enzymes are monotopic
Integral monotopic protein
Integral monotopic proteins, are permanently attached to the membrane from one side.Three-dimensional structures of the following integral monotopic proteins have been determined:*prostaglandin H2 syntheses 1 and 2...
membrane proteins; the membrane-binding domain consists of a series of amphipathic α helices with several hydrophobic amino acids exposed to a membrane monolayer. COX-1 and COX-2 are bifunctional enzymes that carry out two consecutive chemical reactions in spatially distinct but mechanistically coupled
Coupling
A coupling is a device used to connect two shafts together at their ends for the purpose of transmitting power. Couplings do not normally allow disconnection of shafts during operation, however there are torque limiting couplings which can slip or disconnect when some torque limit is exceeded.The...
active sites. Both the cyclooxygenase
Cyclooxygenase
Cyclooxygenase is an enzyme that is responsible for formation of important biological mediators called prostanoids, including prostaglandins, prostacyclin and thromboxane. Pharmacological inhibition of COX can provide relief from the symptoms of inflammation and pain...
and the peroxidase
Peroxidase
Peroxidases are a large family of enzymes that typically catalyze a reaction of the form:For many of these enzymes the optimal substrate is hydrogen peroxide, but others are more active with organic hydroperoxides such as lipid peroxides...
active sites are located in the catalytic domain, which accounts for approximately 80% of the protein. The catalytic domain is homologous to mammalian peroxidases such as myeloperoxidase
Myeloperoxidase
Myeloperoxidase is a peroxidase enzyme that in humans is encoded by the MPO gene. Myeloperoxidase is most abundantly expressed in neutrophil granulocytes . It is a lysosomal protein stored in azurophilic granules of the neutrophil...
.
It has been found that human PGHS-2 functions as a conformational heterodimer having a catalytic monomer (E-cat) and an allosteric monomer (E-allo). Heme
Heme
A heme or haem is a prosthetic group that consists of an iron atom contained in the center of a large heterocyclic organic ring called a porphyrin. Not all porphyrins contain iron, but a substantial fraction of porphyrin-containing metalloproteins have heme as their prosthetic group; these are...
binds only to the peroxidase
Peroxidase
Peroxidases are a large family of enzymes that typically catalyze a reaction of the form:For many of these enzymes the optimal substrate is hydrogen peroxide, but others are more active with organic hydroperoxides such as lipid peroxides...
site of E-cat while substrates, as well as certain inhibitors
Enzyme inhibitor
An enzyme inhibitor is a molecule that binds to enzymes and decreases their activity. Since blocking an enzyme's activity can kill a pathogen or correct a metabolic imbalance, many drugs are enzyme inhibitors. They are also used as herbicides and pesticides...
(e.g. celecoxib
Celecoxib
Celecoxib INN is a sulfa non-steroidal anti-inflammatory drug and selective COX-2 inhibitor used in the treatment of osteoarthritis, rheumatoid arthritis, acute pain, painful menstruation and menstrual symptoms, and to reduce numbers of colon and rectum polyps in patients with familial...
), bind the COX site of E-cat. E-cat is regulated by E-allo in a way dependent on what ligand is bound to E-allo. Substrate
Substrate (chemistry)
In chemistry, a substrate is the chemical species being observed, which reacts with a reagent. This term is highly context-dependent. In particular, in biochemistry, an enzyme substrate is the material upon which an enzyme acts....
and non-substrate fatty acid (FAs) and some COX inhibitors (e.g. naproxen
Naproxen
Naproxen sodium is a nonsteroidal anti-inflammatory drug commonly used for the reduction of pain, fever, inflammation and stiffness caused by conditions such as:...
) preferentially bind to the COX site of E-allo. AA
Arachidonic acid
Arachidonic acid is a polyunsaturated omega-6 fatty acid 20:4.It is the counterpart to the saturated arachidic acid found in peanut oil, Arachidonic acid (AA, sometimes ARA) is a polyunsaturated omega-6 fatty acid 20:4(ω-6).It is the counterpart to the saturated arachidic acid found in peanut oil,...
can bind to E-cat and E-allo, but the affinity of AA for E-allo is 25 times that for Ecat. Palmitic acid, an efficacious stimulator of huPGHS-2, binds only E-allo in palmitic acid/murine PGHS-2 co-crystals. Non-substrate FAs can potentiate or attenuate
Attenuation
In physics, attenuation is the gradual loss in intensity of any kind of flux through a medium. For instance, sunlight is attenuated by dark glasses, X-rays are attenuated by lead, and light and sound are attenuated by water.In electrical engineering and telecommunications, attenuation affects the...
COX inhibitors depending on the fatty acid
Fatty acid
In chemistry, especially biochemistry, a fatty acid is a carboxylic acid with a long unbranched aliphatic tail , which is either saturated or unsaturated. Most naturally occurring fatty acids have a chain of an even number of carbon atoms, from 4 to 28. Fatty acids are usually derived from...
and whether the inhibitor binds E-cat or E-allo. Studies suggest that the concentration and composition of the free fatty acid pool in the environment in which PGHS-2 functions in cells, also referred to as the FA tone, is a key factor regulating the activity of PGHS-2 and its response to COX inhibitors.(Figure 1)
Function
Prostaglandin endoperoxide H synthase, COX 2, converts arachidonic acid (AA)Arachidonic acid
Arachidonic acid is a polyunsaturated omega-6 fatty acid 20:4.It is the counterpart to the saturated arachidic acid found in peanut oil, Arachidonic acid (AA, sometimes ARA) is a polyunsaturated omega-6 fatty acid 20:4(ω-6).It is the counterpart to the saturated arachidic acid found in peanut oil,...
to prostaglandin endoperoxide H2. PGHSs are targets for NSAIDs and COX-2 specific inhibitors called coxibs. PGHS-2 is a sequence homodimer. Each monomer
Monomer
A monomer is an atom or a small molecule that may bind chemically to other monomers to form a polymer; the term "monomeric protein" may also be used to describe one of the proteins making up a multiprotein complex...
of the enzyme has a peroxidase
Peroxidase
Peroxidases are a large family of enzymes that typically catalyze a reaction of the form:For many of these enzymes the optimal substrate is hydrogen peroxide, but others are more active with organic hydroperoxides such as lipid peroxides...
and a COX active site
Active site
In biology the active site is part of an enzyme where substrates bind and undergo a chemical reaction. The majority of enzymes are proteins but RNA enzymes called ribozymes also exist. The active site of an enzyme is usually found in a cleft or pocket that is lined by amino acid residues that...
. The COX enzymes catalyze the conversion of arachidonic acid
Arachidonic acid
Arachidonic acid is a polyunsaturated omega-6 fatty acid 20:4.It is the counterpart to the saturated arachidic acid found in peanut oil, Arachidonic acid (AA, sometimes ARA) is a polyunsaturated omega-6 fatty acid 20:4(ω-6).It is the counterpart to the saturated arachidic acid found in peanut oil,...
to prostaglandins in a two steps. First, hydrogen is abstracted from carbon 13 of arachidonic acid, and then two molecules of oxygen are added by the COX-2, giving PGG2. Second, PGG2 is reduced to PGH2 in the peroxidase active site. The synthesized PGH2 is converted to prostaglandins (PGD2, PGE2, PGF2R), prostacyclin
Prostacyclin
Prostacyclin is a member of the family of lipid molecules known as eicosanoids.As a drug, it is also known as "epoprostenol". The terms are sometimes used interchangeably.-History:...
(PGI2), or thromboxane A2
Thromboxane A2
Thromboxane A2 is a thromboxane. It is produced by activated platelets and has prothrombotic properties: it stimulates activation of new platelets as well as increases platelet aggregation. This is achieved by mediating expression of the glycoprotein complex GP IIb/IIIa in the cell membrane of...
by tissue-specific isomerases.(Figure 2)
Both the peroxidase and the cyclooxygenase activities are inactivated during catalysis by mechanism-based, first-order processes, which means that PGHS-2 peroxidase or cyclooxygenase activities fall to zero within 1–2 minutes, even in the presence of sufficient substrates.
Mechanism
The conversion of arachidonic acid to PGG2 can be shown as a series of radical reactions analogous to polyunsaturated fatty acidFatty acid
In chemistry, especially biochemistry, a fatty acid is a carboxylic acid with a long unbranched aliphatic tail , which is either saturated or unsaturated. Most naturally occurring fatty acids have a chain of an even number of carbon atoms, from 4 to 28. Fatty acids are usually derived from...
autoxidation
Autoxidation
Autoxidation is any oxidation that occurs in open air or in presence of oxygen and/or UV radiation and forms peroxides and hydroperoxides. A classic example of autoxidation is that of simple ethers like diethyl ether, whose peroxides can be dangerously explosive. It can be considered to be a slow,...
(Figure 3.). The 13-pro(S) -hydrogen is abstracted and dioxygen traps the pentadienyl radical at carbon 11. The 11-peroxyl radical cyclizes at carbon 9 and the carbon-centered radical generated at C-8 cyclizes at carbon 12, generating the endoperoxide. The allyl
Allyl
An allyl group is a substituent with the structural formula H2C=CH-CH2R, where R is the connection to the rest of the molecule. It is made up of a methylene , attached to a vinyl group . The name is derived from the Latin word for garlic, Allium sativum. Theodor Wertheim isolated an allyl...
ic radical generated is trapped by dioxygen at carbon 15 to form the 15-(S) -peroxyl radical; this radical is then reduced to PGG2 . This is supported by the following evidence: 1) a significant kinetic isotope effect
Kinetic isotope effect
The kinetic isotope effect is the ratio of reaction rates of two different isotopically labeled molecules in a chemical reaction. It is also called "isotope fractionation," although this term is somewhat broader in meaning...
is observed for the abstraction of the 13-pro (S )-hydrogen; 2) carbon-centered radicals are trapped during catalysis
Catalysis
Catalysis is the change in rate of a chemical reaction due to the participation of a substance called a catalyst. Unlike other reagents that participate in the chemical reaction, a catalyst is not consumed by the reaction itself. A catalyst may participate in multiple chemical transformations....
; 3) small amounts of oxidation products are formed due to the oxygen trapping of an allylic radical intermediate at positions 13 and 15. Another mechanism shown in Figure 4 in which the 13-pro (S )-hydrogen is deprotonated
Deprotonation
Deprotonation is the removal of a proton from a molecule, forming the conjugate base.The relative ability of a molecule to give up a proton is measured by its pKa value. A low pKa value indicates that the compound is acidic and will easily give up its proton to a base...
and the carbanion
Carbanion
A carbanion is an anion in which carbon has an unshared pair of electrons and bears a negative charge usually with three substituents for a total of eight valence electrons. The carbanion exists in a trigonal pyramidal geometry. Formally a carbanion is the conjugate base of a carbon acid.where B...
is oxidized to a radical
Radical (chemistry)
Radicals are atoms, molecules, or ions with unpaired electrons on an open shell configuration. Free radicals may have positive, negative, or zero charge...
is theoretically possible. However, oxygenation of 10,10-difluoroarachidonic acid to 11-(S )-hydroxyeicosa-5,8,12,14-tetraenoic acid is not consistent with the generation of a carbanion intermediate because it would eliminate fluoride to form a conjugated diene. The absence of endoperoxide-containing products derived from 10,10-difluoroarachidonic acid has been thought to indicate the importance of a C-10 carbocation in PGG2 synthesis. However, the cationic mechanism requires that endoperoxide formation comes before the removal of the 13-pro (S )-hydrogen. This is not consistent with the results of the isotope experiments of arachidonic acid
Arachidonic acid
Arachidonic acid is a polyunsaturated omega-6 fatty acid 20:4.It is the counterpart to the saturated arachidic acid found in peanut oil, Arachidonic acid (AA, sometimes ARA) is a polyunsaturated omega-6 fatty acid 20:4(ω-6).It is the counterpart to the saturated arachidic acid found in peanut oil,...
oxygenation.
Clinical significance
Cyclooxygenase-2 (COX-2, prostaglandin H synthase-2, PGHS-2) is unexpressed under normal conditions in most cells, but elevated levels are found during inflammation. COX-1 (prostaglandin H2 synthase 1) is constitutively expressed in many tissues and is the predominant form in gastric mucosa and in the kidneys. Inhibition of COX-1 reduces the basal production of cytoprotective PGE2 and PGI2 in the stomachStomach
The stomach is a muscular, hollow, dilated part of the alimentary canal which functions as an important organ of the digestive tract in some animals, including vertebrates, echinoderms, insects , and molluscs. It is involved in the second phase of digestion, following mastication .The stomach is...
, which may contribute to gastric ulceration. Since COX-2 is generally expressed only in cells where prostaglandin
Prostaglandin
A prostaglandin is any member of a group of lipid compounds that are derived enzymatically from fatty acids and have important functions in the animal body. Every prostaglandin contains 20 carbon atoms, including a 5-carbon ring....
s are upregulated (e.g., during inflammation), drug-candidates that selectively inhibit COX-2 are thought to show fewer side-effects.
Non-steroidal anti-inflammatory drug
Non-steroidal anti-inflammatory drug
Nonsteroidal anti-inflammatory drugs, usually abbreviated to NSAIDs or NAIDs, but also referred to as nonsteroidal anti-inflammatory agents/analgesics or nonsteroidal Anti-inflammatory medicines , are drugs with analgesic and antipyretic effects and which have, in higher doses, anti-inflammatory...
s (NSAIDs) inhibit prostaglandin
Prostaglandin
A prostaglandin is any member of a group of lipid compounds that are derived enzymatically from fatty acids and have important functions in the animal body. Every prostaglandin contains 20 carbon atoms, including a 5-carbon ring....
production by cyclooxygenases (COX) 1 and 2. NSAIDs selective for inhibition of COX-2 are less likely than traditional drugs to cause gastrointenstinal adverse effects, but could cause cardiovascular events, such as heart failure, myocardial infarction
Myocardial infarction
Myocardial infarction or acute myocardial infarction , commonly known as a heart attack, results from the interruption of blood supply to a part of the heart, causing heart cells to die...
, and stroke
Stroke
A stroke, previously known medically as a cerebrovascular accident , is the rapidly developing loss of brain function due to disturbance in the blood supply to the brain. This can be due to ischemia caused by blockage , or a hemorrhage...
. Studies with human pharmacology
Pharmacology
Pharmacology is the branch of medicine and biology concerned with the study of drug action. More specifically, it is the study of the interactions that occur between a living organism and chemicals that affect normal or abnormal biochemical function...
and genetics
Genetics
Genetics , a discipline of biology, is the science of genes, heredity, and variation in living organisms....
, genetically manipulated rodents, and other animal models and randomized trials indicate that this is due to suppression of COX-2-dependent cardioprotective prostaglandins, prostacyclin
Prostacyclin
Prostacyclin is a member of the family of lipid molecules known as eicosanoids.As a drug, it is also known as "epoprostenol". The terms are sometimes used interchangeably.-History:...
in particular.
The expression of COX-2 is upregulated in many cancers. The overexpression of COX-2 along with increased angiogenesis and GLUT-1 expression is significantly associated with gallbladder carcinomas. Furthermore the product of COX-2, PGH2
Prostaglandin H2
Prostaglandin H2 is a type of Prostaglandin which is derived from arachidonic acid and is a precursor for many other biologically significant molecules.It is acted upon by:* prostacyclin synthase to create prostacyclin...
is converted by prostaglandin E2 synthase
PTGES2
Prostaglandin E synthase 2 is an enzyme that in humans is encoded by the PTGES2 gene.The protein encoded by this gene is a membrane-associated prostaglandin E synthase, which catalyzes the conversion of prostaglandin H2 to prostaglandin E2...
into PGE2, which in turn can stimulate cancer progression. Consequently inhibiting COX-2 may have benefit in the prevention and treatment of these types of cancer.
The mutant allele PTGS2 5939C carriers among the Han Chinese population have been shown to have a higher risk of gastric cancer. In addition, a connection was found between Helicobacter pylori
Helicobacter pylori
Helicobacter pylori , previously named Campylobacter pyloridis, is a Gram-negative, microaerophilic bacterium found in the stomach. It was identified in 1982 by Barry Marshall and Robin Warren, who found that it was present in patients with chronic gastritis and gastric ulcers, conditions that were...
infection and the presence of the 5939C allele.
Interactions
PTGS2 has been shown to interactProtein-protein interaction
Protein–protein interactions occur when two or more proteins bind together, often to carry out their biological function. Many of the most important molecular processes in the cell such as DNA replication are carried out by large molecular machines that are built from a large number of protein...
with Caveolin 1.