
The PDE2 (phosphodiesterase 2)
enzyme
Enzymes () are proteins that act as biological catalysts by accelerating chemical reactions. The molecules upon which enzymes may act are called substrate (chemistry), substrates, and the enzyme converts the substrates into different molecule ...
is one of 21 different
phosphodiesterase
A phosphodiesterase (PDE) is an enzyme that breaks a phosphodiester bond. Usually, ''phosphodiesterase'' refers to cyclic nucleotide phosphodiesterases, which have great clinical significance and are described below. However, there are many o ...
s (PDE) found in
mammals. These different PDEs can be subdivided to 11 families (PDE1 – PDE11). The different PDEs of the same family are functionally related despite the fact that their
amino acid
Amino acids are organic compounds that contain both amino and carboxylic acid functional groups. Although hundreds of amino acids exist in nature, by far the most important are the alpha-amino acids, which comprise proteins. Only 22 alpha ...
sequences show considerable divergence.
The PDEs have different
substrate
Substrate may refer to:
Physical layers
*Substrate (biology), the natural environment in which an organism lives, or the surface or medium on which an organism grows or is attached
** Substrate (locomotion), the surface over which an organism lo ...
specificities. Some are
cAMP selective
hydrolase
Hydrolase is a class of enzyme that commonly perform as biochemical catalysts that use water to break a chemical bond, which typically results in dividing a larger molecule into smaller molecules. Some common examples of hydrolase enzymes are es ...
s (PDE 4, -7 and -8), others are
cGMP CGMP is an initialism. It can refer to:
*cyclic guanosine monophosphate (cGMP)
*current good manufacturing practice (cGMP)
*CGMP, Cisco Group Management Protocol, the Cisco version of Internet Group Management Protocol
The Internet Group Managem ...
selective hydrolases (PDE 5, -6 and -9) and the rest can hydrolyse both
cAMP and
cGMP CGMP is an initialism. It can refer to:
*cyclic guanosine monophosphate (cGMP)
*current good manufacturing practice (cGMP)
*CGMP, Cisco Group Management Protocol, the Cisco version of Internet Group Management Protocol
The Internet Group Managem ...
(PDE1, -2, -3, -10 and -11).
There is only one
gene
In biology, the word gene (from , ; "... Wilhelm Johannsen coined the word gene to describe the Mendelian units of heredity..." meaning ''generation'' or ''birth'' or ''gender'') can have several different meanings. The Mendelian gene is a b ...
family coding for the PDE2, which is the PDE2A. Three
splice variant
Alternative splicing, or alternative RNA splicing, or differential splicing, is an alternative splicing process during gene expression that allows a single gene to code for multiple proteins. In this process, particular exons of a gene may be i ...
s have been found, the PDE2A1, PDE2A2 and PDE2A3 (PDE2A2 has only been found in rats). PDE2A1 is
cytosol
The cytosol, also known as cytoplasmic matrix or groundplasm, is one of the liquids found inside cells ( intracellular fluid (ICF)). It is separated into compartments by membranes. For example, the mitochondrial matrix separates the mitochondri ...
ic whereas -A2 and -A3 are membrane bound. It has been suggested that different localization of PDE2A2 and -A3 is due to a unique N-terminal sequence, which is absent in PDE2A1. Despite the PDE2A
splice variant
Alternative splicing, or alternative RNA splicing, or differential splicing, is an alternative splicing process during gene expression that allows a single gene to code for multiple proteins. In this process, particular exons of a gene may be i ...
s being different, there is no known differences in their kinetic behavior.
Crystal structure
The
crystal structure
In crystallography, crystal structure is a description of the ordered arrangement of atoms, ions or molecules in a crystalline material. Ordered structures occur from the intrinsic nature of the constituent particles to form symmetric patterns t ...
of the
active site of the PDE2 enzyme has been reported.
Even though
amino acid
Amino acids are organic compounds that contain both amino and carboxylic acid functional groups. Although hundreds of amino acids exist in nature, by far the most important are the alpha-amino acids, which comprise proteins. Only 22 alpha ...
sequences, for members of the PDE family show considerable difference (25-35% identity), the overall folding, functional and structural elements of the active sites are very similar. The
active site is formed by residues that are highly conserved among all PDEs. The
binding pocket
In biology and biochemistry, the active site is the region of an enzyme where substrate molecules bind and undergo a chemical reaction. The active site consists of amino acid residues that form temporary bonds with the substrate (binding site) a ...
contains metal ion (zinc and magnesium) binding sites. The two
histidine
Histidine (symbol His or H) is an essential amino acid that is used in the biosynthesis of proteins. It contains an α-amino group (which is in the protonated –NH3+ form under biological conditions), a carboxylic acid group (which is in the ...
and two
aspartic acid
Aspartic acid (symbol Asp or D; the ionic form is known as aspartate), is an α- amino acid that is used in the biosynthesis of proteins. Like all other amino acids, it contains an amino group and a carboxylic acid. Its α-amino group is in the pr ...
residues, which bind zinc are conserved among all studied PDEs.
The structure of several other PDE iso-enzymes has been elucidated and among them few
co-crystal
Cocrystals are "solids that are crystalline single phase materials composed of two or more different molecular or ionic compounds generally in a stoichiometric ratio which are neither solvates nor simple salts." A broader definition is that cocryst ...
structures, with inhibitors residing in the active site.
The co-crystal structures for PDE4B, PDE4D and
PDE5
Cyclic guanosine monophosphate-specific phosphodiesterase type 5 is an enzyme () from the phosphodiesterase class. It is found in various tissues, most prominently the corpus cavernosum penis, corpus cavernosum and the retina. It has also been ...
A have revealed two common features of inhibitor binding to PDEs. One is a
planar
Planar is an adjective meaning "relating to a plane (geometry)".
Planar may also refer to:
Science and technology
* Planar (computer graphics), computer graphics pixel information from several bitplanes
* Planar (transmission line technologies), ...
ring structure of the inhibitors, which align in the
active site of the enzymes and the other is a conserved
glutamine
Glutamine (symbol Gln or Q) is an α-amino acid that is used in the biosynthesis of proteins. Its side chain is similar to that of glutamic acid, except the carboxylic acid group is replaced by an amide. It is classified as a charge-neutral ...
residue (the "glutamine switch" mentioned below), which is essential for
nucleotide
Nucleotides are organic molecules consisting of a nucleoside and a phosphate. They serve as monomeric units of the nucleic acid polymers – deoxyribonucleic acid (DNA) and ribonucleic acid (RNA), both of which are essential biomolecul ...
recognition and selectivity.
Substrate selectivity
As mentioned above, PDE2 is able to hydrolyze both
cAMP and cGMP, whereas some other members of the PDE family are selective for either of the two cyclic nucleotides. The variability in selectivity towards either cAMP or cGMP is thought to be determined by a so-called "glutamine switch". The "glutamine switch" is an invariant
glutamine
Glutamine (symbol Gln or Q) is an α-amino acid that is used in the biosynthesis of proteins. Its side chain is similar to that of glutamic acid, except the carboxylic acid group is replaced by an amide. It is classified as a charge-neutral ...
found in all PDEs, for which the
crystal structure
In crystallography, crystal structure is a description of the ordered arrangement of atoms, ions or molecules in a crystalline material. Ordered structures occur from the intrinsic nature of the constituent particles to form symmetric patterns t ...
has been solved. In PDE2, this residue is the Gln859. It has potential to form
hydrogen bonds
In chemistry, a hydrogen bond (or H-bond) is a primarily electrostatic force of attraction between a hydrogen (H) atom which is covalently bound to a more electronegative "donor" atom or group (Dn), and another electronegative atom bearing a ...
with the exocyclic
amino group
In chemistry, amines (, ) are compounds and functional groups that contain a basic nitrogen atom with a lone pair. Amines are formally derivatives of ammonia (), wherein one or more hydrogen atoms have been replaced by a substituent such ...
of cAMP and the exocyclic
carbonyl
In organic chemistry, a carbonyl group is a functional group composed of a carbon atom double-bonded to an oxygen atom: C=O. It is common to several classes of organic compounds, as part of many larger functional groups. A compound containin ...
oxygen of cGMP. In PDEs, which can
hydrolyze
Hydrolysis (; ) is any chemical reaction in which a molecule of water breaks one or more chemical bonds. The term is used broadly for substitution, elimination, and solvation reactions in which water is the nucleophile.
Biological hydrolysis ...
both cAMP and
cGMP CGMP is an initialism. It can refer to:
*cyclic guanosine monophosphate (cGMP)
*current good manufacturing practice (cGMP)
*CGMP, Cisco Group Management Protocol, the Cisco version of Internet Group Management Protocol
The Internet Group Managem ...
, this glutamine is able to rotate freely. In PDEs that are selective for either cAMP or cGMP, this glutamine is constrained by neighboring residues to a position favoring selectivity for either cyclic
nucleotide
Nucleotides are organic molecules consisting of a nucleoside and a phosphate. They serve as monomeric units of the nucleic acid polymers – deoxyribonucleic acid (DNA) and ribonucleic acid (RNA), both of which are essential biomolecul ...
.
Regulation
When cGMP binds to the
allosteric GAF-B domain of the PDE, it causes
conformational change
In biochemistry, a conformational change is a change in the shape of a macromolecule, often induced by environmental factors.
A macromolecule is usually flexible and dynamic. Its shape can change in response to changes in its environment or oth ...
in the
protein
Proteins are large biomolecules and macromolecules that comprise one or more long chains of amino acid residues. Proteins perform a vast array of functions within organisms, including catalysing metabolic reactions, DNA replication, respon ...
structure leading to higher enzyme activity. Increased hydrolysis of
cAMP due to binding of
cGMP CGMP is an initialism. It can refer to:
*cyclic guanosine monophosphate (cGMP)
*current good manufacturing practice (cGMP)
*CGMP, Cisco Group Management Protocol, the Cisco version of Internet Group Management Protocol
The Internet Group Managem ...
to the GAF-B domain is well documented, however there are no known examples for the reverse.
It has been shown that the GAF-B domain has 30-100 fold lower
affinity
Affinity may refer to:
Commerce, finance and law
* Affinity (law), kinship by marriage
* Affinity analysis, a market research and business management technique
* Affinity Credit Union, a Saskatchewan-based credit union
* Affinity Equity Part ...
for
cAMP than for
cGMP CGMP is an initialism. It can refer to:
*cyclic guanosine monophosphate (cGMP)
*current good manufacturing practice (cGMP)
*CGMP, Cisco Group Management Protocol, the Cisco version of Internet Group Management Protocol
The Internet Group Managem ...
.
This information combined with what is currently known about
intracellular
This glossary of biology terms is a list of definitions of fundamental terms and concepts used in biology, the study of life and of living organisms. It is intended as introductory material for novices; for more specific and technical definitions ...
cAMP concentrations, renders it unlikely that activation of
cGMP CGMP is an initialism. It can refer to:
*cyclic guanosine monophosphate (cGMP)
*current good manufacturing practice (cGMP)
*CGMP, Cisco Group Management Protocol, the Cisco version of Internet Group Management Protocol
The Internet Group Managem ...
hydrolysis by
cAMP can take place
in vivo
Studies that are ''in vivo'' (Latin for "within the living"; often not italicized in English) are those in which the effects of various biological entities are tested on whole, living organisms or cells, usually animals, including humans, and ...
.
Clinical value of PDE2
PDE2 is expressed in various tissues, for example:
adrenal medulla
The adrenal medulla ( la, medulla glandulae suprarenalis) is part of the adrenal gland. It is located at the center of the gland, being surrounded by the adrenal cortex. It is the innermost part of the adrenal gland, consisting of chromaffin ce ...
, brain, heart,
platelet
Platelets, also called thrombocytes (from Greek θρόμβος, "clot" and κύτος, "cell"), are a component of blood whose function (along with the coagulation factors) is to react to bleeding from blood vessel injury by clumping, thereby i ...
,
macrophages and
endothelial cells
The endothelium is a single layer of squamous endothelial cells that line the interior surface of blood vessels and lymphatic vessels. The endothelium forms an interface between circulating blood or lymph in the lumen and the rest of the ves ...
. The enzyme is thought to be involved in regulating many different
intracellular
This glossary of biology terms is a list of definitions of fundamental terms and concepts used in biology, the study of life and of living organisms. It is intended as introductory material for novices; for more specific and technical definitions ...
processes, such as:
*
aldosterone
Aldosterone is the main mineralocorticoid steroid hormone produced by the zona glomerulosa of the adrenal cortex in the adrenal gland. It is essential for sodium conservation in the kidney, salivary glands, sweat glands, and colon. It plays ...
secretion 440px
Secretion is the movement of material from one point to another, such as a secreted chemical substance from a cell or gland. In contrast, excretion is the removal of certain substances or waste products from a cell or organism. The classica ...
from the
adrenal gland
The adrenal glands (also known as suprarenal glands) are endocrine glands that produce a variety of hormones including adrenaline and the steroids aldosterone and cortisol. They are found above the kidneys. Each gland has an outer cortex w ...
* intracellular concentrations of cAMP in cardiomyocytes and of cAMP and cGMP in
platelets
Platelets, also called thrombocytes (from Greek θρόμβος, "clot" and κύτος, "cell"), are a component of blood whose function (along with the coagulation factors) is to react to bleeding from blood vessel injury by clumping, thereby i ...
* cGMP in
neurons
A neuron, neurone, or nerve cell is an electrically excitable cell that communicates with other cells via specialized connections called synapses. The neuron is the main component of nervous tissue in all animals except sponges and placozoa. ...
and effect on long-term memory
* barrier function of endothelial cells under
inflammatory
Inflammatory may refer to:
* Inflammation, a biological response to harmful stimuli
* The word ''inflammatory'' is also used to refer literally to fire and flammability, and figuratively in relation to comments that are provocative and arouse pas ...
conditions
Several enzyme functions have been reported for the PDE2. It has been shown that PDE2 lowers cAMP through increased cGMP caused by
atrial natriuretic peptide
Atrial natriuretic peptide (ANP) or atrial natriuretic factor (ANF) is a natriuretic peptide hormone secreted from the cardiac atria that in humans is encoded by the NPPA gene. Natriuretic peptides (ANP, BNP, and CNP) are a family of hormone/pa ...
(ANP) resulting in decreased
aldosterone
Aldosterone is the main mineralocorticoid steroid hormone produced by the zona glomerulosa of the adrenal cortex in the adrenal gland. It is essential for sodium conservation in the kidney, salivary glands, sweat glands, and colon. It plays ...
secretion 440px
Secretion is the movement of material from one point to another, such as a secreted chemical substance from a cell or gland. In contrast, excretion is the removal of certain substances or waste products from a cell or organism. The classica ...
.
It has also been suggested that PDE2 might play an important role in the regulation of elevated
intracellular
This glossary of biology terms is a list of definitions of fundamental terms and concepts used in biology, the study of life and of living organisms. It is intended as introductory material for novices; for more specific and technical definitions ...
concentrations of cAMP and cGMP in platelets. PDE3 is an important player in platelet aggregation. It has been reported that higher concentration of cGMP causes inhibition of
PDE3
PDE3 is a phosphodiesterase. The PDEs belong to at least eleven related gene families, which are different in their primary structure, substrate affinity, responses to effectors, and regulation mechanism.
Most of the PDE families are composed of mo ...
, whereas it stimulates PDE2. Interplay between those two functions seems to mediate an opposing regulation of cAMP in platelets.
PDE2 regulates cardiac L-type Ca
2+ current in
cardiac
The heart is a muscular organ found in most animals. This organ pumps blood through the blood vessels of the circulatory system. The pumped blood carries oxygen and nutrients to the body, while carrying metabolic waste such as carbon di ...
myocyte
A muscle cell is also known as a myocyte when referring to either a cardiac muscle cell (cardiomyocyte), or a smooth muscle cell as these are both small cells. A skeletal muscle cell is long and threadlike with many nuclei and is called a m ...
s, where activation of PDE2 by cGMP lowers cAMP and thereby affecting cardiac function.
However, it has been recently found that different cAMP pools, located within the cardiac myocyte, mediate different (moreover, sometimes opposing) effects. As different PDE types may affect different cAMP pools, the different PDEs may regulate different processes in the cell. PDE2 is expressed in several regions of the brain and rat experiments have indicated that inhibition of PDE2 enhances functions such as memory.
PDE2 is up regulated when
monocyte
Monocytes are a type of leukocyte or white blood cell. They are the largest type of leukocyte in blood and can differentiate into macrophages and conventional dendritic cells. As a part of the vertebrate innate immune system monocytes also i ...
s differentiate into
macrophages
Macrophages (abbreviated as M φ, MΦ or MP) ( el, large eaters, from Greek ''μακρός'' (') = large, ''φαγεῖν'' (') = to eat) are a type of white blood cell of the immune system that engulfs and digests pathogens, such as cancer ce ...
, but the role of PDE2 in matured macrophages is yet to be characterized. Furthermore, PDE2 has been indicated to play a role in
inflammatory
Inflammatory may refer to:
* Inflammation, a biological response to harmful stimuli
* The word ''inflammatory'' is also used to refer literally to fire and flammability, and figuratively in relation to comments that are provocative and arouse pas ...
responses as it has been detected in microvessels, but not in larger vessels. It has been speculated that
tumor necrosis factor-alpha
Tumor necrosis factor (TNF, cachexin, or cachectin; formerly known as tumor necrosis factor alpha or TNF-α) is an adipokine and a cytokine. TNF is a member of the TNF superfamily, which consists of various transmembrane proteins with a homologo ...
(TNFα) might regulate the function of PDE2 in
endothelial cells
The endothelium is a single layer of squamous endothelial cells that line the interior surface of blood vessels and lymphatic vessels. The endothelium forms an interface between circulating blood or lymph in the lumen and the rest of the ves ...
and thereby affecting flow of fluid and cells through the endothelial barrier as
in vitro
''In vitro'' (meaning in glass, or ''in the glass'') studies are performed with microorganisms, cells, or biological molecules outside their normal biological context. Colloquially called "test-tube experiments", these studies in biology and ...
experiments on endothelial cells show up regulation of both PDE2
mRNA
In molecular biology, messenger ribonucleic acid (mRNA) is a single-stranded molecule of RNA that corresponds to the genetic sequence of a gene, and is read by a ribosome in the process of synthesizing a protein.
mRNA is created during the ...
and activity.
Until now PDE2 inhibitors have mostly been used as research tools, but are presently being investigated for improving memory, for decreasing endothelial
permeability under
inflammatory
Inflammatory may refer to:
* Inflammation, a biological response to harmful stimuli
* The word ''inflammatory'' is also used to refer literally to fire and flammability, and figuratively in relation to comments that are provocative and arouse pas ...
conditions,
and for preventing/improving heart failure and cardiac hypertrophy.
PDE2A inhibitors
EHNA
The first specific inhibitor developed for PDE2 was
EHNA
EHNA (erythro-9-(2-hydroxy-3-nonyl)adenine) is a potent adenosine deaminase inhibitor, which also acts as a phosphodiesterase inhibitor
A phosphodiesterase inhibitor is a drug that blocks one or more of the five subtypes of the enzyme phosphodie ...
(''erythro''-9-(2-hydroxy-3-nonyl)adenine). It has been demonstrated to specifically act on PDE2 by inhibiting cGMP-activation of PDE2 with an IC
50 value of ~1μM and an at least 50-fold selectivity over other PDEs.
The core structure of EHNA resembles cAMP but differentiates in the fact that EHNA has a bulky
hydrophobic
In chemistry, hydrophobicity is the physical property of a molecule that is seemingly repelled from a mass of water (known as a hydrophobe). In contrast, hydrophiles are attracted to water.
Hydrophobic molecules tend to be nonpolar and, ...
carbon side chain replacing the phospho-ribose
moiety
Moiety may refer to:
Chemistry
* Moiety (chemistry), a part or functional group of a molecule
** Moiety conservation, conservation of a subgroup in a chemical species
Anthropology
* Moiety (kinship), either of two groups into which a society is ...
in cAMP.
Inhibitory effects of EHNA
In
primary culture
Cell culture or tissue culture is the process by which cells are grown under controlled conditions, generally outside of their natural environment. The term "tissue culture" was coined by American pathologist Montrose Thomas Burrows. This te ...
s of rat
cortical neurons, the inhibition of PDE2A by EHNA potentiates
NMDA
''N''-methyl--aspartic acid or ''N''-methyl--aspartate (NMDA) is an amino acid derivative that acts as a specific agonist at the NMDA receptor mimicking the action of glutamate, the neurotransmitter which normally acts at that receptor. Unlike ...
(''N''-methyl-
D-aspartate) receptor activated increase in cGMP, but has no effect on cAMP concentrations.
EHNA is also a very
potent
Potent may refer to:
* for the heraldic fur
*Warren Potent for the Australian Olympic medalist in shooting
See also:
*Potency (disambiguation)
Potency may refer to:
* Potency (pharmacology), a measure of the activity of a drug in a biologica ...
adenosine deaminase
Adenosine deaminase (also known as adenosine aminohydrolase, or ADA) is an enzyme () involved in purine metabolism. It is needed for the breakdown of adenosine from food and for the turnover of nucleic acids in tissues.
Its primary function i ...
inhibitor with an IC
50 ~2 nM.
This dual inhibition would lead to the accumulation of the two inhibitory
metabolite
In biochemistry, a metabolite is an intermediate or end product of metabolism.
The term is usually used for small molecules. Metabolites have various functions, including fuel, structure, signaling, stimulatory and inhibitory effects on enzymes, ...
s, adenosine
and cGMP,
which may act in
synergy
Synergy is an interaction or cooperation giving rise to a whole that is greater than the simple sum of its parts. The term ''synergy'' comes from the Attic Greek word συνεργία ' from ', , meaning "working together".
History
In Christi ...
to mediate diverse pharmacological responses including anti-viral, anti-tumour and anti-arrhythmic effects.
Although EHNA potently inhibits
adenosine deaminase
Adenosine deaminase (also known as adenosine aminohydrolase, or ADA) is an enzyme () involved in purine metabolism. It is needed for the breakdown of adenosine from food and for the turnover of nucleic acids in tissues.
Its primary function i ...
, it has been successfully used with the proper controls as a tool to probe PDE2 functions. EHNA has been used to study implication of PDE2 in calcium control in cardiac myocytes
and has shown to be effective to reverse hypoxic pulmonary vasoconstriction in perfused lung models.
EHNA has been therefore been used for two purposes:
# to serve as a
lead structure
Lead is a chemical element with the Symbol (chemistry), symbol Pb (from the Latin ) and atomic number 82. It is a heavy metals, heavy metal that is density, denser than most common materials. Lead is Mohs scale of mineral hardness#Intermediate ...
for the rational design of more selective and potent PDE2 inhibitors, and
# to define some of PDE's biological targets.
However, the use of EHNA as a chemical tool in determining the pharmacological role of PDE2 is limited due to its low potency to inhibit PDE2 and high potency in inhibition of adenosine deaminase.
Theoretically, this problem can be resolved if the effect of adenosine accumulated by EHNA, a result of adenosine deaminase inhibition, can be accounted and corrected for. This way, a positive inotropic effect elicited by EHNA, as a result of PDE2 inhibition, was observed.
BAY 60-7550, oxindole and PDP
BAY 60-7550 is an
analog
Analog or analogue may refer to:
Computing and electronics
* Analog signal, in which information is encoded in a continuous variable
** Analog device, an apparatus that operates on analog signals
*** Analog electronics, circuits which use analo ...
of
EHNA
EHNA (erythro-9-(2-hydroxy-3-nonyl)adenine) is a potent adenosine deaminase inhibitor, which also acts as a phosphodiesterase inhibitor
A phosphodiesterase inhibitor is a drug that blocks one or more of the five subtypes of the enzyme phosphodie ...
, which is more than 100-fold more potent and is highly selective for PDE2A.
Other newly discovered selective PDE2 inhibitors are PDP (9-(6-phenyl-2-oxohex-3-yl)-2-(3,4-dimethoxybenzyl)-purin-6-one)
and oxindole.
The table above shows the
potency
Potency may refer to:
* Potency (pharmacology), a measure of the activity of a drug in a biological system
* Virility
* Cell potency, a measure of the differentiation potential of stem cells
* In homeopathic dilutions, potency is a measure of how ...
of PDE2 inhibitors including EHNA. There is a large increase in potency between EHNA, Bay 60-7550 and PDP. The large dimethoxybenzyl group in position 2 of the purine moiety of Bay-60 7550 and PDP might be contributing to the added potency.
Structure and bonding of the inhibitors
Comparison of these inhibitors with the natural
substrate
Substrate may refer to:
Physical layers
*Substrate (biology), the natural environment in which an organism lives, or the surface or medium on which an organism grows or is attached
** Substrate (locomotion), the surface over which an organism lo ...
s of the enzyme, cAMP and cGMP reveal some common characteristics of the molecules. The main characteristic of all the molecules is the flat moiety comprising at least two fused ring structures, a six atom ring and a five atom ring. This ring system in cGMP and cAMP is a purine ring system, and the same is true for EHNA and PDP. Bay 60-7550 and oxindole lack the purine core but do possess a related ring system. Hydrogen bond acceptors, mostly nitrogen but also oxygen, reside in the ring system of the inhibitors. These atoms might interact with hydrogen bond donators, which are part of amino acids in the active site of the enzyme and thereby contribute to the inhibition of the enzyme from hydrolyzing cAMP and cGMP similar to how the natural substrates bind to the active site.
Structural similarity of the inhibitors
The structures of Bay 60-7550 and PDP are very similar. The difference between these molecules is the exocyclic
methyl group on Bay 60-7550, which replaces the nitrogen atom in PDP decreasing the possibility to form
hydrogen bond
In chemistry, a hydrogen bond (or H-bond) is a primarily electrostatic force of attraction between a hydrogen (H) atom which is covalently bound to a more electronegative "donor" atom or group (Dn), and another electronegative atom bearing ...
s with the enzyme in an important site for substrate and inhibitor binding.
The oxindole structure differs from the other inhibitors since it is more divergent from the
purine
Purine is a heterocyclic aromatic organic compound that consists of two rings ( pyrimidine and imidazole) fused together. It is water-soluble. Purine also gives its name to the wider class of molecules, purines, which include substituted purin ...
ring system and has less hydrogen binding possibilities. The molecule also lacks the large side group, analogous to the dimethoxybenzyl group of Bay 60-7550 and PDP. It is difficult to predict possible interactions to the enzyme without a co-crystal structure of the phenomenon.
Possible structure-activity relationship for PDE2 inhibitors
There is a lack of co-crystal structures of inhibitors bound in the
active site of PDE2. However, a computerized
docking model of the inhibitor EHNA and the substrates cAMP and cGMP bound in the catalytic site have been made.
The
docking model of EHNA showed that the
mutation
In biology, a mutation is an alteration in the nucleic acid sequence of the genome of an organism, virus, or extrachromosomal DNA. Viral genomes contain either DNA or RNA. Mutations result from errors during DNA or viral replication, m ...
s of the amino acids Asp811 to Ala (Asp811Ala) and Ile826 to Val (Ile826Val) at the
active site, where the only amino acid substitutions that significantly affected the inhibition by EHNA. The Asp811 mutation to alanine increased IC
50 value for EHNA 6-fold and the Ile826 mutation to valine leads to a 7-fold increased IC
50 value for EHNA compared to wild type PDE2A.
Upper binding pocket: Gln859 and Asp811
EHNA is in close proximity to Gln859 at the active site, which could donate two hydrogen bonds to N1 and N6 of the nitrogen atoms in the adenine ring of EHNA. On the other site of the binding pocket the Asp811 could donate another hydrogen bond to N7 in the adenine ring in order to stabilize the bond inhibitor. This hypothesis is supported by the fact that the Asp811Ala mutant has decreased activity toward cAMP, whereas activity toward cGMP is unchanged.
The lower binding pocket: Ile 826
The residues in the lower binding pocket may lie too far away for interaction with the inhibitor and therefore might be irrelevant for EHNA selectivity.
However the residues may play an indirect role of EHNA selectivity. Ile826 is positioned below the purine ring of EHNA and thereby limits the space for EHNA. Substitution with smaller valine (Ile826Val mutation) could increase the space for EHNA and cause the loss of hydrogen binding with residues in the upper binding pocket, while improving hydrogen binding within the lower binding pocket. This shift of interactions could destabilize binding of the adenine ring of EHNA, which could be the reason for higher IC
50 value.
No models are available for the other inhibitors than EHNA, which align in the active site. Therefore, it is more difficult to interpret the molecular binding. When looking at the inhibitors and their overall similarity, it is likely that they bind with a similar mechanism to the active site and that the different side groups determine potency of the inhibitor.
The determinants of inhibitor specificity within the PDE2 active site are not very well known and with better understanding of these determinants it would facilitate the development of inhibitors with increased potency.
References
{{Esterases
Enzymes