Isocitrate lyase
Encyclopedia
Isocitrate lyase or ICL, is an enzyme
in the glyoxylate cycle
that catalyzes
the cleavage of isocitrate to succinate and glyoxylate. Together with malate synthase
, it bypasses the two decarboxylation steps of the tricarboxylic acid cycle (TCA cycle) and is used by bacteria, fungi, and plants.
The systematic name of this enzyme class is isocitrate glyoxylate-lyase (succinate-forming). Other names in common use include isocitrase, isocitritase, isocitratase, threo-Ds-isocitrate glyoxylate-lyase, and isocitrate glyoxylate-lyase. This enzyme participates in glyoxylate and dicarboxylate metabolism
.
s, specifically the oxo-acid-lyases, which cleave carbon-carbon bonds. Other enzymes also belong to this family including carboxyvinyl-carboxyphosphonate phosphorylmutase
which catalyses the conversion of 1-carboxyvinyl carboxyphosphonate to 3-(hydrohydroxyphosphoryl) pyruvate carbon dioxide, and phosphoenolpyruvate mutase
, which is involved in the biosynthesis of phosphinothricin tripeptide antibiotics.
During catalysis, isocitrate is deprotonated
, and an aldol cleavage
results in the release of succinate and glyoxylate. This reaction mechanism functions much like that of aldolase
in glycolysis
, where a carbon-carbon bond is cleaved and an aldehyde is released.
In the glyoxylate cycle, malate synthase then catalyzes the condensation of glyoxylate and acetyl-CoA
to form malate
so the cycle can continue.
ICL competes with isocitrate dehydrogenase
, an enzyme found in the TCA cycle, for isocitrate processing. Flux through these enzymes is controlled by phosphorylation
of isocitrate dehydrogenase, which has a much higher affinity for isocitrate as compared to ICL. Deactivation of isocitrate dehydrogenase by phosphorylation thus leads to increased isocitrate channeling through ICL, as seen when bacteria are grown on acetate
, a two-carbon compound.
have been solved for this class of enzymes, with PDB
accession codes , , , , and .
ICL is composed of four identical chains and requires a Mg2+ or Mn2+ and a thiol
for activity. In Escherichia coli
, Lys-193, Lys-194, Cys-195, His-197, and His-356 are thought to be catalytic residues, while His-184 is thought to be involved in the assembly of the tetrameric enzyme.
Between prokaryote
s and eukaryote
s, a difference in ICL structure is the addition of approximately 100 amino acids near the center of the eukaryotic enzyme. In eukaryotes, the additional amino acids are thought to function in the localization of ICL to single-membrane-bound organelle
s called glyoxysome
s. These additional amino acids account for the difference in molecular mass: the prokaryotic ICL is 48kDa, while the eukaryotic ICL is 67 kDa. Only one cysteine residue is conserved between the sequences of the fungal, plant and bacterial enzymes; it is located in the middle of a conserved hexapeptide.
, bacteria
, protist
s, plant
s, fungi, and nematodes. Although the gene has been found in genomes of nematodes and cnidaria, it has not been found in the genomes of placental mammals.
By diverting isocitrate from the TCA cycle, the actions of ICL and malate synthase in the glyoxylate cycle result in the net assimilation of carbon from 2-carbon compounds. Thus, while the TCA cycle yields no net carbon assimilation, the glyoxylate cycle generates intermediates that can be used to synthesize glucose
(via gluconeogenesis
), and other biosynthetic products. As a result, organisms that use ICL and malate synthase are able to synthesize glucose and metabolic intermediates from acetyl-CoA derived from acetate or from the degradation of ethanol, fatty acids or poly-β-hydroxybutyrate.
This function is especially important for higher plants which use oilseeds. In these germinating seeds, the breakdown of oils generates acetyl-CoA. This serves as a substrate for the glyoxylate cycle, which generates other cyclic intermediates and serves as a primary nutrient source prior to the production of sugars from photosynthesis
.
upon infection of canola. Inactivation of the icl1 gene leads to reduced pathogenicity of the fungus, which is thought to be a result of the inability of the fungus to use carbon sources provided by the plant.
Additionally, upregulation of the glyoxylate cycle has been seen for pathogens that attack humans. This is the case for fungi such as Candida albicans
, which inhabits the skin, mouth, GI tract, gut and vagina of mammals and can lead to systemic infections of immunocompromised patients; as well as for the bacterium Mycobacterium tuberculosis
, the major causative agent of tuberculosis
. In this latter case, ICL has been found to be essential for survival in the host.
Because of its use by pathogenic fungi and bacteria, specific inhibitors
are being sought for ICL and malate synthase. Although some inhibitors have already been identified, including itaconate
, itaconic anhydride, bromopyruvate
, nitropropionate, oxalate
and malate
, these are non-specific and would also inhibit other enzymes essential for host function. More research is needed to identify inhibitors that selectively target enzymes in the glyoxylate cycle.
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...
in the glyoxylate cycle
Glyoxylate cycle
The glyoxylate cycle, a variation of the Tricarboxylic Acid Cycle, is an anabolic metabolic pathway occurring in plants, bacteria, protists, fungi and several microorganisms, such as E. coli and yeast. The glyoxylate cycle centers on the conversion of acetyl-CoA to succinate for the synthesis of...
that catalyzes
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....
the cleavage of isocitrate to succinate and glyoxylate. Together with malate synthase
Malate synthase
In enzymology, a malate synthase is an enzyme that catalyzes the chemical reactionThe 3 substrates of this enzyme are acetyl-CoA, H2O, and glyoxylate, whereas its two products are -malate and CoA....
, it bypasses the two decarboxylation steps of the tricarboxylic acid cycle (TCA cycle) and is used by bacteria, fungi, and plants.
The systematic name of this enzyme class is isocitrate glyoxylate-lyase (succinate-forming). Other names in common use include isocitrase, isocitritase, isocitratase, threo-Ds-isocitrate glyoxylate-lyase, and isocitrate glyoxylate-lyase. This enzyme participates in glyoxylate and dicarboxylate metabolism
Glyoxylate and dicarboxylate metabolism
Glyoxylate and dicarboxylate metabolism describes a variety of reactions involving glyoxylate or dicarboxylates. Glyoxylate is the conjugate base of glyoxylic acid, and within a buffered environment of known pH such as the cell cytoplasm these terms can be used almost interchangeably, as the gain...
.
Mechanism
This enzyme belongs to the family of lyaseLyase
In biochemistry, a lyase is an enzyme that catalyzes the breaking of various chemical bonds by means other than hydrolysis and oxidation, often forming a new double bond or a new ring structure...
s, specifically the oxo-acid-lyases, which cleave carbon-carbon bonds. Other enzymes also belong to this family including carboxyvinyl-carboxyphosphonate phosphorylmutase
Carboxyvinyl-carboxyphosphonate phosphorylmutase
In enzymology, a carboxyvinyl-carboxyphosphonate phosphorylmutase is an enzyme that catalyzes the chemical reactionHence, this enzyme has one substrate, 1-carboxyvinyl carboxyphosphonate, and two products, 3-pyruvate and CO2....
which catalyses the conversion of 1-carboxyvinyl carboxyphosphonate to 3-(hydrohydroxyphosphoryl) pyruvate carbon dioxide, and phosphoenolpyruvate mutase
Phosphoenolpyruvate mutase
In enzymology, a phosphoenolpyruvate mutase is an enzyme that catalyzes the chemical reactionHence, this enzyme has one substrate, phosphoenolpyruvate , and one product, 3-phosphonopyruvate , which are structural isomers....
, which is involved in the biosynthesis of phosphinothricin tripeptide antibiotics.
During catalysis, isocitrate 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 an aldol cleavage
Aldol reaction
The aldol reaction is a powerful means of forming carbon–carbon bonds in organic chemistry.Discovered independently by Charles-Adolphe Wurtz and Alexander Porfyrevich Borodin in 1872, the reaction combines two carbonyl compounds to form a new β-hydroxy carbonyl compound...
results in the release of succinate and glyoxylate. This reaction mechanism functions much like that of aldolase
Aldolase
Aldolase A is an enzyme that catalyses a reverse aldol reaction: The substrate, fructose 1,6-bisphosphate is broken down into glyceraldehyde 3-phosphate and dihydroxyacetone phosphate . This reaction is a part of glycolysis. Three aldolase isozymes , encoded by three different genes, are...
in glycolysis
Glycolysis
Glycolysis is the metabolic pathway that converts glucose C6H12O6, into pyruvate, CH3COCOO− + H+...
, where a carbon-carbon bond is cleaved and an aldehyde is released.
In the glyoxylate cycle, malate synthase then catalyzes the condensation of glyoxylate and acetyl-CoA
Acetyl-CoA
Acetyl coenzyme A or acetyl-CoA is an important molecule in metabolism, used in many biochemical reactions. Its main function is to convey the carbon atoms within the acetyl group to the citric acid cycle to be oxidized for energy production. In chemical structure, acetyl-CoA is the thioester...
to form malate
Malate
Malate is the ionized form of malic acid. It is an important chemical compound in biochemistry. In the C4 carbon fixation process, malate is a source of CO2 in the Calvin cycle....
so the cycle can continue.
ICL competes with isocitrate dehydrogenase
Isocitrate dehydrogenase
Isocitrate dehydrogenase and , also known as IDH, is an enzyme that participates in the citric acid cycle. It catalyzes the third step of the cycle: the oxidative decarboxylation of isocitrate, producing alpha-ketoglutarate and CO2 while converting NAD+ to NADH...
, an enzyme found in the TCA cycle, for isocitrate processing. Flux through these enzymes is controlled by phosphorylation
Phosphorylation
Phosphorylation is the addition of a phosphate group to a protein or other organic molecule. Phosphorylation activates or deactivates many protein enzymes....
of isocitrate dehydrogenase, which has a much higher affinity for isocitrate as compared to ICL. Deactivation of isocitrate dehydrogenase by phosphorylation thus leads to increased isocitrate channeling through ICL, as seen when bacteria are grown on acetate
Acetate
An acetate is a derivative of acetic acid. This term includes salts and esters, as well as the anion found in solution. Most of the approximately 5 billion kilograms of acetic acid produced annually in industry are used in the production of acetates, which usually take the form of polymers. In...
, a two-carbon compound.
Enzyme Structure
As of late 2007, 5 structuresTertiary structure
In biochemistry and molecular biology, the tertiary structure of a protein or any other macromolecule is its three-dimensional structure, as defined by the atomic coordinates.-Relationship to primary structure:...
have been solved for this class of enzymes, with PDB
Protein Data Bank
The Protein Data Bank is a repository for the 3-D structural data of large biological molecules, such as proteins and nucleic acids....
accession codes , , , , and .
ICL is composed of four identical chains and requires a Mg2+ or Mn2+ and a thiol
Thiol
In organic chemistry, a thiol is an organosulfur compound that contains a carbon-bonded sulfhydryl group...
for activity. In Escherichia coli
Escherichia coli
Escherichia coli is a Gram-negative, rod-shaped bacterium that is commonly found in the lower intestine of warm-blooded organisms . Most E. coli strains are harmless, but some serotypes can cause serious food poisoning in humans, and are occasionally responsible for product recalls...
, Lys-193, Lys-194, Cys-195, His-197, and His-356 are thought to be catalytic residues, while His-184 is thought to be involved in the assembly of the tetrameric enzyme.
Between prokaryote
Prokaryote
The prokaryotes are a group of organisms that lack a cell nucleus , or any other membrane-bound organelles. The organisms that have a cell nucleus are called eukaryotes. Most prokaryotes are unicellular, but a few such as myxobacteria have multicellular stages in their life cycles...
s and eukaryote
Eukaryote
A eukaryote is an organism whose cells contain complex structures enclosed within membranes. Eukaryotes may more formally be referred to as the taxon Eukarya or Eukaryota. The defining membrane-bound structure that sets eukaryotic cells apart from prokaryotic cells is the nucleus, or nuclear...
s, a difference in ICL structure is the addition of approximately 100 amino acids near the center of the eukaryotic enzyme. In eukaryotes, the additional amino acids are thought to function in the localization of ICL to single-membrane-bound organelle
Organelle
In cell biology, an organelle is a specialized subunit within a cell that has a specific function, and is usually separately enclosed within its own lipid bilayer....
s called glyoxysome
Glyoxysome
Glyoxysomes are specialized peroxisomes found in plants and also in filamentous fungi....
s. These additional amino acids account for the difference in molecular mass: the prokaryotic ICL is 48kDa, while the eukaryotic ICL is 67 kDa. Only one cysteine residue is conserved between the sequences of the fungal, plant and bacterial enzymes; it is located in the middle of a conserved hexapeptide.
Biological Function
The ICL enzyme has been found to be functional in various archaeaArchaea
The Archaea are a group of single-celled microorganisms. A single individual or species from this domain is called an archaeon...
, bacteria
Bacteria
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...
, protist
Protist
Protists are a diverse group of eukaryotic microorganisms. Historically, protists were treated as the kingdom Protista, which includes mostly unicellular organisms that do not fit into the other kingdoms, but this group is contested in modern taxonomy...
s, plant
Plant
Plants are living organisms belonging to the kingdom Plantae. Precise definitions of the kingdom vary, but as the term is used here, plants include familiar organisms such as trees, flowers, herbs, bushes, grasses, vines, ferns, mosses, and green algae. The group is also called green plants or...
s, fungi, and nematodes. Although the gene has been found in genomes of nematodes and cnidaria, it has not been found in the genomes of placental mammals.
By diverting isocitrate from the TCA cycle, the actions of ICL and malate synthase in the glyoxylate cycle result in the net assimilation of carbon from 2-carbon compounds. Thus, while the TCA cycle yields no net carbon assimilation, the glyoxylate cycle generates intermediates that can be used to synthesize glucose
Glucose
Glucose is a simple sugar and an important carbohydrate in biology. Cells use it as the primary source of energy and a metabolic intermediate...
(via gluconeogenesis
Gluconeogenesis
Gluconeogenesis is a metabolic pathway that results in the generation of glucose from non-carbohydrate carbon substrates such as lactate, glycerol, and glucogenic amino acids....
), and other biosynthetic products. As a result, organisms that use ICL and malate synthase are able to synthesize glucose and metabolic intermediates from acetyl-CoA derived from acetate or from the degradation of ethanol, fatty acids or poly-β-hydroxybutyrate.
This function is especially important for higher plants which use oilseeds. In these germinating seeds, the breakdown of oils generates acetyl-CoA. This serves as a substrate for the glyoxylate cycle, which generates other cyclic intermediates and serves as a primary nutrient source prior to the production of sugars from photosynthesis
Photosynthesis
Photosynthesis is a chemical process that converts carbon dioxide into organic compounds, especially sugars, using the energy from sunlight. Photosynthesis occurs in plants, algae, and many species of bacteria, but not in archaea. Photosynthetic organisms are called photoautotrophs, since they can...
.
Disease Relevance
ICL has found to be important in human, animal, and plant pathogenesis. For several agricultural crops including cereals, cucumbers, and melons, increased expression of the gene encoding ICL is important for fungal virulence. For instance, increased gene expression of icl1 has been seen in the fungus Leptosphaeria maculansLeptosphaeria maculans
Leptosphaeria maculans is a fungal pathogen that is the causal agent of blackleg disease on Brassica crops. The major yield loss is due to stem canker....
upon infection of canola. Inactivation of the icl1 gene leads to reduced pathogenicity of the fungus, which is thought to be a result of the inability of the fungus to use carbon sources provided by the plant.
Additionally, upregulation of the glyoxylate cycle has been seen for pathogens that attack humans. This is the case for fungi such as Candida albicans
Candida albicans
Candida albicans is a diploid fungus that grows both as yeast and filamentous cells and a causal agent of opportunistic oral and genital infections in humans. Systemic fungal infections including those by C...
, which inhabits the skin, mouth, GI tract, gut and vagina of mammals and can lead to systemic infections of immunocompromised patients; as well as for the bacterium Mycobacterium tuberculosis
Mycobacterium tuberculosis
Mycobacterium tuberculosis is a pathogenic bacterial species in the genus Mycobacterium and the causative agent of most cases of tuberculosis . First discovered in 1882 by Robert Koch, M...
, the major causative agent of tuberculosis
Tuberculosis
Tuberculosis, MTB, or TB is a common, and in many cases lethal, infectious disease caused by various strains of mycobacteria, usually Mycobacterium tuberculosis. Tuberculosis usually attacks the lungs but can also affect other parts of the body...
. In this latter case, ICL has been found to be essential for survival in the host.
Because of its use by pathogenic fungi and bacteria, specific 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...
are being sought for ICL and malate synthase. Although some inhibitors have already been identified, including itaconate
Itaconic acid
Itaconic acid, or methylenesuccinic acid, is an organic compound. Itaconic acid is a white crystalline powder. Itaconic acid is a naturally occurring compound, non-toxic, and readily biodegradable. The name itaconic was devised as an anagram of aconitic.Historically, itaconic acid was obtained by...
, itaconic anhydride, bromopyruvate
Bromopyruvic acid
Bromopyruvic acid, or bromopyruvate, is a synthetic brominated derivative of pyruvic acid. It is being studied as a potential treatment for certain types of cancer...
, nitropropionate, oxalate
Oxalate
Oxalate , is the dianion with formula C2O42− also written 22−. Either name is often used for derivatives, such as disodium oxalate, 2C2O42−, or an ester of oxalic acid Oxalate (IUPAC: ethanedioate), is the dianion with formula C2O42− also written (COO)22−. Either...
and malate
Malate
Malate is the ionized form of malic acid. It is an important chemical compound in biochemistry. In the C4 carbon fixation process, malate is a source of CO2 in the Calvin cycle....
, these are non-specific and would also inhibit other enzymes essential for host function. More research is needed to identify inhibitors that selectively target enzymes in the glyoxylate cycle.