Enoyl-CoA-(∆) isomerase (, also known as dodecenoyl-CoA-(∆) isomerase, 3,2-trans-enoyl-CoA isomerase, ∆3(cis),∆2(trans)-enoyl-CoA isomerase, or acetylene-allene isomerase, is an

enzyme Enzymes () are proteins that act as biological catalysts by accelerating chemical reactions. The molecules upon which enzymes may act are called substrates, and the enzyme converts the substrates into different molecules known as products ...

that catalyzes the conversion of cis- or trans-

double bond In chemistry, a double bond is a covalent bond between two atoms involving four bonding electrons as opposed to two in a single bond. Double bonds occur most commonly between two carbon atoms, for example in alkenes. Many double bonds exist betwee ...

s of coenzyme A (CoA) bound

fatty acids In chemistry, particularly in biochemistry, a fatty acid is a carboxylic acid with an aliphatic chain, which is either saturated and unsaturated compounds#Organic chemistry, saturated or unsaturated. Most naturally occurring fatty acids have an B ...

at gamma-

carbon Carbon () is a chemical element with the symbol C and atomic number 6. It is nonmetallic and tetravalent—its atom making four electrons available to form covalent chemical bonds. It belongs to group 14 of the periodic table. Carbon mak ...

(position 3) to trans

s at beta-

(position 2) as below: This enzyme has an important role in the

metabolism Metabolism (, from el, μεταβολή ''metabolē'', "change") is the set of life-sustaining chemical reactions in organisms. The three main functions of metabolism are: the conversion of the energy in food to energy available to run c ...

unsaturated fatty acid In chemistry, particularly in biochemistry, a fatty acid is a carboxylic acid with an aliphatic chain, which is either saturated or unsaturated. Most naturally occurring fatty acids have an unbranched chain of an even number of carbon atoms, ...

s in

beta oxidation In biochemistry and metabolism, beta-oxidation is the catabolic process by which fatty acid molecules are broken down in the cytosol in prokaryotes and in the mitochondria in eukaryotes to generate acetyl-CoA, which enters the citric acid cyc ...

Mechanism

Enoyl-CoA

isomerase Isomerases are a general class of enzymes that convert a molecule from one isomer to another. Isomerases facilitate intramolecular rearrangements in which bonds are broken and formed. The general form of such a reaction is as follows: A–B � ...

is involved in the beta-oxidation, one of the most frequently used pathways in

fatty acid degradation Fatty acid degradation is the process in which fatty acids are broken down into their metabolites, in the end generating acetyl-CoA, the entry molecule for the citric acid cycle, the main energy supply of living organisms, including bacteria and ani ...

, of

s with

s at odd-numbered

positions. It does so by shifting the position of the

s in the acyl-CoA intermediates and converting 3-cis or trans-enoyl-CoA to 2-trans-enoyl-CoA. Since the key step in the degradation of

with

s at even-numbered

positions also produces 3-trans-enoyl-CoA in mammals and

yeast Yeasts are eukaryotic, single-celled microorganisms classified as members of the fungus kingdom. The first yeast originated hundreds of millions of years ago, and at least 1,500 species are currently recognized. They are estimated to constit ...

s, enoyl-CoA

is technically required for their

as well. The reaction mechanism is detailed in figure 1, and the base that initiates the

isomerization In chemistry, isomerization or isomerisation is the process in which a molecule, polyatomic ion or molecular fragment is transformed into an isomer with a different chemical structure. Enolization is an example of isomerization, as is tautomeriz ...

and NH groups that stabilize the intermediate are located on the active site of enoyl-coA

. As it functions in the step immediately preceding the actual beta-oxidation and forms a

extending from the beta-

(position 2), enoyl-CoA

is involved in both the NADPH-dependent and NADPH-independent pathways of beta-oxidation. The

serves as the target of

oxidation Redox (reduction–oxidation, , ) is a type of chemical reaction in which the oxidation states of substrate change. Oxidation is the loss of electrons or an increase in the oxidation state, while reduction is the gain of electrons or a ...

and

-to-

bond cleavage, thereby shortening the

fatty acid In chemistry, particularly in biochemistry, a fatty acid is a carboxylic acid with an aliphatic chain, which is either saturated or unsaturated. Most naturally occurring fatty acids have an unbranched chain of an even number of carbon atoms, ...

chain.

Sub-classification

Enoyl-CoA isomerases can be categorized into three classes: *monofunctional mitochondrial *monofunctional

peroxisomal A peroxisome () is a membrane-bound organelle, a type of microbody, found in the cytoplasm of virtually all eukaryotic cells. Peroxisomes are oxidative organelles. Frequently, molecular oxygen serves as a co-substrate, from which hydrogen per ...

*multifunctional The monofunctional mitochondrial and

s are found in the mitochondria and peroxisomes of eukaryotes, respectively. The multifunctional

s are found in

bacteria Bacteria (; singular: bacterium) are ubiquitous, mostly free-living organisms often consisting of one Cell (biology), biological cell. They constitute a large domain (biology), domain of prokaryotic microorganisms. Typically a few micrometr ...

and in the peroxisomes of some eukaryotes, but they serve two functions: the N-terminal domain works the same as the other classes of enoyl-CoA

s and the

C-terminal The C-terminus (also known as the carboxyl-terminus, carboxy-terminus, C-terminal tail, C-terminal end, or COOH-terminus) is the end of an amino acid chain (protein or polypeptide), terminated by a free carboxyl group (-COOH). When the protein is ...

domain works as a

dehydrogenase A dehydrogenase is an enzyme belonging to the group of oxidoreductases that oxidizes a substrate by reducing an electron acceptor, usually NAD+/NADP+ or a flavin coenzyme such as FAD or FMN. Like all catalysts, they catalyze reverse as well as ...

, specifically, to 3-hydroxyactyl-CoA. There are two divisions among the mitochondrial enoyl Co-A isomerase: short-chain and long-chain In an immunoblot, antibodies were run against all enoyl CoA isomerase. However, two of these

s had

antibody An antibody (Ab), also known as an immunoglobulin (Ig), is a large, Y-shaped protein used by the immune system to identify and neutralize foreign objects such as pathogenic bacteria and viruses. The antibody recognizes a unique molecule of the ...

attachment: the short chain isomerase and the peroxisomal multifunctional enzyme. There was one

which did not have binding specificity to this

: mitochondrial long-chain isomerase. Long-chain isomerase was found when it eluted at a lower potassium phosphate

concentration In chemistry, concentration is the abundance of a constituent divided by the total volume of a mixture. Several types of mathematical description can be distinguished: '' mass concentration'', ''molar concentration'', '' number concentration'', ...

in the gradient. Thus, the discovery of three sub-classes of enoyl CoA isomerase was made. Although all three classes of

s have the same function, there is little overlap among 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 a ...

sequences. For example, only 40 out of 302

sequences (13%) are the same between monofunctional

and mitochondrial

s in

human Humans (''Homo sapiens'') are the most abundant and widespread species of primate, characterized by bipedalism and exceptional cognitive skills due to a large and complex brain. This has enabled the development of advanced tools, cultu ...

s. In fact, in mammals, the

has an extra N-terminal domain that is not present in the mitochondrial counterpart. Also, it has been found to be a subunit of the peroxisomal trifunctional

(pTFE) and contributes only to minor cleavages of the

chain. In that sense, for many higher organisms, the mitochondrial

is essential for deriving maximum

energy In physics, energy (from Ancient Greek: ἐνέργεια, ''enérgeia'', “activity”) is the quantitative property that is transferred to a body or to a physical system, recognizable in the performance of work and in the form of hea ...

from

lipid Lipids are a broad group of naturally-occurring molecules which includes fats, waxes, sterols, fat-soluble vitamins (such as vitamins A, D, E and K), monoglycerides, diglycerides, phospholipids, and others. The functions of lipids includ ...

s and fueling muscles. Mitochondria (both short- and long-chain) of rat

liver The liver is a major organ only found in vertebrates which performs many essential biological functions such as detoxification of the organism, and the synthesis of proteins and biochemicals necessary for digestion and growth. In humans, it ...

contain more than one enoyl Co-A isomerase. To further support the idea that short- and long-chain isomerases elute at different concentration of potassium phosphate concentration, they do not share similar primary polypeptide structure, hence they must not be evolutionarily related. Peroxisomes of

plants Plants are predominantly photosynthetic eukaryotes of the kingdom Plantae. Historically, the plant kingdom encompassed all living things that were not animals, and included algae and fungi; however, all current definitions of Plantae exclude ...

and of rat

are very different in the way they operate. Despite their primary structure similarities, there are differences among the different specimen. To begin with, the peroxisomes of rat

are a multifunctional

including enoyl-CoA isomerase, enoyl-CoA hydratase, and L-(−)-3-hydroxyacyl-CoA

. Three different

s reside on this entity (multifunctional protein) allowing this

to perform isomerization, hydration, and dehydration. Isomerase activity on the multifunctional

occurs at the amino-terminal

catalytic Catalysis () is the process of increasing the rate of a chemical reaction by adding a substance known as a catalyst (). Catalysts are not consumed in the reaction and remain unchanged after it. If the reaction is rapid and the catalyst recyc ...

half of the protein along with the hydratase activity. The

activity of enoyl-CoA occurs in the carboxyl-terminal. Upon further investigation of the CoA

binding site In biochemistry and molecular biology, a binding site is a region on a macromolecule such as a protein that binds to another molecule with specificity. The binding partner of the macromolecule is often referred to as a ligand. Ligands may includ ...

on the amino-terminal half of the multifunctional

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, res ...

, the CoA substrate is not transferred through the aqueous phase from the isomerization phase to the site of hydration or does not have a bulk phase. This removes the need for a substrate transferring

. On the other hand, the

cotyledon A cotyledon (; ; ; , gen. (), ) is a significant part of the embryo within the seed of a plant, and is defined as "the embryonic leaf in seed-bearing plants, one or more of which are the first to appear from a germinating seed." The num ...

s convert long-chain 3-trans-enoyl-CoA, long-chain 3-cis-enoyl-CoA, and short-chain 3-cis-enoyl-CoA species into their 2- trans-enoyl-CoA respective forms. As previously mentioned,

plant Plants are predominantly photosynthetic eukaryotes of the kingdom Plantae. Historically, the plant kingdom encompassed all living things that were not animals, and included algae and fungi; however, all current definitions of Plantae exclu ...

enoyl-CoA isomerase exclusively forms the 2-trans isomer as

product Product may refer to: Business * Product (business), an item that serves as a solution to a specific consumer problem. * Product (project management), a deliverable or set of deliverables that contribute to a business solution Mathematics * Produ ...

. It does not act on 4-cis-enoyl-CoA species or 2-trans- 4-trans-dienoyl-CoA species. In comparing the products of the plant peroxisome and the multifunctional enzyme of rat

, the plant has no hydratase activity. The Plant form did not form a 2-cis-isomer (from enoyl-CoA hydratase) or D- or L- 3 hydroxy derivative (L-(−)-3-hydroxyacyl-CoA dehydrogenase): products of multifunctional enzyme of rat

. The turnover rates of these the two sub divisions of peroxisomes are very different. The Kcat/Km ratio in

s is 10^6 M-1s-1 which outperforms the ratio .07 * 10^6 M-1s-1. Due to a high turnover rate, the plant peroxisomes contain a lesser amount of enoyl-CoA isomerase than their counterparts in the rat

. In rat liver, mitochondrial enoyl CoA isomerase and peroxisomal enoyl CoA isomerase embedded in the multifunctional enzyme have similarities in the primary structure sequence. When comparing the amino-terminal half of E. coli against the amino-terminal half of rat

, there were primary and secondary structure similarities towards the middle of the amino-terminal end. This conserved region must be important for structure and function of this specific enzyme since showing equally in both E. coli and rat

Structure

All classes of enoyl-CoA

s belong to a family of

s, the hydratase/

or crotonase superfamily, and when examined with

x-ray crystallography X-ray crystallography is the experimental science determining the atomic and molecular structure of a crystal, in which the crystalline structure causes a beam of incident X-rays to diffract into many specific directions. By measuring the angles ...

, exhibit a common structural feature of the family, the N-terminal core with a spiral fold composed of four turns, each turn consisting of two

beta-sheet The beta sheet, (β-sheet) (also β-pleated sheet) is a common motif of the regular protein secondary structure. Beta sheets consist of beta strands (β-strands) connected laterally by at least two or three backbone hydrogen bonds, forming a ge ...

s and one alpha-helix. In enoyl-CoA

, the two

s are part of the

catalytic site 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 ...

, since the NH groups of residues following the

s attach to the

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 containi ...

oxygen Oxygen is the chemical element with the symbol O and atomic number 8. It is a member of the chalcogen group in the periodic table, a highly reactive nonmetal, and an oxidizing agent that readily forms oxides with most elements as ...

of the acyl-CoA intermediate. The formation of this

oxyanion hole An oxyanion hole is a pocket in the active site of an enzyme that stabilizes transition state negative charge on a deprotonated oxygen or alkoxide. The pocket typically consists of backbone amides or positively charged residues. Stabilising the t ...

stabilizes the

transition state In chemistry, the transition state of a chemical reaction is a particular configuration along the reaction coordinate. It is defined as the state corresponding to the highest potential energy along this reaction coordinate. It is often marked ...

of the

-catalyzed reaction. Moreover, a glutamate

residue Residue may refer to: Chemistry and biology * An amino acid, within a peptide chain * Crop residue, materials left after agricultural processes * Pesticide residue, refers to the pesticides that may remain on or in food after they are applied ...

located next to body cavities filled with water molecules and lined with hydrophobic or apolar side chains has also been identified as a part of the

. In its deprotonated form, the glutamate can act as a base and remove a proton from the acyl-CoA intermediate. The body cavities aid in rearranging the glutamate side chain to retain the proton and later deliver it back to the acyl-CoA, on a different

position. The NH-containing residues have been identified as Ala70 and Leu126 and the glutamate as Glu158 in

s in a

species, ''Saccharomyces cerevisiae''. Their relative locations on the enzyme can be compared in figure 2. The

s of the hydratase/

or crotonase superfamily are typically trimer (chemistry), trimeric disks dimer (chemistry), dimerized into hexamers. The wide range of their substrate-

specificity derives from the variations in the distances between the trimer (chemistry), trimeric disks and their orientation. However, the

mitochondrial enoyl-CoA

is a trimer (chemistry), trimer and orients the

tail in a completely different direction from that seen in the hexamers. The trimer (chemistry), trimeric disk of

enzymes in ''Saccharomyces cerevisiae'' is displayed in figure 3.

History

Enoyl-CoA

was first identified and purified from rat

mitochondria in the 1960s and 1970s via gel filtration and ion exchange chromatography. Since then, all classes of enoyl-CoA

, mitochondrial,

and multifunctional, have been identified in different organisms, including more mammals,

s, and unicellular organisms. By 1994, using the rat enoyl-CoA

cDNA as a hybridization probe,

enoyl-CoA

cDNA could be sequenced and cloned. In the same year, the protein itself was isolated, not by Chemistry, affinity to rat

or cDNA Atom probe, probes, but by coList of purification methods in chemistry, purification with a transferase, human glutathione S-transferases. In the attempts to examine the

enoyl-CoA

in detail, the mitochondrial

in the mammalian liver was identified as a potential biological marker for metabolic diseases due to its elevated levels in defective cell (biology), cells, and linked defects in

beta-oxidation to

diseases, to be specified in the next section.

Clinical significance

s, defects in the beta-oxidation mechanism result in hypoketotic hyperglycemia, a symptom of starvation, due to the inefficient utilization of

s as a primary source of

. The metabolic disease was found to be on a genetics, genetic level: rats without the genes for enoyl-CoA

also displayed high blood glucose level. Moreover, a biological marker for this condition may have been identified as the urine of these rats included high concentrations of medium chain Saturated and unsaturated compounds, unsaturated dicarboxylic acids, a condition called dicarboxylic aciduria. More recent studies link hepatitis C virus (HCV) infection to defects in

, specifically, to that in enoyl-CoA

. Hepatitis C virus, HCV is the leading cause of chronic hepatitis, cirrhosis, and liver cancer, and more than 180 million people are affected globally. Due to the prolonged latency period, latency of the virus and no existing cures to rid the virus specifically, Hepatitis C virus, HCV is a serious problem that is causing more deaths than HIV/AIDS in the United States, but its threat still do not receive adequate attention. The need for a Hepatitis C virus, HCV-specific treatment is essential, and according to John Ward, the director of the Centers for Disease Control and Prevention, CDC Hepatitis Division, it can save up to 120,000 lives. According to

profiling in the

biopsies of Hepatitis C virus, HCV patients, a correlation was initially discovered between dysfunctional mitochondrial processes, which include beta-oxidation, and Hepatitis C virus, HCV. As a matter of fact,

s play an important role in the DNA replication, replication cycle of Hepatitis C virus, HCV, and in the "in vivo" samples from Hepatitis C virus, HCV patients, many

s were found in abundance to aid Hepatitis C virus, HCV in virus uptake, RNA replication, and secretion from host cells. Enzymes that regulate fatty acid metabolism, including enoyl-CoA

, were also similarly upregulated. Gene silencing techniques revealed that enoyl-CoA

is essential in Hepatitis C virus, HCV RNA replication, and opened ways to stop Hepatitis C virus, HCV infection on an intracellular level.

References