In biochemistry, 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:

:<math chem>\ce{A-B} \quad \xrightarrow[\text{ isomerase }]{} \quad \ce{B-A}</math>

There is only one substrate yielding one product. This product has the same molecular formula as the substrate but differs in bond connectivity or spatial arrangement. Isomerases catalyze reactions across many biological processes, such as in glycolysis and carbohydrate metabolism.

Isomerization

Isomerases catalyze changes within one molecule. They convert one isomer to another, meaning that the end product has the same molecular formula but a different physical structure. Isomers themselves exist in many varieties but can generally be classified as structural isomers or stereoisomers. Structural isomers have a different ordering of bonds and/or different bond connectivity from one another, as in the case of hexane and its four other isomeric forms (2-methylpentane, 3-methylpentane, 2,2-dimethylbutane, and 2,3-dimethylbutane).

Stereoisomers have the same ordering of individual bonds and the same connectivity but the three-dimensional arrangement of bonded atoms differ. For example, 2-butene exists in two isomeric forms: cis-2-butene and trans-2-butene. The sub-categories of isomerases containing racemases, epimerases and cis-trans isomers are examples of enzymes catalyzing the interconversion of stereoisomers. Intramolecular lyases, oxidoreductases and transferases catalyze the interconversion of structural isomers.

The prevalence of each isomer in nature depends in part on the isomerization energy, the difference in energy between isomers. Isomers close in energy can interconvert easily and are often seen in comparable proportions. The isomerization energy, for example, for converting from a stable cis isomer to the less stable trans isomer is greater than for the reverse reaction, explaining why in the absence of isomerases or an outside energy source such as ultraviolet radiation a given cis isomer tends to be present in greater amounts than the trans isomer. Isomerases can increase the reaction rate by lowering the isomerization energy.

Calculating isomerase kinetics from experimental data can be more difficult than for other enzymes because the use of product inhibition experiments is impractical. That is, isomerization is not an irreversible reaction since a reaction vessel will contain one substrate and one product so the typical simplified model for calculating reaction kinetics does not hold. There are also practical difficulties in determining the rate-determining step at high concentrations in a single isomerization. Instead, tracer perturbation can overcome these technical difficulties if there are two forms of the unbound enzyme. This technique uses isotope exchange to measure indirectly the interconversion of the free enzyme between its two forms. The radiolabeled substrate and product diffuse in a time-dependent manner. When the system reaches equilibrium the addition of unlabeled substrate perturbs or unbalances it. As equilibrium is established again, the radiolabeled substrate and product are tracked to determine energetic information.

The earliest use of this technique elucidated the kinetics and mechanism underlying the action of phosphoglucomutase, favoring the model of indirect transfer of phosphate with one intermediate and the direct transfer of glucose. This technique was then adopted to study the profile of proline racemase and its two states: the form which isomerizes L-proline and the other for D-proline. At high concentrations it was shown that the transition state in this interconversion is rate-limiting and that these enzyme forms may differ just in the protonation at the acidic and basic groups of the active site.

Classification

Enzyme-catalyzed reactions each have a uniquely assigned classification number. Isomerase-catalyzed reactions have their own EC category: EC 5. Sub-categories of this class are:

thumbnail|reaction catalyzed by phosphoribosylanthranilate isomerase

{| class="wikitable"

|+ Intramolecular oxidoreductases:

! EC number || Description || Examples

|-

| align=center | EC 5.3.1 || Interconverting Aldoses and Ketoses || Triose-phosphate isomerase, Ribose-5-phosphate isomerase

|-

| align=center | EC 5.3.2 || Interconverting Keto- and Enol-Groups || Phenylpyruvate tautomerase, Oxaloacetate tautomerase

|-

| align=center | EC 5.3.3 || Transposing C=C Double Bonds || Steroid Delta-isomerase, L-dopachrome isomerase

|-

| align=center | EC 5.3.4 || Transposing S-S Bonds || Protein disulfide-isomerase

|-

| align=center | EC 5.3.99 || Other Intramolecular Oxidoreductases || Prostaglandin-D synthase, Allene-oxide cyclase

|-

|}

Intramolecular transferases

This category (EC 5.4) includes intramolecular transferases (mutases). These isomerases catalyze the transfer of functional groups from one part of a molecule to another. This sub-class can be broken down according to the functional group the enzyme transfers:

thumbnail|reaction catalyzed by phosphoenolpyruvate mutase

{| class="wikitable"

|+ Intramolecular transferases:

! EC number || Description || Examples

|-

| align=center | EC 5.4.1 || Transferring Acyl Groups || Lysolecithin acylmutase, Precorrin-8X methylmutase

|-

| align=center | EC 5.4.2 || Phosphotransferases (Phosphomutases) || Phosphoglucomutase, Phosphopentomutase

|-

| align=center | EC 5.4.3 || Transferring Amino Groups || Beta-lysine 5,6-aminomutase, Tyrosine 2,3-aminomutase

|-

| align=center | EC 5.4.4 || Transferring hydroxy groups || (hydroxyamino)benzene mutase, Isochorismate synthase

|-

| align=center | EC 5.4.99 || Transferring Other Groups || Methylaspartate mutase, Chorismate mutase

|-

|}

Intramolecular lyases

This category (EC 5.5) includes intramolecular lyases. These enzymes catalyze "reactions in which a group can be regarded as eliminated from one part of a molecule, leaving a double bond, while remaining covalently attached to the molecule."

Epimerization

thumbnail|left|The conversion of ribulose-5-phosphate to xylulose-5-phosphate

An example of epimerization is found in the Calvin cycle when D-ribulose-5-phosphate is converted into D-xylulose-5-phosphate by ribulose-phosphate 3-epimerase. The substrate and product differ only in stereochemistry at the third carbon in the chain. The underlying mechanism involves the deprotonation of that third carbon to form a reactive enolate intermediate. The enzyme's active site contains two Asp residues. After the substrate binds to the enzyme, the first Asp deprotonates the third carbon from one side of the molecule. This leaves a planar sp<sup>2</sup>-hybridized intermediate. The second Asp is located on the opposite side of the active side and it protonates the molecule, effectively adding a proton from the back side. These coupled steps invert stereochemistry at the third carbon.

Intramolecular transfer

thumbnail|A proposed mechanism for chorismate mutase. Clark, T., Stewart, J.D. and Ganem, B. Transition-state analogue inhibitors of chlorismate mutase. Tetrahedron 46 (1990) 731–748. © IUBMB 2001

Chorismate mutase is an intramolecular transferase and it catalyzes the conversion of chorismate to prephenate, used as a precursor for L-tyrosine and L-phenylalanine in some plants and bacteria. This reaction is a Claisen rearrangement that can proceed with or without the isomerase, though the rate increases 10<sup>6</sup> fold in the presence of chorismate mutase. The reaction goes through a chair transition state with the substrate in a trans-diaxial position. Experimental evidence indicates that the isomerase selectively binds the chair transition state, though the exact mechanism of catalysis is not known. It is thought that this binding stabilizes the transition state through electrostatic effects, accounting for the dramatic increase in the reaction rate in the presence of the mutase or upon addition of a specifically-placed cation in the active site.

Intramolecular oxidoreduction

thumbnail|left|Conversion by IPP isomerase

Isopentenyl-diphosphate delta isomerase type I (also known as IPP isomerase) is seen in cholesterol synthesis and in particular it catalyzes the conversion of isopentenyl diphosphate (IPP) to dimethylallyl diphosphate (DMAPP). In this isomerization reaction a stable carbon-carbon double bond is rearranged top create a highly electrophilic allylic isomer. IPP isomerase catalyzes this reaction by the stereoselective antarafacial transposition of a single proton. The double bond is protonated at C4 to form a tertiary carbocation intermediate at C3. The adjacent carbon, C2, is deprotonated from the opposite face to yield a double bond. In effect, the double bond is shifted over.

The role of isomerase in human disease

Isomerase plays a role in human disease. Deficiencies of this enzyme can cause disorders in humans.

Phosphohexose isomerase deficiency

Phosphohexose Isomerase Deficiency (PHI) is also known as phosphoglucose isomerase deficiency or Glucose-6-phosphate isomerase deficiency, and is a hereditary enzyme deficiency. PHI is the second most frequent erthoenzyopathy in glycolysis besides pyruvate kinase deficiency, and is associated with non-spherocytic haemolytic anaemia of variable severity. PHI is the result of a dimeric enzyme that catalyses the reversible interconversion of fructose-6-phosphate and gluose-6-phosphate.

Triosephosphate isomerase deficiency

The disease referred to as triosephosphate isomerase deficiency (TPI), is a severe autosomal recessive inherited multisystem disorder of glycolytic metabolism. It is characterized by hemolytic anemia and neurodegeneration, and is caused by anaerobic metabolic dysfunction. This dysfunction results from a missense mutation that effects the encoded TPI protein. The most common mutation is the substitution of gene, Glu104Asp, which produces the most severe phenotype, and is responsible for approximately 80% of clinical TPI deficiency. It is a congenital disease that most often occurs with hemolytic anemia and manifests with jaundice.

Individuals with TPI show obvious symptoms after 6–24 months of age. These symptoms include: dystonia, tremor, dyskinesia, pyramidal tract signs, cardiomyopathy and spinal motor neuron involvement.

Supportive measures such as red cell transfusions in cases of severe anaemia can be taken to treat TPI as well. In some cases, spleen

removal (splenectomy) may improve the anaemia. There is no treatment to prevent progressive

neurological impairment of any other non-haematological clinical manifestation of the diseases.

Industrial applications

By far the most common use of isomerases in industrial applications is in sugar manufacturing. Glucose isomerase (also known as xylose isomerase) catalyzes the conversion of D-xylose and D-glucose to D-xylulose and D-fructose. Like most sugar isomerases, glucose isomerase catalyzes the interconversion of aldoses and ketoses.

The conversion of glucose to fructose is a key component of high-fructose corn syrup production. Isomerization is more specific than older chemical methods of fructose production, resulting in a higher yield of fructose and no side products.), its relatively low cost and its inability to crystallize. Fructose is also used as a sweetener for use by diabetics. The enzyme requires a divalent cation such as Co<sup>2+</sup> and Mg<sup>2+</sup> for peak activity, an additional cost to manufacturers. Glucose isomerase also has a much higher affinity for xylose than for glucose, necessitating a carefully controlled environment. The use of glucose isomerase very efficiently converts xylose to xylulose, which can then be acted upon by fermenting yeast. Overall, extensive research in genetic engineering has been invested into optimizing glucose isomerase and facilitating its recovery from industrial applications for re-use.

Glucose isomerase is able to catalyze the isomerization of a range of other sugars, including D-ribose, D-allose and L-arabinose. The most efficient substrates are those similar to glucose and xylose, having equatorial hydroxyl groups at the third and fourth carbons. The current model for the mechanism of glucose isomerase is that of a hydride shift based on X-ray crystallography and isotope exchange studies. for example isomerases with the thioredoxin domain, and certain prolyl isomerases.

References

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