Aconitase
Overview
 
Aconitase is an enzyme that catalyses the stereo-specific
Stereochemistry
Stereochemistry, a subdiscipline of chemistry, involves the study of the relative spatial arrangement of atoms within molecules. An important branch of stereochemistry is the study of chiral molecules....

 isomerization of citrate
Citrate
A citrate can refer either to the conjugate base of citric acid, , or to the esters of citric acid. An example of the former, a salt is trisodium citrate; an ester is triethyl citrate.-Other citric acid ions:...

 to isocitrate via cis-aconitate in the tricarboxylic acid cycle, a non-redox
Redox
Redox reactions describe all chemical reactions in which atoms have their oxidation state changed....

-active process.
In contrast with the majority of iron-sulfur protein
Iron-sulfur protein
Iron-sulfur proteins are proteins characterized by the presence of iron-sulfur clusters containing sulfide-linked di-, tri-, and tetrairon centers in variable oxidation states...

s that function as electron carriers, the iron-sulfur cluster
Iron-sulfur cluster
For biological Fe-S clusters, see iron-sulfur proteins.Iron-sulfur clusters are ensembles of iron and sulfide centres. Fe-S clusters are most often discussed in the context of the biological role for iron-sulfur proteins. Many Fe-S clusters are known in the area of organometallic chemistry and as...

 of aconitase reacts directly with an enzyme substrate. Aconitase has an active [Fe4S4]2+ cluster, which may convert to an inactive [Fe3S4]+ form.
Encyclopedia
Aconitase is an enzyme that catalyses the stereo-specific
Stereochemistry
Stereochemistry, a subdiscipline of chemistry, involves the study of the relative spatial arrangement of atoms within molecules. An important branch of stereochemistry is the study of chiral molecules....

 isomerization of citrate
Citrate
A citrate can refer either to the conjugate base of citric acid, , or to the esters of citric acid. An example of the former, a salt is trisodium citrate; an ester is triethyl citrate.-Other citric acid ions:...

 to isocitrate via cis-aconitate in the tricarboxylic acid cycle, a non-redox
Redox
Redox reactions describe all chemical reactions in which atoms have their oxidation state changed....

-active process.

Function

In contrast with the majority of iron-sulfur protein
Iron-sulfur protein
Iron-sulfur proteins are proteins characterized by the presence of iron-sulfur clusters containing sulfide-linked di-, tri-, and tetrairon centers in variable oxidation states...

s that function as electron carriers, the iron-sulfur cluster
Iron-sulfur cluster
For biological Fe-S clusters, see iron-sulfur proteins.Iron-sulfur clusters are ensembles of iron and sulfide centres. Fe-S clusters are most often discussed in the context of the biological role for iron-sulfur proteins. Many Fe-S clusters are known in the area of organometallic chemistry and as...

 of aconitase reacts directly with an enzyme substrate. Aconitase has an active [Fe4S4]2+ cluster, which may convert to an inactive [Fe3S4]+ form. Three cysteine
Cysteine
Cysteine is an α-amino acid with the chemical formula HO2CCHCH2SH. It is a non-essential amino acid, which means that it is biosynthesized in humans. Its codons are UGU and UGC. The side chain on cysteine is thiol, which is polar and thus cysteine is usually classified as a hydrophilic amino acid...

 (Cys) residues have been shown to be ligands of the [Fe4S4] centre. In the active state, the labile iron
Iron
Iron is a chemical element with the symbol Fe and atomic number 26. It is a metal in the first transition series. It is the most common element forming the planet Earth as a whole, forming much of Earth's outer and inner core. It is the fourth most common element in the Earth's crust...

 ion of the [Fe4S4] cluster is not coordinated by Cys but by water molecules.

The iron-responsive element-binding protein (IRE-BP) and 3-isopropylmalate dehydratase
3-isopropylmalate dehydratase
3-Isopropylmalate dehydratase is an aconitase homologue....

 (α-isopropylmalate isomerase; ), an enzyme catalysing the second step in the biosynthesis of leucine
Leucine
Leucine is a branched-chain α-amino acid with the chemical formula HO2CCHCH2CH2. Leucine is classified as a hydrophobic amino acid due to its aliphatic isobutyl side chain. It is encoded by six codons and is a major component of the subunits in ferritin, astacin and other 'buffer' proteins...

, are known aconitase homologues. Iron regulatory elements (IREs) constitute a family of 28-nucleotide, non-coding, stem-loop structures that regulate iron storage, 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...

 synthesis and iron uptake. They also participate in ribosome
Ribosome
A ribosome is a component of cells that assembles the twenty specific amino acid molecules to form the particular protein molecule determined by the nucleotide sequence of an RNA molecule....

 binding and control the mRNA turnover (degradation). The specific regulator protein, the IRE-BP, binds to IREs in both 5' and 3' regions, but only to RNA in the apo form, without the Fe-S cluster. Expression of IRE-BP in cultured cells has revealed that the protein functions either as an active aconitase, when cells are iron-replete, or as an active RNA-binding protein, when cells are iron-depleted. Mutant IRE-BPs, in which any or all of the three Cys residues involved in Fe-S formation are replaced by serine
Serine
Serine is an amino acid with the formula HO2CCHCH2OH. It is one of the proteinogenic amino acids. By virtue of the hydroxyl group, serine is classified as a polar amino acid.-Occurrence and biosynthesis:...

, have no aconitase activity, but retain RNA-binding properties.

Aconitase is inhibited by fluoroacetate, therefore fluoroacetate is poisonous. The iron sulfur cluster is highly sensitive to oxidation by superoxide
Superoxide
A superoxide, also known by the obsolete name hyperoxide, is a compound that possesses the superoxide anion with the chemical formula O2−. The systematic name of the anion is dioxide. It is important as the product of the one-electron reduction of dioxygen O2, which occurs widely in nature...

.

Enzyme Structure

Aconitase, displayed in the structures in the right margin of this page, has two slightly different structures, depending on whether it is activated or inactivated. In the inactive form, its structure is divided into four domains. Counting from the N-terminus, only the first three of these domains are involved in close interactions with the [3Fe-4S] cluster, but the 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...

 consists of residues from all four domains, including the larger C-terminal domain. The Fe-S cluster and a SO42- anion also reside in the active site. When the enzyme is activated, it gains an additional iron atom, creating a [4Fe-4S] cluster. However, the structure of the rest of the enzyme is nearly unchanged; the conserved atoms between the two forms are in essentially the same positions, up to a difference of 0.1 angstroms.

Enzyme Mechanism

Aconitase employs a dehydration-hydration mechanism. The catalytic residues involved are His-101 and Ser-642. His-101 protonates the hydroxyl group on C3 of citrate, allowing it to leave as water, and Ser-642 concurrently abstracts the proton on C2, forming a double bond between C2 and C3, forming a cis-aconitate intermediate. At this point, the intermediate is rotated 180º. This rotation is referred to as a "flip." Because of this flip, the intermediate is said to move from a "citrate mode" to a "isocitrate mode."

How exactly this flip occurs is debatable. One theory is that, in the rate-limiting step of the mechanism, the cis-aconitate is released from the enzyme, then reattached in the isocitrate mode to complete the reaction. This rate-liming step ensures that the right stereochemistry
Stereochemistry
Stereochemistry, a subdiscipline of chemistry, involves the study of the relative spatial arrangement of atoms within molecules. An important branch of stereochemistry is the study of chiral molecules....

, specifically (2R,3S), is formed in the final product. Another hypothesis is that cis-aconitate stays bound to the enzyme while it flips from the citrate to the isocitrate mode.

In either case, flipping cis-aconitate allows the dehydration and hydration steps to occur on opposite faces of the intermediate. Aconitase catalyzes trans elimination/addition of water, and the flip guarantees that the correct stereochemistry is formed in the product. To complete the reaction, the serine and histidine residues reverse their original catalytic actions: the histidine, now basic, abstracts a proton from water, priming it as a nucleophile
Nucleophile
A nucleophile is a species that donates an electron-pair to an electrophile to form a chemical bond in a reaction. All molecules or ions with a free pair of electrons can act as nucleophiles. Because nucleophiles donate electrons, they are by definition Lewis bases.Nucleophilic describes the...

 to attack at C2, and the protonated serine is deprotonated by the cis-aconitate double bond to complete the hydration, producing isocitrate.

Disease Relevance

A serious ailment associated with aconitase is known as aconitase deficiency. It is caused by a mutation in the gene for iron-sulfur cluster scaffold protein (ISCU
ISCU
Iron-sulfur cluster assembly enzyme ISCU, mitochondrial is a protein that in humans is encoded by the ISCU gene.-External links:* -External links:*...

), which helps build the Fe-S cluster on which the activity of aconitase depends. The main symptoms are myopathy
Myopathy
In medicine, a myopathy is a muscular disease in which the muscle fibers do not function for any one of many reasons, resulting in muscular weakness. "Myopathy" simply means muscle disease...

 and exercise intolerance
Exercise intolerance
Exercise intolerance is a condition where the patient is unable to do physical exercise at the level or for the duration that would be expected of someone in his or her general physical condition, or experiences unusually severe post-exercise pain, fatigue, or other negative effects...

; physical strain is lethal for some patients because it can lead to circulatory shock. There are no known treatments for aconitase deficiency.

Another disease associated with aconitase is Friedreich's ataxia
Friedreich's ataxia
Friedreich's ataxia is an inherited disease that causes progressive damage to the nervous system, resulting in symptoms ranging from gait disturbance to speech problems; it can also lead to heart disease and diabetes....

 (FRDA), which is caused when the Fe-S proteins in aconitase and succinate dehydrogenase have decreased activity. A proposed mechanism for this connection is that decreased Fe-S activity in aconitase and succinate dehydrogenase is correlated with excess iron concentration in the mitochondria and insufficient iron in the cytoplasm, disrupting iron homeostasis. This deviance from homeostasis causes FRDA, a neurodegenerative disease for which no effective treatments have been found.

Finally, aconitase is thought to be associated with diabetes. Although the exact connection is still being determined, multiple theories exist. In a study of organs from mice with alloxan diabetes (experimentally induced diabetes) and genetic diabetes, lower aconitase activity was found to decrease the rates of metabolic reactions involving citrate, pyruvate, and malate. In addition, citrate concentration was observed to be unusually high. Since these abnormal data were found in diabetic mice, the study concluded that low aconitase activity is likely correlated with genetic and alloxan diabetes. Another theory is that, in diabetic hearts, accelerated phosphorylation of heart aconitase by protein kinase C causes aconitase to speed up the final step of its reverse reaction relative to its forward reaction. That is, it converts isocitrate back to cis-aconitate more rapidly than usual, but the forward reaction proceeds at the usual rate. This imbalance may contribute to disrupted metabolism in diabetics.

Family members

Aconitases are expressed in bacteria to humans. Humans express the following two aconitase isozyme
Isozyme
Isozymes are enzymes that differ in amino acid sequence but catalyze the same chemical reaction. These enzymes usually display different kinetic parameters Isozymes (also known as isoenzymes) are enzymes that differ in amino acid sequence but catalyze the same chemical reaction. These enzymes...

s:

External links

- the Aconitase structure in interactive 3D
The source of this article is wikipedia, the free encyclopedia.  The text of this article is licensed under the GFDL.
 
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