Isocitrate isomerization

Perhaps the best characterized example of a subsite differentiated [4Fe-4S] protein is aconitase, which catalyzes the citrate-isocitrate isomerization in the citric acid cycle (257). Aconitase isolated aerobically is inactive and contains a [3Fe-4S] cluster. Activity is restored by incubation with Fe and this also reconstitutes the [4Fe-4S] cluster. Oxidation of the core results in loss of the fourth iron atom, regenerating the [3Fe-4S] form. Mossbauer studies have demonstrated that only one of the four iron sites is exchanged (258). X-ray studies on both [3Fe-4S] and [4Fe-4S] forms of pig heart aconitase 258a) showed that insertion of iron into [3Fe-4S] occurs isomorphously. The positions of the common atoms in the two forms of the core agree to within 0.1 A, supporting the view of the [3Fe-4S] cluster as an iron-voided cubane. A similar result was obtained for the seven iron ferredoxin from Azo-... 

Assign configurations, using the sequence rule, to each chiral center of the stereo-isomeric isocitric acids and alloisocitric acids ... 

Citrate is isomerized to isocitrate by aconitase in a two-step process involving aconitate as an intermediate (Figure 20.7). In this reaction, the elements... 

Schossberger has recently isolated the isomeric form of citral which is knowm as isocitral, corresponding to the isogeraniol resulting from enolised citral acetate. 

Citrate is isomerized by dehydration and rehydration to give isocitrate. 

Step 2 of Figure 29.12 Isomerization Citrate, a prochiral tertiary alcohol, is next converted into its isomer, (2, 35)-isocitrate, a chiral secondary alcohol. The isomerization occurs in two steps, both of which are catalyzed by the same aconitase enzyme. The initial step is an ElcB dehydration of a /3-hydroxy acid to give cfs-aconitate, the same sort of reaction that occurs in step 9 of glycolysis (Figure 29.7). The second step is a conjugate nucleophilic addition of water to the C=C bond (Section 19.13). The dehydration of citrate takes place specifically on the pro-R arm—the one derived from oxaloacetate—rather than on the pro-S arm derived from acetyl CoA. 

Citrate is isomerized to isocitrate by the enzyme aconitase (aconitate hydratase) the reaction occurs in two steps dehydration to r-aconitate, some of which remains bound to the enzyme and rehydration to isocitrate. Although citrate is a symmetric molecule, aconitase reacts with citrate asymmetrically, so that the two carbon atoms that are lost in subsequent reactions of the cycle are not those that were added from acetyl-CoA. This asymmetric behavior is due to channeling— transfer of the product of citrate synthase directly onto the active site of aconitase without entering free solution. This provides integration of citric acid cycle activity and the provision of citrate in the cytosol as a source of acetyl-CoA for fatty acid synthesis. The poison fluo-roacetate is toxic because fluoroacetyl-CoA condenses with oxaloacetate to form fluorocitrate, which inhibits aconitase, causing citrate to accumulate. 

This interpretation offered by Ogston was confirmed by further isotope experiments (Potter and Heidelberger, 1949 Martius and Schorre, 1950). Ochoa and his colleagues also demonstrated that in pigeon-liver preparations which had been freed from aconitase, thus preventing isomerization between citrate and isocitrate, citrate was the product of the condensation of oxaloacetate and acetate. 

There have been a number of isolated studies of metal-ion catalyzed nucleophilic reactions of other groupings. Particularly interesting is the induced nucleophilic attack on olefins. Hydration is normally very sluggish. Enzymes can speed up such reactions. Aconitase, an iron-containing enzyme, catalyzes the isomerization of citric acid to isocitric acid, through the intermediacy of cis-aconitic acid. A possible mechanism has been suggested based on the following Co(III) model chemistry. Rapid cyclization of the maleate ester produces Ai and AS chelated malate half ester ... 

Citrate is subsequently isomerized to isocitrate this involves dehydration and rehydration via the intermediate cis-aconitate. Both reactions are... 

Isomers are molecules with the same composition (i. e. the same molecular formula), but with different chemical and physical properties. If isomers differ in the way in which their atoms are bonded in the molecule, they are described as structural isomers (cf citric acid and isocitric acid, D). Other forms of isomerism are based on different arrangements of the substituents of bonds (A, B) or on the presence of chiral centers in the molecule (C). 

Enzymes usually function stereospedfically. In chiral substrates, they only accept one of the enantiomers, and the reaction products are usually also sterically uniform. Aconitate hydratase (aconitase) catalyzes the conversion of citric acid into the constitution isomer isocitric acid (see p.l36). Although citric acid is not chiral, aconitase only forms one of the four possible isomeric forms of isocitric acid (2i ,3S-isocitric acid). The intermediate of the reaction, the unsaturated tricarboxylic acid aconitate, only occurs in the cis form in the reaction. The trans form of aconitate is found as a constituent of certain plants. 

Citrate is isomerized to isocitrate by aconitase (see Figure 9.5). [Note Aconitase is inhibited by fluoroacetate, a compound that is used as a rat poison. Fluoroacetate is converted to fluoroacetyl CoA, which condenses with oxaloacetate to form fluorocitrate—a potent inhibitor of aconitase—resulting in citrate accumulation.]... 

Citrate is synthesized from oxaloacetate (OAA) and acetyl CoA by citrate synthase. This enzyme is allosterically activated by ADP, and inhibited by ATP, NADH, succinyl CoA, and fatty acyl CoA derivatives. Citrate is isomerized to isocitrate by aconitase, an enzyme that is targeted by the rat poison, fluoroacetate. 

Citrate Synthase Is the Gateway to the TCA Cycle Aconitase Catalyzes the Isomerization of Citrate to Isocitrate... 

Aconitase, the enzyme that catalyzes this isomerization, is named for the fact that the unsaturated compound formed by removing H20 from either citrate or isocitrate, cA-aconitate, can also serve as substrate or product. Aconitase catalyzes the attainment of equilibrium between citrate, isocitrate, and civ-acon itate. The three compounds may be considered as belonging to the same metabolic pool. 

The enzyme aconitase catalyzes the isomerization of citric acid to isocitric acid via the intermediate cis-aconitic acid (Scheme 46),530 and various attempts have been made to model this reaction.21 The cobalt Ill) complexes derived from methyl maleate (171) and methyl fumarate (172) have been prepared531 to study intramolecular attack by coordinated hydroxide on the alkene. Generation of the hydroxo species of the maleic acid complex leads to rapid cyclization to give the... 

Aconitase is the trivial name for citrate dehydratase cw-aconitate hydratase (EC 4.2.1.3). It catalyzes the reversible isomerization reaction of citrate into isocitrate via the intermediate cA-aconitate (Figure 2). It is a water-soluble, monomeric protein. In eukaryotic cells aconitase is located in the mitochondrial matrix. In prokaryotes the enzyme occurs in the cytoplasma. The pig heart enzyme consists of 754 amino-acid residues, providing a molecular mass of 83 kDa [27], Aconitase from other sources has similar size. The porcine protein is synthesized with a mitochondrial targeting sequence. The mature, functional protein can be (over)expressed in Escherichia coli [28],... 

Citrate is converted to isocitrate (6C) by an isomerization catalyzed by aconi-tase. This is actually a two-step reaction during which c/s-aconitate is formed as an intermediate. It is the c/s-aconitate which gives the enzyme its name. 

Fig. 3. Condensation of acetyl-CoA and oxaloacetate to citrate and isomerization of citrate into isocitrate. The numbers next to the carbon atoms indicate the enumeration of the metabolite carbon atoms. OAA oxaloacetate AcCoA acetyl-coenzyme A...

The tertiary hydroxyl group is not properly located in the citrate molecule for the oxidative decarboxylations that follow. Thus, citrate is isomerized into isocitrate to enable the six-carbon unit to undergo oxidative decarboxylation. The isomerization of citrate is accomplished by a dehydration step followed by a hydration step. The result is an interchange of a hydrogen atom and a hydroxyl group. The enzyme catalyzing both steps is called aconitase because cis-aconitate is an intermediate. 

The cycle starts with the condensation of oxaloacetate (C4) and acetyl CoA (C2) to give citrate (Cg), which is isomerized to isocitrate (Cg). Oxidative decarboxylation of this intermediate gives a-ketoglutarate (C5). The second molecule of... 

Aconitase catalyzes the isomerization of citrate to isodtrate, isocitrate dehydrogenase catalyzes the oxidative decarboxylation of isocitrate to a-ketoglutarate, and a-ketoglutarate dehydrogenase catalyzes the oxidative decarboxylation of a-keto-glutarate to succinyl-CoA. Succinyl-CoA and the remaining intermediates are the 4-carbon intermediates of the Krebs cycle. Succinyl thiokinase catalyzes the release of coenzyme A from succinyl-CoA and the production of GTP. Succinate dehydro-... 

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