Isocitric acid

Aconitic acid involved in the TCA and glyoxylate cycles and the acid commonly occurring in nature has the c/5-configuration. The trans- om x has also been isolated from some plant materials - for example, sugarcane Saccharum offi-cinarum) juice (17), tomato (Lycopersicon esculentum) (56) or moss (Bryophyta) (34). However, some of the occurrences might be artifacts of the isolation procedures, for an interconversion between two isomers of aconitic acid has been reported (14, 81). In both cycles, cw-aconitic acid is formed upon dehydration of citric acid catalyzed by aconitase (aconitate hydratase) which also catalyzes the rehydration of cw-aconitate to isocitric acid. 

Of the four stereoisomers, erythro-Ti - and Ls-, and threo-T - and Lg-isodtric acid, it is the t/j/ieo-Dg-isomer that occurs in nature. The formation of isocitric acid from c/s-aconitic acid is catalyzed by aconitase. This acid is also accumulated in Crassulacean plants as a result of dark CO2 fixation. 

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

The enzyme aconitase catalyzes the hydration of aconitic acid to two products citric acid and isocitric acid. Isocitric acid is optically active citric acid is not. What are the respective constitutions of citric acid and isocitric acid ... 

Isocitric acid, an intermediate in the citric acid cycle of food metabolism, has the systematic name (2/ ,3S)-3-carboxy-2-hydroxypentanedioic acid. Draw the structure. 

This cycle is a series of reactions starting from the reaction of oxaloacetic acid with acetylcoenzyme A and finally regenerates oxaloacetic acid again, forming two moles of carbon dioxide during one cycle. The first step of the decarboxylation is conjugated with the oxidation of isocitric acid (Fig. 3). 

As mentioned in the introductory part, stereochemical course of the conversion of isocitric acid to a-ketoglutaric acid in TCA cycle is completely enantiose-lective although the reaction does not form an asymmetric carbon in the usual metabolic path. If such type of oxidative decarboxylation can be applied to synthetic compounds, it is expected that an entirely new type of asymmetric biotransformation will be developed. 

An attempt was made to convert aconitic acid (2 g) to isocitric acid using 25 ml. of 30% aqueous hydrogen peroxide, time and temperature unspecified. A crystalline product precipitated which exploded on grinding in a mortar. 

Polarographic studies of a mitochondrial fraction from Hymenolepis diminuta showed that of four substrates tested, DL-glycerol-3-phosphate was the most rapidly oxidized, but the highest respiratory control ratio (1.7) was obtained with dl-isocitric acid. With isocitrate as substrate oxyclozanide at 1.61 nM stimulated O uptake and relieved oligomycin inhibition of adinosine diphosphate-stimulated respiration, but at concentrations above 2 pM progressively inhibited O uptake. Rafoxanide, niclosamide, 3,4,5-tribromo-salicylanilide, nitroxynil, resorantel, di-chlorophen, and 2,4-dinitrophenol exhibited effects similar to those of oxyclozanide on the respiration in cestode mitochondria. The relative potencies were compared and the possible mode of action discussed [38]. 

Also, the carboxylation of oxoglutaric acid to form isocitric acid catalyzed by isocitric acid dehydrogenase (ICDH) can be driven electrochemically using methyl viologen as mediator without reduced pyridine nucleotides as cofactors. Thus, electrolysis at — 0.95 V vs SCE of a 0.2 M tris buffer solution (pH 7.7)... 

Fig. 20. Electroenzymatic carboxylation of oxoglutaric acid with isocitric acid dehydrogenase (ICDH) using ferredoxin (Fd) as mediator and poly-L-lysine as promoter... 

Isocitric acid, in citric acid cycle, 6 633 Isocontour surfaces, discrete, 10 340 Isocorrosion diagrams, 23 784 Isocratic chromatography, 3 827 Isocrotonic acid, physical properties,... 

Aconitase, an unstable enzyme,4 is concerned with the reversible conversion of cis-aconitate to either citric acid or isocitric acid. It may be noted that the entire system of tricarboxylic cycle enzymes are present in the mitochondria separated from cells, and, furthermore, it has been found that the mitochondrial enzymes differ from the isolated enzymes in that the former require no addition of D.P.N. (co-enzyme I) or T.P.N. (co-enzyme II) for activity. Peters suggests that the citrate accumulation is caused by the competitive reaction of the fluorocitrate with aconitase required for the conversion of citrate to isocitrate. This interference with the tricarboxylic acid... 

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 ... 

Gawron et al. (13,14) determined the stereochemistry of natural isocitric acid by chemical means. The results require the rrons-addition of water across the cis-aconitate intermediate double bond to produce either citrate or 2R,3S-isodtrate. Mass and NMR analyses of isotopically labeled citrate and isocitrate in the early 1960 s (15-17), defined the stercospedficities of the dehydration steps. These results led Gawron to propose the binding of cis-aconitate to the active site in two orientations differing by a 180° rotation about the double bond, as shown in Equation 2. This allows for the protonation by a base (-BH) and hydroxylation of the double bond to occur on aconitase at single, separate loci for the formation of either citrate or isocitrate. 

The enzyme isocitrate dehydrogenase is one of the enzymes of the Krebs or citric acid cycle, a major feature in carbohydrate metabolism (see Section 15.3). This enzyme has two functions, the major one being the dehydrogenation (oxidation) of the secondary alcohol group in isocitric acid to a ketone, forming oxalosuccinic acid. This requires the cofactor NAD+ (see Section 11.2). For convenience, we are showing non-ionized acids here, e.g. isocitric acid, rather than anions, e.g. isocitrate. 

The second function, and the one pertinent to this section, is the decarboxylation of oxalosuccinic acid to 2-oxoglutaric acid. This is simply a biochemical example of the ready decarboxylation of a P-ketoacid, involving an intramolecular hydrogen-bonded system. This reaction could occur chemically without an enzyme, but it is known that isocitric acid, the product of the dehydrogenation, is still bound to the enzyme isocitrate dehydrogenase when decarboxylation occurs. 

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. 

Certain microorganisms have a modification of this cycle in which isocitric acid is cleaved to succinic acid and glyoxylic acid. The latter acid is condensed with acetyl-CoA to form malic acid. In this modification (the glvoxvlic acid cvcle), oxalsuccinic acid and alpha-ketoglularic acid are not involved. This is sometimes referred to as the glyoxylate shunt pathway. 

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