Decarboxylation Of Oxaloacetic Acid Research Articles

Computational insights into the binding modes, keto-enol tautomerization and stereo-electronically controlled decarboxylation of oxaloacetate in the active site of macrophomate synthase.

Oxaloacetic acid (OAA) is a β-ketocarboxylic acid, which plays an important role as an intermediate in some metabolic pathways, including the tricarboxylic acid cycle, gluconeogenesis and fatty acid biosynthesis. Animal studies have indicated that supplementing oxaloacetic acid shows an increase of lifespan and other substantial health benefits including mitochondrial DNA protection, and protection of retinal, neural and pancreatic tissues. Most of the chemical transformations of OAA in the metabolic pathways have been extensively studied; however, the understanding of decarboxylation of OAA at the atomic level is relatively lacking. Here, we carried out MD simulations and combined quantum mechanical/molecular mechanical (QM/MM) calculations as an example to systematically elucidate the binding modes, keto-enol tautomerization and decarboxylation of OAA in the active site of macrophomate synthase (MPS), which is a Mg(II)-dependent bifunctional enzyme that catalyzes both the decarboxylation of OAA and [4+2] cycloaddition of 2-pyrone with the decarboxylated intermediate of OAA (pyruvate enolate). On the basis of our calculations, it was found that the Mg2+-coordinated oxaloacetate may exist in enol forms and keto forms. The four keto forms can be transformed into each other by simply rotating the C2-C3 single bond, nevertheless, the keto-enol tautomerization strictly requires the assistance of pocket water molecules. In addition, the decarboxylation is stereo-electronically controlled, i.e., it is the relative orientation of the terminal carboxyl anion that determines the rate of decarboxylation. As such, the chemistry of oxaloacetate in the active site of MPS is complex. On one hand, the most stable binding mode (K-I) may undergo enol-keto tautomerization to isomerize to the enol form, which may further react with the second substrate; on the other hand, K-I may isomerize to another binding mode K-II to proceed decarboxylation to generate pyruvate enolate and CO2. Starting from K-I, the enol-keto tautomerization corresponds to a barrier of 16.2 kcal mol-1, whereas the decarboxylation is associated with an overall barrier of 19.7 kcal mol-1. These findings may provide useful information for understanding the chemistry of OAA and the catalysis of related enzymes, and they are basically in agreement with the available experimental kinetic data.

Effect of Ammonium Salts on the Decarboxylation of Oxaloacetic Acid in Atmospheric Particles

Oxaloacetic acid (OAA) is a 3-oxocarboxylic acid formed from the oxidation of succinic acid. OAA and other 3-oxocarboxylic acids experience a decarboxylation reaction in aqueous solutions, which can be catalyzed by ammonium and amines. This catalysis has not been studied under atmospherically relevant conditions despite previous interest in OAA in the atmosphere. To address this, 1 mM solutions of OAA were prepared with varying concentrations of ammonium sulfate, ammonium bisulfate, ammonium chloride, and sodium sulfate to simulate various atmospheric conditions. The extent of the decarboxylation was monitored using UV–visible absorption spectroscopy. OAA’s uncatalyzed decarboxylation lifetime was around 5 h. Under moderately acidic conditions representative of aerosol particles (pH = 3–4), the decarboxylation rate increased linearly with ammonium concentration up to about 2.7 M, after which additional ammonium had no effect. The effective lifetime of OAA reduced to approximately 1 h under these conditions. Density functional theory calculations support the proposed catalytic mechanism, predicting the free energy barrier height for decarboxylation to be approximately 21 kcal/mol lower after OAA has reacted with ammonium. In more acidic solutions (pH < 1), OAA’s decarboxylation was suppressed, with lifetimes of tens of hours, even in the presence of ammonium. A comparison of the decarboxylation rate with the expected rate of oxidation by OH suggests that decarboxylation will be the dominant loss mechanism for OAA, and presumably other 3-oxocarboxylic acids, in aerosol particles and cloud/fog droplets. This result explains why OAA is hard to detect in field measurements even though it is a known oxidation product of succinic acid.

Ethylenediamine Catalyzed Decarboxylation of Oxaloacetic Acid: A DFT Investigation

The decarboxylation mechanism of oxaloacetic acid aided with ethylenediamine or without any catalyst is investigated employing Density Functional Theory (DFT). DFT calculations for both the gas phase and in water solution indicate a stepwise mechanism for each of the steps of the reactions. In the catalyzed mechanism, the dehydration of carbinolamine (IM1) is via a seven-membered ring transition structure (TS5), which is consistent with the structure proposed by Thalji, et al. The decarboxylation of the imine (IM6) is the rate determining step with an energy barrier of 16.46 kcal/mol, lower than the reaction without any catalysts or catalyzed with ions.

Structures of Rat Cytosolic PEPCK: Insight into the Mechanism of Phosphorylation and Decarboxylation of Oxaloacetic Acid,

The structures of the rat cytosolic isoform of phosphoenolpyruvate carboxykinase (PEPCK) reported in the PEPCK-Mn2+, -Mn2+-oxaloacetic acid (OAA), -Mn2+-OAA-Mn2+-guanosine-5'-diphosphate (GDP), and -Mn2+-Mn2+-guanosine-5'-tri-phosphate (GTP) complexes provide insight into the mechanism of phosphoryl transfer and decarboxylation mediated by this enzyme. OAA is observed to bind in a number of different orientations coordinating directly to the active site metal. The Mn2+-OAA and Mn2+-OAA-Mn2+GDP structures illustrate inner-sphere coordination of OAA to the manganese ion through the displacement of two of the three water molecules coordinated to the metal in the holo-enzyme by the C3 and C4 carbonyl oxygens. In the PEPCK-Mn2+-OAA complex, an alternate bound conformation of OAA is present. In this conformation, in addition to the previous interactions, the C1 carboxylate is directly coordinated to the active site Mn2+, displacing all of the waters coordinated to the metal in the holo-enzyme. In the PEPCK-Mn2+-GTP structure, the same water molecule displaced by the C1 carboxylate of OAA is displaced by one of the gamma-phosphate oxygens of the triphosphate nucleotide. The structures are consistent with a mechanism of direct in-line phosphoryl transfer, supported by the observed stereochemistry of the reaction. In the catalytically competent binding mode, the C1 carboxylate of OAA is sandwiched between R87 and R405 in an environment that would serve to facilitate decarboxylation. In the reverse reaction, these two arginines would form the CO2 binding site. Comparison of the Mn2+-OAA-Mn2+GDP and Mn2+-Mn2+GTP structures illustrates a marked difference in the bound conformations of the nucleotide substrates in which the GTP nucleotide is bound in a high-energy state resulting from the eclipsing of all three of the phosphoryl groups along the triphosphate chain. This contrasts a previously determined structure of PEPCK in complex with a triphosphate nucleotide analogue in which the analogue mirrors the conformation of GDP as opposed to GTP. Last, the structures illustrate a correlation between conformational changes in the P-loop, the nucleotide binding site, and the active site lid that are important for catalysis.

Stability constants and rate coefficients for decarboxylation of oxalacetic acid and some metal oxalacetates

Data collected at constant ionic strengths, from e.m.f. measurements of a glass electrode–calomel electrode cell, u.v. spectrophotometry, and a pressure-sensitive transducer, DVM, micro-computer system, have been used to calculate the stability constants of oxalacetic acid and some metal oxalacetates ML (L = oxalacetate, M = CuII, NiII, ZnII, CoII, or MnII) and the rate coefficients for decarboxylation of the species H2L, HL, L, and ML. The stability constants of the ML species fit the Irving–Williams order (Zn Ni > Co > Mn) but the order of the rate coefficients is Zn Ni ≈ Co > Mn, and while the stability constant order is Ni > Zn the rate coefficient order is Ni ≈ Zn.

Cerium(IV)-induced decarboxylation of penta-ammineoxaloacetatocobalt(III) perchlorate

Cerium(IV) induces electron transfer in penta-ammineoxaloacetatocobalt(III) perchlorate, yielding CO2 and cobalt(II) quantitatively as ultimate products due to carbon–carbon bond fission. Enzymatic decarboxylation of oxaloacetic acid yields only pyruvic acid, whereas cerium(IV) oxidises oxaloacetic acid to CO2, probably because of radical intervention.

Micellar Effects upon p-n-Butylaniline Catalyzed Decarboxylation of Oxalacetic Acid

p-n-Butylaniline ([nBA] = 1 × 10−3 M) incorporated in cationic micelles of cetyltrimethylammonium bromide ([CTABr] = 6 × 10−3M) was found to enhance the rate of decarboxylation of oxalacetic acid (OAA) 45-fold while nBA alone only increases the rate six-fold as compared to the rate of spontaneous decarboxylation at the same pH. Anionic micelles of sodium dodecyl sulfate ([SDS] = 6 × 10−3 M) completely inhibited the catalytic effect of nBA. Together with the effects of added salts and the change of pH on the rate of the reaction the role of micelles in the decarboxylation of OAA is discussed.

Decarboxylation Reaction of Oxalacetic Acid by Metal Chelates

Decarboxylation of oxalacetic acid (OAA) by metal chelates was examined, using copper (II) as a metal and seven kinds of ligand, which forms N, N-coordinated five membered ring chelate, used were 6, 6'-dimethyl-α, α'-dipyridyl, 4, 4'-dimethyl-α, α'-dipyridyl, α, α'-dipyridyl, 5-nitro-ο-phenanthroline, ο-phenanthroline, 4, 7-dimethyl-ο-phenanthroline, and ethylenediamine. When the concentration ratio of dipyridyl to copper (II) was varied, 1 : 1 ratio of copper to dipyridyl effected the fastest decarboxylation rate of OAA, and the rate was faster than that by copper (II) ion alone. Effect of each ligand on the reaction rate was examined and the reaction rate fell in the order of α, α'-dipyridyl, ο-phenanthroline, and ethylenediamine ; 5-nitro-ο-phenanthroline, ο-phenanthroline, and 4, 7-dimethyl-ο-phenanthroline ; α, α'-dipyridyl, 6, 6'-dimethyl-α, α'-dipyridyl, and 4, 4'-dimethyl-α, α'-dipyridyl. When the acetate buffer concentration was 0.1M, the reaction rate was faster, the smaller the pKa, and greater the catalytic effect. When the buffer concentration was 1.0M, the above order did not hold. It was also revealed that steric effect of the ligand affected this reaction. The intermediate formed in this decarboxylation reaction was examined by the measurement of ultraviolet and infrared spectra, and by isolation of this intermediate, and formation of a mixed chelate compound of OAA, copper, and ligand was confirmed.

Phosphate esters as activators of decarboxylation catalytic reactions

The possibilities for increasing the catalytic activity of magnesium and calcium in the decarboxylation of oxaloacetic acid was studied. The alkyl and aryl esters of phosphoric acid were used as activators of the reactions. The application of TBP as activator in 50% ethanolic medium permits the determination of 50μg of magnesium.

Application of decarboxylation reactions in catalytic analysis determination of aluminium

The activation of the catalytic decarboxylation of oxaloacetic acid with some pyridine derivatives was studied in order to select suitable activators for increasing the sensitivity of this type of analytical catalytic reactions. The catalytic activity of some bivalent metal ions (Cu2+, Zn2s+, Cd2+, Pb2+) as well as that of Al3+ was also studied. On that ground a catalytic method for determination of aluminium was developed. The sensitivity of the method is 0.5μg of aluminium in the sample.

Selectivity in the metal-complex-catalyzed decarboxylation of oxaloacetic acid and a role of metal ion in an enzyme system.

Abstract The catalytic action of metal ions in the presence of various coordinating agents and a role of metal ions for the activations of an enzyme in a decarboxylation reaction were investigated. Many of the ligands studied decreased the catalytic activity of metal ions. However, α,α′-dipyridyl selectively enhanced the catalytic activities of divalent metal ions, except Cu2+. The activity of Cu2+ was specifically enhanced only with histidine and its derivatives. Histidine destroyed Ni2+ and Co2+ catalysis. The specific enhancement may be due to the increase in the electronegativities of the metal ions with dπ (metal)-pπ(ligand) back donation and to the stereo-configuration of the metal complexes. The catalytic behavior in these metal ion coordinating agent systems was different from that in the metal-enzyme systems. It was assumed that the activation of enzyme resulted from the change in the configuration of enzyme by the coordination of metal ions, and not from the catalytic action of metal ions enhanced by the coordination of enzyme. This was supported by the fact that the catalysis of compounds containing amino groups, active sites of enzyme, remarkably depended on the stereo-configuration. In the enzyme-model system, Mn2+ was generally preferable to Cu2+ and Ni2+. The catalytic actions of Cu2+ and Ni2+ were lowered by the complex formation with the active sites. This may solve the question why, in enzymatic reactions, Mn2+ having small complex-forming ability is more selective and effective than Cu2+ and Ni2+ with large complex-forming abilities.

Catalytic determination of microamounts of zinc

A catalytic method for determination of micro amounts of zinc has been developed, based on its catalytic action on the decarboxylation of oxalacetic acid. The high sensitivity of the reaction is due to the increase in the effective charge on the metal ion catalyst, when dioxan-water is used as a reaction medium. The determination of more than 5μg of zinc is possible with sufficient accuracy. The method has been applied to the determination of zinc in plants.

Carbon isotope effects in the decarboxylation of oxaloacetic acid. Part 2.—Studies using 146C

Oxaloacetic acid (OAA) enriched in OAA-1-14C was decarboxylated at 25° in aqueous solutions, alone, and also in presence of various rare-earth cations. These catalyze the reaction through complex formation. Normal isotope effects were found in each case, the reactions producing 14CO2 being slower than those producing 12CO2. The effects for the complexes were similar to one another and differed significantly from the result for the uncatalyzed reaction, due to the inductive effects of the cations.

LITHIUM CHLORIDE CATALYZED DECARBOXYLATION OF OXALACETIC ACID IN ETHANOL

The monovalent metal salt lithium chloride promotes the decarboxylation of oxalacetic acid in ethanol. The absorption peak which appears at 230 mμ during the reaction is interpreted to be the enol form of pyruvic acid. Hydrogen chloride has a similar effect on the decarboxylation.

Carbon isotope effects in the decarboxylation of oxaloacetic acid. Part 1.—Studies using 136C

Carbon isotope effects in the decarboxylation of oxaloacetic acid. Part 1.—Studies using 136C

Metal-ion-catalysed Decarboxylation of Oxalacetic Acid in Water–Ethanol Mixtures

OXALACETIC acid (OAA) decarboxylates spontaneously in water1,2 and in various organic solvents with a rate that is dependent on the polarity of the solvent3. Additions of a number of polyvalent metal ions to aqueous solutions of oxalacetic acid promote decarboxylation4–9. The degree of this rate promotion depends on the nature of the metal, on its concentration and on the pH. Addition of ethanol to the aqueous system of the metal ion-promoted decarboxylation of oxalacetic acid decreases the polarity, diminishing both dissociation of the acid and the extent of hydration of the metal ion. These polarity effects are demonstrated by changes in chelation and decarboxylation of oxalacetic acid using magnesium and manganese ion.

L-Malate: NADP oxidoreductase (decarboxylating) from germinating flax rust uredospores

l-Malate: NADP oxidoreductase (decarboxylating) (here called “malic” enzyme) from germinating flax rust uredospores was purified more than twenty-fold and partially characterized. This enzyme preparation was found to catalyze both the oxidative decarboxylation of l-malic acid to pyruvic acid at pH 7·6 and the decarboxylation of oxalacetic acid at pH 5. The oxidative decarboxylation of l-malate was found to specifically need NADP and required Mn 2+ for maximum activity. Optimal pH for enzyme activity was 7·6. A Michaelis constant for l-malate of 1·5 × 10 −4 M was found for this partially purified enzyme preparation. The stability and heat lability of the enzyme are reported in addition to the effect of a number of inhibitors.

Magnetic Catalysis of the Decarboxylation of Oxaloacetic Acid

KINETIC studies of the decarboxylation of phenylmalonic acid in aqueous solution in the presence of high concentrations of rare-earth ions show that diamagnetic ions have little effect on the rate of reaction, while the paramagnetic dysprosium ion leads to an acceleration in rate of the order of 10 per cent1,2. Rare-earth ions do not appear to become co-ordinated appreciably with phenylmalonic acid. Since any magnetic effect must be a very sensitive function of the distance between catalyst and substrate, a study is now being made of the kinetics of decarboxylation of an acid, the anions of which are known to become co-ordinated more strongly with metal ions.

Amine Catalysis in the Decarboxylation of Oxalacetic Acid.

Amine Catalysis in the Decarboxylation of Oxalacetic Acid.

Metal Ion Catalysis of the Decarboxylation of Oxalo-Acetic Acid

IN order to explain the catalysis of oxalo-acetic acid decarboxylation by metal ions, intermediate complex formation between the metal ion and the acid anion has been postulated. In an attempt to determine the nature of this complex, Steinberger and Westheimer1 studied the kinetics of the decarboxylation of the dimethyl substituted acid. In this way they showed that the catalysis did not proceed through an enolic form of the acid. From a further consideration of the general characteristics of decarboxylation reactions, they suggested that the metal complex involved in the catalysis had the formula, MA, which was formed from MHA+, by the loss of a proton.