Pyruvate decarboxylation, enzymatic

A new development is the industrial production of L-phenylalanine by converting phenylpyruvic add with pyridoxalphosphate-dependent phenylalanine transaminase (see Figure A8.16). The biotransformation step is complicated by an unfavourable equilibrium and the need for an amino-donor (aspartic add). For a complete conversion of phenylpyruvic add, oxaloacetic add (deamination product of aspartic add) is decarboxylated enzymatically or chemically to pyruvic add. The use of immobilised . coli (covalent attachment and entrapment of whole cells with polyazetidine) is preferred in this process (Figure A8.17). 

Scheme 3. Reaction path of enzymatic pyruvate decarboxylation and formation of a-hydroxy ketones...

The simplest example of such reactions is the decarboxylation of pyruvate. Both model and enzyme studies have shown the intermediacy of covalent complexes formed between the cofactor and the substrate. Kluger and coworkers have studied extensively the chemical and enzymatic behavior of the pyruvate and acetaldehyde complexes of ThDP (2-lactyl or LThDP, and 2-hydroxyethylThDP or HEThDP, respectively) . As Scheme 1 indicates, the coenzyme catalyzes both nonoxidative and oxidative pathways of pyruvate decarboxylation. The latter reactions are of immense consequence in human physiology. While the oxidation is a complex process, requiring an oxidizing agent (lipoic acid in the a-keto acid dehydrogenases , or flavin adenine dinucleotide, FAD or nicotinamide adenine dinucleotide , NAD " in the a-keto acid oxidases and Fe4.S4 in the pyruvate-ferredoxin oxidoreductase ) in addition to ThDP, it is generally accepted that the enamine is the substrate for the oxidation reactions. 

The authors chose pyruvic acid as their model compound this C3 molecule plays a central role in the metabolism of living cells. It was recently synthesized for the first time under hydrothermal conditions (Cody et al., 2000). Hazen and Deamer carried out their experiments at pressures and temperatures similar to those in hydrothermal systems (but not chosen to simulate such systems). The non-enzymatic reactions, which took place in relatively concentrated aqueous solutions, were intended to identify the subsequent self-selection and self-organisation potential of prebiotic molecular species. A considerable series of complex organic molecules was tentatively identified, such as methoxy- or methyl-substituted methyl benzoates or 2, 3, 4-trimethyl-2-cyclopenten-l-one, to name only a few. In particular, polymerisation products of pyruvic acid, and products of consecutive reactions such as decarboxylation and cycloaddition, were observed the expected tar fraction was not found, but water-soluble components were found as well as a chloroform-soluble fraction. The latter showed similarities to chloroform-soluble compounds from the Murchison carbonaceous chondrite (Hazen and Deamer, 2007). 

Rates have been monitored by ultraviolet absorption,60 following the decay of oxalacetate, and by enzymatic assays61,62 responding to the production of pyruvate [see (32)]. Figure 19 illustrates the rapid drop in absorbance with time for oxalacetate in the presence of the primary-amine-containing polymer. Even a sixtyfold excess of substrate over primary-amine concentration was completely decarboxylated. This and... 

It has also been possible to confirm the presence of the reduction product of a Schiff base on the polymer by proton magnetic resonance. For this purpose we have used unmodified poly(ethylenimine), since it too catalyzes the decarboxylation of oxalacetate to its product, pyruvate. Unmodified polyethylenimine was mixed with oxalacetate-4-ethyl ester. One-half of this solution was treated with NaBH4 the second half was exposed to a similar environment but no NaBH4 was added. The borohydride-treated polymer exhibited a strong triplet in the nmr spectrum centered at 3.4 ppm upfield from the HOD resonance. This new feature would be expected from the terminal methyl protons of the oxalacetate ester attached to the polymer. Only a very weak triplet was found in the control sample not treated with borohydride. These observations are strong evidence for the formation of Schiff bases with the polymer primary amine groups. Evidently the mechanistic pathway for decarboxylation by the polymer catalyst is similar to that used enzymatically. 

The pyruvate formed by decarboxylation of malate in bundle-sheath cells is transferred back to the mesophyll cells, where it is converted to PEP by an unusual enzymatic reaction catalyzed by pyruvate phosphate dikinase (Fig. 20-23b). This enzyme is called a dikinase because two different molecules are simultaneously phosphorylated by one molecule of ATP pyruvate to PEP, and phosphate to pyrophosphate. The pyrophosphate is subsequently hydrolyzed to phosphate, so two high-energy phosphate groups of ATP are used in regenerating PEP. The PEP is now ready to receive another molecule of C02 in the mesophyll cell. 

Practical and fundamental aspects of malo-lactic fermentation are given. Conditions which winemakers can use for better control of the fermentation, including detailed procedures for inoculation with Leuconostoc oenos ML 34 and for inhibition with fumaric acid, are presented. New information on the role of malic acid decarboxylation in bacterial metabolism and on the enzymatics of malic acid decarboxylation are reviewed. The malic acid decarboxylation seems to involve two pathways a direct decarboxylation of malic to lactic acid with NAD as a coenzyme and a concurrent but small oxidative decarboxylation to pyruvic acid and NADH. How these pathways can bring about the marked stimulation of bacterial growth rate by the malo-lactic reaction and their negligible effect on growth yield are discussed. 

As a side activity, many decarboxylases catalyze the formation of C-C bonds. In the reaction of two pyruvate molecules, catalyzed by pyruvate decarboxylase (PDC, E.C. 4.1.1.1), a-acetolactate is formed, an important intermediate of valine biosynthesis. In turn, a-acetolactale can be oxidatively decarboxylated by oxygen to diacetyl or enzymatically decarboxylated by acetolactate decarboxylase (ADC, E.C. 4.1.1.5) to (] )-acetoin (Figure 7.29). 

No 3-carboxy-substituted TBCs, derived from L-tryptophan by the Pic-tet-Spengler route, have yet been isolated from mammalian tissues. The same is also true for the dicarboxylic acid 23a derived from the condensation of L-tryptophan with pyruvic acid (36). The 1-carboxy-substituted TBCs 37 and 38, on the other hand, occur in mammalian systems (70,71) and are metabolically decarboxylated (65,S5). Whether a direct enzymatic decarboxylation of racemic material, occurring with the (S) and (R) enantiomers at a different rate, could account for the formation of unequal amounts of the enantiomers of TBC has not been investigated so far. The pyruvic acid route to optically active TBC (Fig. 12) leading from TBC 38a to TBC 29a via DBC 34 is at tifie moment the preferred pathway (85,86,89), although the enzymes involved in the asymmetric reduction leading to TBC 29a and the hydroxylated metabolites TBCs 30a and 33a have been neither isolated nor characterized. 

The reaction of optically active carbinolamines formed by an enzymatically controlled addition of acetaldehyde to amines, illustrated in Fig. 2, may be of theoretical interest, but lacks experimental verification it also would require the presence of acetaldehyde. The more likely pyruvic acid route to optically active TIQs, however, also remains inconclusive. If it indeed proceeds through TIQ-1-carboxylic acids to DIQ intermediates by an oxidative decarboxylation (176,217,218), it requires that it be followed by an asymmetric enzymatic reduction. Although achieved in vitro (35), this reaction has not been realized in vivo. The formation of unequal amounts of the optical isomers of salsolinol and other TIQs in vivo could arise from racemic 1-carboxy-TIQ in an enzymatic decarboxylation, proceeding with (S) and (R) enantiomers at a different rate and thus affording different amounts of (5)- and (/ )-TIQ. With the availability of optically active TIQ-1-carboxylic acids, this possibility can now be tested. 

Do optically active 1-methyl-TIQs, as sketched in Fig. 32 for the synthesis of (7 )-salsolinol, originate from a Pictet-Spengler reaction of dopamine with acetaldehyde derive from ethanol, or are they the result of a Pictet-Spengler reaction of biogenic amines with pyruvic acid, as sketched in Fig. 33 Based on the accumulated data it seems reasonable to propose that optically active TIQs are formed by the pyruvic acid pathway, and that the pyruvic acids may be derived from an impaired glucose metabolism or an impaired amino acid metabolism. Whether the intermediate TIQ-1-carboxylic acids 91a,b are enzymatically decarboxylated to afford 64a,b in a different enantiomeric ratio, or whether optically active TIQs are formed by oxidative decarboxylation of TIQ 91 to DIQ 120, followed by an asymmetric reduction, remains open to question. 

A biomimetic synthesis of solerone (5-oxo-4-hexanolide) 1 using both enzymatic and acid-catalyzed reactions was performed. Starting from L-glutamic acid 5-ethyl ester 2 enzymatic oxidative deamination followed by subsequent decarboxylation of the corresponding 2-oxoglutaric acid 5-ethyl ester 3 led to ethyl 4-oxobutanoate 4. In the presence of pyruvate,... 

While in the presence of 2-oxoglutaric acid neither decarboxylation nor acyloin condensation had been observed, as expected from previously published results (75), we succeeded in the enzymatic conversion of the mono ethyl ester 3 to ethyl 4-oxobutanoate 4, using both whole yeast cells (Saccharomyces cerevisiae) and purified PDC. The oxo ester 4 served as substrate for a second reaction catalyzed by PDC. Formation of a new carbon-carbon bond was accomplished in the presence of pyruvic acid which acted as donor of a C2-unit. Thus, ethyl 4-hydroxy-5-oxohexanoate 5 was obtained for the first time as the result of an enzymatic acyloin condensation. Finally, traces of acid induced the lactonization of hydroxyester 5, indicating it as direct precursor of solerone 1 (Figure 1). 

TCA cycle. (tricarboxylic acid cycle Krebs cycle citric acid cycle). A series of enzymatic reactions occurring in living cells of aerobic organisms, the net result of which is the conversion of pyruvic acid, formed by anaerobic metabolism of carbohydrates, into carbon dioxide and water. The metabolic intermediates are degraded by a combination of decarboxylation and dehydrogenation. It is the major terminal pathway of oxidation in animal, bacterial, and plant cells. Recent research indicates that the TCA cycle may have predated life on earth and may have provided the pathway for formation of amino acids. 

Terpenoids do not necessarily contain exact multiples of five carbons and allowance has to be made for the loss or addition of one or more fragments and possible molecular rearrangements during biosynthesis. In reality the terpenoids are biosynthesized from acetate units derived from the primary metabolism of fatty acids, carbohydrates and some amino acids (see Fig. 2.10). Acetate has been shown to be the sole primary precursor of the terpenoid cholesterol. The major route for terpenoid biosynthesis, the mevalonate pathway, is summarized in Fig. 2.16. Acetyl-CoA is involved in the generation of the C6 mevalonate unit, a process that involves reduction by NADPH. Subsequent decarboxylation during phosphorylation (i.e. addition of phosphate) in the presence of ATP yields the fundamental isoprenoid unit, isopentenyl pyrophosphate (IPP), from which the terpenoids are synthesized by enzymatic condensation reactions. Recently, an alternative pathway has been discovered for the formation of IPP in various eubacteria and plants, which involves the condensation of glyceraldehyde 3-phosphate and pyruvate to form the intermediate 1-deoxy-D-xylulose 5-phosphate (Fig. 2.16 e.g. Eisenreich et al. 1998). We consider some of the more common examples of the main classes of terpenoids below. 

Kumagai and coworkers11131 developed an enzymatic procedure to produce d-alanine from fumarate by means of aspartase (E. C. 4.3.1.1), aspartate racemase, and D-amino acid aminotransferase (Fig. 17-12). Aspartase catalyzes conversion of fumarate into L-aspartate, which is racemized to form D-aspartate. D-Amino acid aminotransferase catalyzes transamination between D-aspartate and pyruvate to produce D-alanine and oxalacetate. This 2-oxo acid is easily decarboxylated spontaneously to form pyruvate in the presence of metals. Thus, the transamination proceeds exclusively toward the direction of D-alanine synthesis, and total conversion of fumarate into D-alanine was achieved. 

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