Pyruvate-dependent decarboxylations

Figure 2. Mechanism of PDH. The three different subunits of the PDH complex in the mitochondrial matrix (E, pyruvate decarboxylase E2, dihydrolipoamide acyltrans-ferase Ej, dihydrolipoamide dehydrogenase) catalyze the oxidative decarboxylation of pyruvate to acetyl-CoA and CO2. E, decarboxylates pyruvate and transfers the acetyl-group to lipoamide. Lipoamide is linked to the group of a lysine residue to E2 to form a flexible chain which rotates between the active sites of E, E2, and E3. E2 then transfers the acetyl-group from lipoamide to CoASH leaving the lipoamide in the reduced form. This in turn is oxidized by E3, which is an NAD-dependent (low potential) flavoprotein, completing the catalytic cycle. PDH activity is controlled in two ways by product inhibition by NADH and acetyl-CoA formed from pyruvate (or by P-oxidation), and by inactivation by phosphorylation of Ej by a specific ATP-de-pendent protein kinase associated with the complex, or activation by dephosphorylation by a specific phosphoprotein phosphatase. The phosphatase is activated by increases in the concentration of Ca in the matrix. The combination of insulin with its cell surface receptor activates PDH by activating the phosphatase by an unknown mechanism.

These pyridoxal-phosphate-dependent (or pyruvate-dependent) enzymes [EC 4.1.1.65] catalyze the decarboxylation of phosphatidyl-L-serine to produce phospha-tidylethanolamine and carbon dioxide. 

One important subgroup of the lyases are the decarboxylases. The decarboxylation of amino acids is assisted by pyridoxal phosphate as a prosthetic group, whereas in the decarboxylation of pyruvate to acetaldehyde, thiamine pyrophosphate (TPP) plays that role. Oxidative decarboxylation, lastly, depends on the cooperation of no fewer than five cofactors thiamine pyrophosphate, lipoic acid, coenzyme A, flavin-adenine dinucleotide, and nicotinamide-adenine dinucleotide. 

The first step of this reaction, decarboxylation of pyruvate and transfer of the acetyl group to lipoic acid, depends on accumulation of negative charge on the carbonyl carbon of pyruvate. This is facilitated by the quaternary nitrogen on the thiazolium group of thiamine pyrophosphate. As shown in (c), this cationic... 

How many of the 14 NADPH needed to form one palmitate (Eq. 25.1) can be made in this way The answer depends on the status of malate. Every citrate entering the cytosol produces one acetyl-CoA and one malate (Figure 25.1). Every malate oxidized by malic enzyme produces one NADPH, at the expense of a decarboxylation to pyruvate. Thus, when malate is oxidized, one NADPH is produced for every acetyl-CoA. Conversion of 8 acetyl-CoA units to one palmitate would then be accompanied by production of 8 NADPH. (The other 6 NADPH required [Eq. 25.1] would be provided by the pentose phosphate pathway.) On the other hand, for every malate returned to the mitochondria, one NADPH fewer is produced. 

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). 

Biotin is involved in carboxylation and decarboxylation reactions. It is covalently bound to its enzyme. In the carboxylase reaction, C02 is first attached to biotin at the ureido nitrogen, opposite the side chain in an ATP-dependent reaction. The activated C02 is then transferred from carboxybiotin to the substrate. The four enzymes of the intermediary metabolism requiring biotin as a prosthetic group are pyruvate carboxylase (pyruvate oxaloacetate), propionyl-CoA-carboxylase (propionyl-CoA methylmalonyl-CoA), 3-methylcroto-nyl-CoA-carboxylase (metabolism of leucine), and actyl-CoA-carboxylase (acetyl-CoA malonyl-CoA) [1]. 

Biochemical reactions include several types of decarboxylation reactions as shown in Eqs. (1)-(5), because the final product of aerobic metabolism is carbon dioxide. Amino acids result in amines, pyruvic acid and other a-keto acids form the corresponding aldehydes and carboxylic acids, depending on the cooperating coenzymes. Malonyl-CoA and its derivatives are decarboxylated to acyl-CoA. -Keto carboxylic acids, and their precursors (for example, the corresponding hydroxy acids) also liberate carbon dioxide under mild reaction conditions. 

The first designed catalyst where there was some understanding of the relationship between structure and function was oxaldie 1, a 14-residue peptide that folds in solution to form helical bundles [11] (Fig. 12). Oxaldie 1 was designed to catalyze the decarboxylation of oxaloacetate, the a-keto acid of aspartic acid, via a mechanism where a primary amine reacts with the ketone carbonyl group to form a carbinolamine that is decarboxylated to form pyruvate. The reaction is piCj dependent and proceeds faster the lower the piC of the primary amine if the reaction is carried out at a pH that is lower than the piCj, of the reactive amine. The sequence contains five lysine residues that in the folded state form... 

Other organisms are equipped to produce ethanol, by employing a thiamine diphosphate-dependent decarboxylation of pyruvate to acetaldehyde (see Section 15.8) and NAD+ is regenerated by reducing the acetaldehyde to ethanol. This is a characteristic of baker s yeast, and forms the essential process for both bread making (production of CO2) and the brewing industry (formation of ethanol). 

The tricarboxylic acid cycle not only takes up acetyl CoA from fatty acid degradation, but also supplies the material for the biosynthesis of fatty acids and isoprenoids. Acetyl CoA, which is formed in the matrix space of mitochondria by pyruvate dehydrogenase (see p. 134), is not capable of passing through the inner mitochondrial membrane. The acetyl residue is therefore condensed with oxaloacetate by mitochondrial citrate synthase to form citrate. This then leaves the mitochondria by antiport with malate (right see p. 212). In the cytoplasm, it is cleaved again by ATP-dependent citrate lyase [4] into acetyl-CoA and oxaloacetate. The oxaloacetate formed is reduced by a cytoplasmic malate dehydrogenase to malate [2], which then returns to the mitochondrion via the antiport already mentioned. Alternatively, the malate can be oxidized by malic enzyme" [5], with decarboxylation, to pyruvate. The NADPH+H formed in this process is also used for fatty acid biosynthesis. 

Reddy et al. (1983) concluded that NO inactivation of iron-sulfur proteins was the probable mechanism of botulinal inhibition in nitrite-tteated foods. In support of this conclusion, Carpenter et al. (1987) observed decreased activity of clostridial pyruvate-ferredoxin oxidoteductase and lower cytochrome c reducing ability by ferredoxin in extracts of cells treated with nitrite. NO tteatment also inhibits yeast pyruvate decarboxylase (a non-iron-sulfur protein) and py-ruvate-ferredoxin oxidoteductase from C. perfringens (McMindes and Siedler, 1988). They suggested that thiamine-dependent decarboxylation of pyruvate may be an additional site for antimicrobial effects of NO. 

In a series of transition metal oxide semiconductor powders, photochemical activity in the decarboxylation of oxalic acid was controlled by surface properties and the presence of recombination centers, which in turn depended on the preparation method Similar effects have also been noted in the photodecarboxylation of pyruvic acid and formic acid... 

TPP-dependent enzymes catalyze either simple decarboxylation of a-keto acids to yield aldehydes (i.e. replacement of C02 with H+), or oxidative decarboxylation to yield acids or thioesters. The latter type of reaction requires a redox coenzyme as well (see below). The best known example of the former non-oxidative type of decarboxylation is the pyruvate decarboxylase-mediated conversion of pyruvate to acetaldehyde and C02. The accepted pathway for this reaction is shown in Scheme 10 (69MI11002, B-70MI11003, B-77MI11001>. 

Linked oxidation and decarboxylation. Metabolic pathways often make use of oxidation of a (3-hydroxy acid to a (3-oxoacid followed by decarboxylation in the active site of the same enzyme. An example is conversion of L-malate to pyruvate (Eq. 13-45). The Mg2+ or Mn2+-dependent decarboxylating malic dehydrogenase that catalyzes the reaction is usually called the malic enzyme. It is found in most organisms.237-240 While a concerted decarboxylation and dehydrogenation may sometimes occur,241-242 the enzymes of this group appear usually to operate with bound oxoacid intermediates as in Eq. 13-45. 

Biotin-dependent decarboxylases act as sodium ion pumps in Klebsiella74 and in various anaerobes.22 75 For example, oxaloacetate is converted to pyruvate and bound carboxybiotin.74 743 The latter is decarboxylated... 

Most known thiamin diphosphate-dependent reactions (Table 14-2) can be derived from the five halfreactions, a through e, shown in Fig. 14-3. Each halfreaction is an a cleavage which leads to a thiamin- bound enamine (center, Fig. 14-3) The decarboxylation of an a-oxo acid to an aldehyde is represented by step b followed by a in reverse. The most studied enzyme catalyzing a reaction of this type is yeast pyruvate decarboxylase, an enzyme essential to alcoholic fermentation (Fig. 10-3). There are two 250-kDa isoenzyme forms, one an a4 tetramer and one with an ( P)2 quaternary structure. The isolation of ohydroxyethylthiamin diphosphate from reaction mixtures of this enzyme with pyruvate52 provided important verification of the mechanisms of Eqs. 14-14,14-15. Other decarboxylases produce aldehydes in specialized metabolic pathways indolepyruvate decarboxylase126 in the biosynthesis of the plant hormone indoIe-3-acetate and ben-zoylformate decarboxylase in the mandelate pathway of bacterial metabolism (Chapter 25).1243/127... 

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