Acetaldehyde activated

The reaction has been shown to be carried out by a pyravate decarboxylase and involves thiamine pyrophosphate in the formation of activated acetaldehyde from pyravate, which then condenses with benzaldehyde. Evidently, pyravate decarboxylase, a crucial enzyme for ethanol biosynthesis, is nsed in an urmatural way... 

Another example is the commercialized formation of R-phenylacetylcarbinol (R-PAC) from benzaldehyde and pyravate by fermenting brewer s yeast (see chapter 4.16 Neuberg Hirsch, 1921). Pymvate decarboxylase is responsible for the formation of an activated acetaldehyde from the decarboxylation of pyravate and the condensation of this activated acetaldehyde with benzaldehyde. Nowadays, about 1000 tons per aimum (tpa) R-PAC is produced this way. 

A patented process for the production of green notes applying bakers yeast for in situ reduction of enzymatically produced aldehydes [67, 68] has been called into question regarding the effective production of (Z)-3-hexenol. According to Gatfield s report [69] the isomerisation of (Z)-3-hexenol to (E)-2-hexenal is a very fast process. The latter undergoes facile conversion to hexanol. Beside this, baker s yeast can add activated acetaldehyde to ( )-2-hexenal, forming 4-octen-2,3-diol. 

Thiamine Pyrophosphate Carries Active Acetaldehyde Groups... 

Thiamine pyrophosphate plays an important role in the cleavage of bonds adjacent to a carbonyl group, such as the decarboxylation of a-lceto acids, and in chemical rearrangements in which an activated acetaldehyde group is transferred from one carbon atom to another (Table 14-1). The functional part of TPP, the thiazolium ring, has a relatively acidic proton at C-2. Loss of this... 

Formation of a-ketols from a-oxo acids also starts with step b of Fig. 14-3 but is followed by condensation with another carbonyl compound in step c, in reverse. An example is decarboxylation of pyruvate and condensation of the resulting active acetaldehyde with a second pyruvate molecule to give R-a-acetolactate, a reaction catalyzed by acetohydroxy acid synthase (acetolactate synthase).128 Acetolactate is the precursor to valine and leucine. A similar ketol condensation, which is catalyzed by the same synthase, is... 

Ketols can also be formed enzymatically by cleavage of an aldehyde (step a, Fig. 14-3) followed by condensation with a second aldehyde (step c, in reverse). An enzyme utilizing these steps is transketolase (Eq. 17-15),132b which is essential in the pentose phosphate pathways of metabolism and in photosynthesis. a-Diketones can be cleaved (step d) to a carboxylic acid plus active aldehyde, which can react either via a or c in reverse. These and other combinations of steps are often observed as side reactions of such enzymes as pyruvate decarboxylase. A related thiamin-dependent reaction is that of pyruvate and acetyl-CoA to give the a-diketone, diacetyl, CH3COCOCH3.133 The reaction can be viewed as a displacement of the CoA anion from acetyl-CoA by attack of thiamin-bound active acetaldehyde derived from pyruvate (reverse of step d, Fig. 14-3 with release of CoA). 

Although the direct reaction of a lipoyl group with the thiamin-bound enamine (active aldehyde) is generally accepted, and is supported by recent studies,3153 an alternative must be considered.315 Hexacyanoferrate (III) can replace NAD+ as an oxidant for pyruvate dehydrogenase and is also able to oxidize nonenzymatically thiamin-bound active acetaldehyde... 

The first step in valine biosynthesis is a condensation between pyruvate and active acetaldehyde (probably hy-droxyethyl thiamine pyrophosphate) to yield a-acetolactate. The enzyme acetohydroxy acid synthase usually has a requirement for FAD, which, in contrast to most flavopro-teins, is rather loosely bound to the protein. The very same enzyme transfers the acetaldehyde group to a-ketobutyrate to yield a-aceto-a-hydroxybutyrate, an isoleucine precursor. Unlike pyruvate, the a-ketobutyrate is not a key intermediate of the central metabolic routes rather it is produced for a highly specific purpose by the action of a deaminase on L-threonine as shown in figure 21.10. 

First steps to elucidate the reaction mechanism of PDC were achieved by the investigation of model reactions using ThDP or thiamine [36,37], Besides the identification of C2-ThDP as the catalytic center of the cofactor [36], the mechanism of the ThDP-catalyzed decarboxylation of a-keto acids as well as the formation of acyloins was explained by the formation of a common reaction intermediate, active acetaldehyde . This active species was first identified as HEThDP 7 (Scheme 3) [38,39]. Later studies revealed the a-carbanion/enamine 6 as the most likely candidate for the active acetaldehyde [40 47] (for a comprehensive review see [48]). The relevance of different functional groups in the ThDP-molecule for the enzymatic catalysis was elucidated by site-directed substitutions of the cofactor ThDP by chemical means (for a review see... 

Meaden, P. G., Dickinson, F. M., Mifsud, A., Tessier, W., Westwater, J., Bussey, H., Midgley, M. (1997) The ALD6 gene of Saccharomyces cerevisiae encodes a cytosolic, Mg -activated acetaldehyde dehydrogenase. Yeast, 13, 1319-1327. 

Another pharmaceutical administered as a racemic mixture to treat liver diseases is a-Iipoic acid (Thioctacid). Practically nothing is known about the activity of the individual enantiomers, although naturally occur-ring (+)-a-lipoic acid has an R-configuration. The first enantiomer separation of a-lipoic acid methyl ester was demonstrated on colunms with Lipodex D (Fig. 12,46). a-Lipoic acid is involved in the oxidative decarboxylation of pyruvic add to activated acetaldehyde, which is transferred to coenz3une A to form acetyl coenzyme A. 

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 half-reaction 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 h followed by fl 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 tetramer and one with an (aP)2 quaternary structure. The isolation of a-hydroxyethylthiamin diphosphate from reaction mixtures of this enzyme with pyruvate provided important verification of the mechanisms of Eqs. 14-14,14-15. Other decarboxylases produce aldehydes in specialized metabolic pathways indolepyruvate decarboxylase in the biosynthesis of the plant hormone indole-3-acetate and ben-zoylformate decarboxylase in the mandelate pathway of bacterial metabolism (Chapter 25). Formation of a-ketols from a-oxo acids also starts with step h of Fig. 14-3 but is followed by condensation with another carbonyl compound in step c, in reverse. An example is decarboxylation of pyruvate and condensation of the resulting active acetaldehyde with a second pyruvate molecule to give l -a-acetolactate, a reaction catalyzed by acetohydroxy acid synthase (acetolactate synthase). Acetolactate is the precursor to valine and leucine. A similar ketol condensation, which is catalyzed by the same S5mthase, is... 

Protonation of the eneamine to form an active acetaldehyde called hydroxyethyl-TPP. 

The active acetaldehyde can then be oxidized (as in the pyruvate dehydrogenase and oi-ketoglutarate dehydrogenase complexes) or an elimination reaction (non-oxidative) can occur. In either case, the decarboxylated compound is released, yielding free TPP. 

The formation of IPP/DMAPP via an alternative, nonmevalonate, pathway has been described (93). In this pathway, a TPP-activated acetaldehyde (generated by pyruvate decarboxylation) is coupled to the C-2 carbonyl... 

McNelis, 1959). This subject has been reviewed recently by Metzler (1960) and will not be discussed in detail here. In brief, Breslow postulated that thiamine pyropho.sphate ionizes at the 2-position of the thiazole ring, and that the thiazolium dipolar ion (IV) reacts with pyruvate to form an intermediate (2-laetylthiamine pyrophosphate) (V) which undergoes decarboxylation to produce 2-hydroxyethylthiamine pyrophosphate (VI, VII) (Fig. 2). Species (VI) is regarded as active acetaldehyde, and can... 

Acetylthiamine pjrrophosphate appears to be yet another form of active acetate. It has been assigned a key role in the lipoic acid-Unked oxidative decarboxylation of pyruvate as the primary product of the oxidation of active acetaldehyde, i.e., 2-hydroxyethylthiamine pyrophosphate. It has been proposed that 2-acetylthiamine pyrophosphate is an intermediate in all oxidative transformations of pyruvate and that 2-succinylthiamine pyrophosphate plays a similar role in oxidation of a-ketoglutarate. Further evaluation of this proposal is anticipated in the near future. 

However, failing incoporations of C-labeled aeetate and sueeessful ones of Relabeled glycerol as well as pyruvate in hopanes and ubiquinones showed isopen-tenyldiphosphate (IPP) to originate not only from the acetate mevalonate pathway, but also from activated acetaldehyde (C2, by reaction of pyruvate and thiamine diphosphate) and glyceraldehyde-3-phosphate (C3) R. In this way, 1-deoxy-pentulose-5-phosphate is generated as the first unbranched C5 preeursor of IPP. 

Quite similar considerations apply to the other types of thiamine-catalyzed reactions. In particular, the structure of active acetaldehyde is sinular to that of active benzaldehyde . 

Diacetyl can be produced by either homolactic or heterolactic pathways of sugar metabolism (via free pyruvate) or by utilization of citric acid (see Figs. 1-1 lA and 1-1 IB). In this case, citric acid is first converted to oxaloacetic and acetic acids. The former is then decarboxylated to pyruvate which undergoes a second decarboxylation and condensation with thiamine pyrophosphate (TPP) to yield active acetaldhyde, which reacts with another pyruvate to yield a-acetolactate which undergoes oxidative decarboxylation to yield diacetyl and its equilibrium products see Fig. 1-11 A. In the case of other LAB, the precursor, a-acetolactate is not produced. Here active acetaldehyde, produced as described above, reacts with acetyl CoA to yield diacetyl see Fig. 1-1 IB. 

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