The Malonic Semialdehyde Pathways

The arguments in favor of the existence of the malonyl semialdehyde pathway outlined in the upper part of figure include the following observations the discovery of acrylyl CoA in bacteria the fact that acrylyl CoA is a good substrate for crotonase and the existence in bacteria and mammalian tissues of a specific hydroxypropionyl dehydrogenase. The enzyme is NAD-dependent for activity and has been purified 200 times. The product of the reaction is malonyl semialdehyde which can be either oxidized to malonate or split to yield acetyl-CoA and CO2. 

The most obvious route of metabolism of propionyl-CoA is further (1 oxidation which leads to 3-hydroxypropionyl-CoA (Fig. 17-3, step a). This appears to be the major pathway in green plants.17 Continuation of the (1 oxidation via steps a-c of Fig. 17-3 produces the CoA derivative of malonic semialdehyde. The latter can, in turn, be oxidized to malonyl-CoA, a P-oxoacid which can be decarboxylated to acetyl-CoA. The necessary enzymes have been found in Clostridium kluyveri,70 but the pathway appears to be little used. 

Nevertheless, malonyl-CoA is a major metabolite. It is an intermediate in fatty acid synthesis (see Fig. 17-12) and is formed in the peroxisomal P oxidation of odd chain-length dicarboxylic acids.703 Excess malonyl-CoA is decarboxylated in peroxisomes, and lack of the decarboxylase enzyme in mammals causes the lethal malonic aciduria.703 Some propionyl-CoA may also be metabolized by this pathway. The modified P oxidation sequence indicated on the left side of Fig. 17-3 is used in green plants and in many microorganisms. 3-Hydroxypropionyl-CoA is hydrolyzed to free P-hydroxypropionate, which is then oxidized to malonic semialdehyde and converted to acetyl-CoA by reactions that have not been completely described. Another possible pathway of propionate metabolism is the direct conversion to pyruvate via a oxidation into lactate, a mechanism that may be employed by some bacteria. Another route to lactate is through addition of water to acrylyl-CoA, the product of step a of Fig. 17-3. Tire water molecule adds in the "wrong way," the OH ion going to the a carbon instead of the P (Eq. 17-8). An enzyme with an active site similar to that of histidine ammonia-lyase (Eq. 14-48) could... 

The product of acetyl-CoA carboxylase reaction, malonyl-CoA, is reduced via malonate semialdehyde to 3-hydroxypropionate, which is further reductively converted to propionyl-CoA. Propionyl-CoA is carboxylated to (S)-methylmalonyl-CoA by the same carboxylase. (S)-Methylmalonyl-CoA is isomerized to (R)-methylmal-onyl-CoA, followed by carbon rearrangement to succinyl-CoA by coenzyme B 12-dependent methylmalonyl-CoA mutase. Succinyl-CoA is further reduced to succinate semialdehyde and then to 4-hydroxybutyrate. The latter compound is converted into two acetyl-CoA molecules via 4-hydroxybutyryl-CoA dehydratase, a key enzyme of the pathway. 4-Hydroxybutyryl-CoA dehydratase is a [4Fe-4S] cluster and FAD-containing enzyme that catalyzes the elimination of water from 4-hydroxybutyryl-CoA by a ketyl radical mechanism to yield crotonyl-CoA [34]. Conversion of the latter into two molecules of acetyl-CoA proceeds via normal P-oxidation steps. Hence, the 3-hydroxypropionate/4-hydroxybutyrate cycle (as illustrated in Figure 3.5) can be divided into two parts. In the first part, acetyl-CoA and two bicarbonate molecules are transformed to succinyl-CoA, while in the second part succinyl-CoA is converted to two acetyl-CoA molecules. 

CaaD is part of a pathway that is responsible for the degradation of the nematocide 1,3-dichloropropene in the soil bacterium Pseudomonas pavonaceae 170 (Figure l(b)). Its metabolic function is to convert trans-l>-chloroacrylate into malonate semialdehyde (4 at 3 s, 1.2 x lO moH Is ), which is probably the... 

Therefore, it is not surprising that several studies have reported successful 3-HP production by employing this pathway [27, 30-32, 35]. Overexpression of acetyl-CoA carboxylase encoded by the accABCD genes, and C. aurantiacus malonyl-CoA reductase encoded by the mcr gene in E. coli led to the production of 1.6 mM 3-HP. Malonyl-CoA reductase catalyzes the two-step conversion of malonyl-CoA to malonic-semialdehyde and further to 3-HP. It was expected that additional expression of E. coli pntAB genes, to meet the NADPH requirement... 

Figure 14.5 Malonyl-coA pathway for the production of 3-HP from glucose. Deletions and overexpression of genes beneficial for malonyl-coA pathway for the production of 3-HP from glucose [31, 32, 35]. Enzymes 1, acetyl-coA carboxylase [accABCD) 2, malonyl-coA reductase (mcr) 3, malonic semialdehyde reductase (msr) 4, lactalde-hyde [aldA) 5, (aldB) 6, [puuQ 7, lactate...

Pathways 8-10 are all thermodynamically favorable and produce 1 mol of ATP. Malonyl-CoA and malonic-semialdehyde can be derived from oxaloacetate by employing novel enzymes, with CoA-dependent oxaloacetate dehydrogenase and 2-keto acid decarboxylase activity, respectively. Malate can be converted to 3-HP using a novel enzyme with malate decarboxylase activity (Figure 14.4). These enzymes do not exist in nature and, because of this, it has been proposed that malate decarboxylase activity can be created by enzyme engineering in order to increase their specificity toward oxaloacetate and ability to produce the metabolic intermediates [33]. 

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