Fatty acid-condensing enzyme

INHIBITION OF FATTY ACID CONDENSING ENZYMES IN PLANTS... 

Snapp, A.R., Kang, J., Qi, X., Lu, C., 2014. A fatty acid condensing enzyme from Physaria fendleri increases hydroxy fatty acid accumulation in transgenic oilseeds of Camelirta saliva. Planta 240, 599-610. 

Acetyl-CoA is the source of the carbon atoms for the synthesis of cholesterol, steroid hormones, and fatty acids. Various enzymes catalyze biological versions of aldol reactions and Claisen condensations during the syntheses of these compounds. 

Once condensation has taken place, the /3-ketoacyl-ACP undergoes a cycle of reduction, dehydration, and reduction to yield a new fatty acid. The enzymes concerned are )8-ketoacyl-ACP reductase, )8-hydroxyacyl-ACP dehydrase, and enoyl reductase, and detailed information is provided in recent reviews. Both of the reductases have been reported in isoforms from several tissues. Thus, in avocado, an NADH form of )8-ketoacyl-ACP reductase has been resolved from an NADPH form. In seed tissues, two forms of enoyl reductase appear to be present they differ in the chain-length activity and also in whether they use NADH only or use NADPH in preference to NADH only the NADH form has been purified from leaves. ... 

It is also worth noting that the carbon of the carboxyl group that was added to drive this reaction is the one removed by the condensing enzyme. Thus, all the carbons of acetoacetyl-ACP (and of the fatty acids to be made) are derived from acetate units of acetyl-CoA. 

Citrate is isomerized to isocitrate by the enzyme aconitase (aconitate hydratase) the reaction occurs in two steps dehydration to r-aconitate, some of which remains bound to the enzyme and rehydration to isocitrate. Although citrate is a symmetric molecule, aconitase reacts with citrate asymmetrically, so that the two carbon atoms that are lost in subsequent reactions of the cycle are not those that were added from acetyl-CoA. This asymmetric behavior is due to channeling— transfer of the product of citrate synthase directly onto the active site of aconitase without entering free solution. This provides integration of citric acid cycle activity and the provision of citrate in the cytosol as a source of acetyl-CoA for fatty acid synthesis. The poison fluo-roacetate is toxic because fluoroacetyl-CoA condenses with oxaloacetate to form fluorocitrate, which inhibits aconitase, causing citrate to accumulate. 

Nature gives us some illustrative examples of iterative methodologies in its biochemical mechanisms. The fatty acid-polyketide biosynthesis is one of them. The assembly of acyl units by sequential Claisen-type condensations to form a polyketide or fatty acid takes place at a multi-enzyme complex, at which the initial molecule is lengthened by one C2-unit per pass of a reaction cycle (Fig. 2). 

The first formation of a carbon-carbon bond occurs between malonyl and acetyl units bound to fatty acid synthase. After reduction, dehydration, and further reduction, the acyl enzyme is condensed with more malonyl-CoA and the cycle is repeated until the acyl chain grows to C16. When the growing fatty acid reaches a chain length of 16 carbons, the acyl group is hydrolyzed to give the free fatty acid. 

Each CHS monomer consists of two structural domains (Fig. 12.5, left). The upper domain exhibits the a-p-a-p-a pseudo-symmetric motif observed in fatty acid P-ketoacyl synthases (KASs) (Fig. 12.5, right).20 Both CHS and KAS use a cysteine as a nucleophile in the condensation reaction, and shuttle reaction intermediates via CoA thioester-linked molecules or ACPs, respectively. The conserved architecture of the upper domain maintains the three-dimensional position of the catalytic residues of each enzyme Cysl64, His303, and Asn336 in CHS correspond to a Cys, His, and His in KAS I and II. 

SCHRODER, J., The chalcone/stilbene synthase-type family of condensing enzymes. In Comprehensive Natural Products Chemistry, vol. 1, Polyketides and Other Secondary Metabolites Including Fatty Acids and Their Derivatives (U. Sankawa ed.), Elsevier, Amersterdam, 1999, pp. 749-771. 

By 1960 it was clear that acetyl CoA provided its two carbon atoms to the to and co—1 positions of palmitate. All the other carbon atoms entered via malonyl CoA (Wakil and Ganguly, 1959 Brady et al. 1960). It was also known that 3H-NADPH donated tritium to palmitate. It had been shown too that fatty acid synthesis was very susceptible to inhibition by p-hydroxy mercuribenzoate, TV-ethyl maleimide, and other thiol reagents. If the system was pre-incubated with acetyl CoA, considerable protection was afforded against the mercuribenzoate. In 1961 Lynen and Tada suggested tightly bound acyl-S-enzyme complexes were intermediates in fatty acid synthesis in the yeast system. The malonyl-S-enzyme complex condensed with acyl CoA and the B-keto-product reduced by NADPH, dehydrated, and reduced again to yield the (acyl+2C)-S-enzyme complex. Lynen and Tada thought the reactions were catalyzed by a multifunctional enzyme system. 

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. 

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