Figure 1. Biosynthesis of 4¢-phosphopantetheine and its conversion into coenzyme A. Valine is initially transformed, in a three-reaction sequence, into pantoic acid. This is then linked to -alanine to give pantothenic acid (vitamin B₅) in a reaction with pantoyl adenylate as an enzyme-bound intermediate. Animals can phosphorylate pantothenic acid to 4¢-phosphopantothenic acid for conversion first into 4¢-phosphopantothenoyl cysteine and then, by decarboxylation, into 4¢-phosphopantetheine. Coenzyme A is synthesized by linking 4¢-phosphopantetheine to the nucleotide of ATP followed finally by phosphorylation of the resulting dephospho-coenzyme A to give coenzyme A.

Figure 2. Activation of fatty acids with ATP and CoA to give acyl-CoA derivatives. The enzyme reaction proceeds in two stages with the initial displacement of pyrophosphate from ATP by the acyl oxygen atom to form an enzyme–acyladenylate mixed anhydride complex. In the second stage, the thiolate anion of CoAS^– then reacts with the mixed anhydride, displacing AMP to form the thiol ester product.

Figure 3. Formation of an acyl-CoA by the coupled oxidative decarboxylation of an -keto acid. The reaction shown is catalysed by pyruvate dehydrogenase. Decarboxylation of the adduct formed between the carbanion of thiamin pyrophosphate and pyruvate generates an enamine that reacts with oxidized lipoamide to form a thiohemiacetal intermediate. This intermediate can then fragment to regenerate the thiamin pyrophosphate anion and the acetyl thiol ester of lipoamide. The transfer of the acetyl moiety to CoAS^– to give acetyl-CoA, followed by the reoxidation of the reduced lipoamide by NADH (not shown), completes the reaction. The formation of succinyl-CoA from 2-oxoglutarate, catalysed by 2-oxoglutarate dehydrogenase, proceeds along similar lines.

Figure 4. Reaction mechanism of CoA-transferase enzymes. Reaction of the donor acyl-CoA with a transferase glutamyl residue (Glu344 in succinyl-CoA transferase) results in the formation of a putative anhydride intermediate between the glutamyl carbonyl and the donor acyl group and displacement of CoAS^–. Reaction of CoAS^– with the glutamyl carbonyl of the anhydride displaces the donor acyl group as a free carboxylic acid (R¹) and results in the formation of an enzyme glutamyl-CoA thiol ester intermediate that may be isolated and characterized. The CoAS^– is then transferred from the enzyme–CoA intermediate to the acceptor carboxylic acid (R²), essentially by the reverse of the above process.

Figure 5. The two major classes of reaction in which acyl thiol esters of CoA and 4¢-phosphopantetheine participate. (a) An acyl thiol ester acting as an electrophile for acyl transfer to a nucleophile. The reaction proceeds by the initial reaction of the nucleophile X^– to form a tetrahedral intermediate from which the thiol is displaced to yield the acyl product RCOX. (b) An acyl thiol ester acting as a nucleophile. The ketonic properties of the thiol ester allow enolization to the thiohemiketal (shown also as a carbanion), which can act as a nucleophile, reacting typically with an electrophilic carbon atom to form a C–C bond.

Figure 6. Examples of reactions in which a CoA thiol ester acts as an electrophile in acyl transfer. (a) Acyl transfer to oxygen in choline acetyltransferase. (b) Acyl transfer to nitrogen as exemplified by N-acetyl-d-glucosamine-6-phosphate synthase. (c) Acyl transfer to carbon catalysed by a pyridoxal 5¢-phosphate-dependent enzyme catalysing -aminoketone biosynthesis where the nucleophile is the -carbanion of an amino acid stabilized by resonance with the positively charged pyridine ring nitrogen. In 5-aminolaevulinic acid synthase the glycine -carbanion reacts with succinyl-CoA. Enzymic decarboxylation of the resulting -keto acid and reprotonation yields 5-aminolaevulinic acid. The enzymic base HB-Enz is a lysine residue. (d) Acyl transfer to carbon catalysed by thiolase in which the acetyl group is transferred first to an enzyme thiol before reacting with an acetyl-CoA to yield acetoacetyl-CoA. (e) Succinyl-CoA catalysed acyl transfer to phosphate to form a mixed anhydride followed by phosphoryl transfer to GDP via an enzyme N-phosphohistidine. (f) Acyl-CoA reductase-catalysed reduction of acetyl-CoA to yield a thiohemiacetal intermediate that eliminates CoAS^– to yield acetaldehyde.

Figure 7. Examples of reactions in which a CoA thiol ester acts as a nucleophile. (a) Citrate synthase involves the nucleophilic addition of the carbanion of acetyl-CoA to the si face of acetoacetate to yield S-citryl-CoA and finally citrate. (b) Biotin-dependent carboxylation catalysed by acetyl-CoA carboxylase utilizes the carbanion of acetyl-CoA, which reacts with carboxybiotin to yield malonyl-CoA. (c) Thiolase-catalysed formation of acetoacetyl-CoA nucleophilic reaction of the carbanion of acetyl-CoA with an enzyme-bound acetyl thiol ester. (d) Coupled decarboxylation and nucleophilic addition of C2 of malonyl-CoA to any of a number of acyl thiol esters at the active site of polyketide and fatty acid synthases. (e) Utilization of the acidic nature of protons at the -position of thiol esters (i) to assist in -oxidation of fatty acids and (ii) in the dehydration and allylic rearrangement catalysed by -hydroxydecanoyl thiol ester dehydratase. B represents a catalytic enzyme group.

Figure 8. Synthesis of chalcone and resveratrol. These enzymes function in a similar way to other polyketide synthases, both using one acyl-CoA and three malonyl-CoA units, but contain no acyl carrier protein functionality using the acyl-CoA thiol ester itself. The initial stages are common to both enzymes but the final folding of intermediates gives the different end products.