Acetyl-CoA carboxylases

Zn-+ DNA polymerase Coenzyme A (CoA) Acyl groups Acetyl-CoA carboxylase... 

Rittenberg and Bloch showed in the late 1940s that acetate units are the building blocks of fatty acids. Their work, together with the discovery by Salih Wakil that bicarbonate is required for fatty acid biosynthesis, eventually made clear that this pathway involves synthesis of malonyl-CoA. The carboxylation of acetyl-CoA to form malonyl-CoA is essentially irreversible and is the committed step in the synthesis of fatty acids (Figure 25.2). The reaction is catalyzed by acetyl-CoA carboxylase, which contains a biotin prosthetic group. This carboxylase is the only enzyme of fatty acid synthesis in animals that is not part of the multienzyme complex called fatty acid synthase. 

Acetyl-CoA Carboxylase Is Biotin-Dependent and Displays Ping-Pong Kinetics... 

In animals, acetyl-CoA carboxylase (ACC) is a filamentous polymer (4 to 8 X 10 D) composed of 230-kD protomers. Each of these subunits contains the biotin carboxyl carrier moiety, biotin carboxylase, and transcarboxylase activities, as well as allosteric regulatory sites. Animal ACC is thus a multifunctional protein. The polymeric form is active, but the 230-kD protomers are inactive. The activity of ACC is thus dependent upon the position of the equilibrium between these two forms ... 

FIGURE 25.2 (a) The acetyl-CoA carboxylase reaction produces malonyl-CoA for fatty acid synthesis, (b) A mechanism for the acetyl-CoA carboxylase reaction. Bicarbonate is activated for carboxylation reactions by formation of N-carboxybiotin. ATP drives the reaction forward, with transient formation of a carbonylphosphate intermediate (Step 1). In a typical biotin-dependent reaction, nncleophilic attack by the acetyl-CoA carbanion on the carboxyl carbon of N-carboxybiotin—a transcarboxylation—yields the carboxylated product (Step 2). 

Because this enzyme catalyzes the committed step in fatty acid biosynthesis, it is carefully regulated. Palmitoyl-CoA, the final product of fatty acid biosynthesis, shifts the equilibrium toward the inactive protomers, whereas citrate, an important allosteric activator of this enzyme, shifts the equilibrium toward the active polymeric form of the enzyme. Acetyl-CoA carboxylase shows the kinetic behavior of a Monod-Wyman-Changeux V-system allosteric enzyme (Chapter 15). 

The regulatory effects of citrate and palmitoyl-CoA are dependent on the phosphorylation state of acetyl-CoA carboxylase. The animal enzyme is phosphorylated at 8 to 10 sites on each enzyme subunit (Figure 25.4). Some of these sites are reg-... 

FIGURE 25.3 In the acetyl-CoA carboxylase reaction, the biotin ring, on its flexible tether, acquires carboxyl groups from carbonylphos-phate on the carboxylase subunit and transfers them to acyl-CoA molecules on the transcarboxylase subunits. 

FIGURE 25.4 Models of the acetyl-CoA carboxylase polypeptide, with phosphorylation sites indicated, along with the protein kinases responsible. Phosphorylation at Ser " is primarily responsible for decreasing the affinity for citrate. 

Dephospho-acetyl-CoA carboxylase (Low [citrate] activates, high [fatty acyl-CoA] inhibits)... 

FIGURE 25.5 The activity of acetyl-CoA carboxylase is modulated by phosphorylation and dephosphoryladon. The dephospho form of the enzyme is activated by low [citrate] and inhibited only by high levels of fatty acyl-CoA. 

FIGURE 25.17 Hormonal signals regulate fatty acid synthesis, primarily through actions on acetyl-CoA carboxylase. Availability of fatty acids also depends upon hormonal activation of triacylglycerol lipase. 

Based on the information presented in the text and in Figures 25.4 and 25.5, suggest a model for the regulation of acetyl-CoA carboxylase. Consider the possible roles of subunit interactions, phosphorylation, and conformation changes in your model. 

Lopa.schnk, G. D., and Gamble, J., 1994. The 1993 Merck Fro.s.st Award. Acetyl-CoA carboxylase an important regulator of fatty acid oxidation in die heart. Canadian Journal of Physiology and Pharmacology 72 1101 — 1109. 

Fatty acid oxida- T Acetyl-CoA carboxylase-2 l Activity, J, malonyl-CoA Liver, muscle,... 

Fatty acid synthesis 1 Acetyl-CoA carboxylase-1 l Activity All cells ... 

Acetyl-CoA carboxylase T Insulin Citrate, insulin Long-chain acyl-CoA, cAMP, glucagon... 

Bicarbonate as a source of CO2 is required in the initial reaction for the carboxylation of acetyl-CoA to mal-onyl-CoA in the presence of ATP and acetyl-CoA carboxylase. Acetyl-CoA carboxylase has a requirement for the vitamin biotin (Figure 21-1). The enzyme is a multienzyme protein containing a variable number of identical subunits, each containing biotin, biotin carboxylase, biotin carboxyl carrier protein, and transcarboxylase, as well as a regulatory allosteric site. The reaction takes place in two steps (1) carboxylation of biotin involving ATP and (2) transfer of the carboxyl to acetyl-CoA to form malonyl-CoA. 

Acetyl-CoA Carboxylase Is the Most Important Enzyme in the Regulation of Lipogenesis... 

Acetyl-CoA carboxylase is an allosteric enzyme and is activated by citrate, which increases in concentration in the well-fed state and is an indicator of a plentiful supply of acetyl-CoA. Citrate converts the enzyme from an inactive dimer to an active polymeric form, having a molecular mass of several milhon. Inactivation is promoted by phosphorylation of the enzyme and by long-chain acyl-CoA molecules, an example of negative feedback inhibition by a product of a reaction. Thus, if acyl-CoA accumulates because it is not esterified quickly enough or because of increased lipolysis or an influx of free fatty acids into the tissue, it will automatically reduce the synthesis of new fatty acid. Acyl-CoA may also inhibit the mitochondrial tricarboxylate transporter, thus preventing activation of the enzyme by egress of citrate from the mitochondria into the cytosol. 

Acetyl-CoA carboxylase is also regulated by hormones such as glucagon, epinephrine, and insulin via... 

Figure 21-6. Regulation of acetyl-CoA carboxylase by phosphorylation/dephosphorylation.The enzyme is inactivated by phosphorylation by AMP-activated protein kinase (AMPK), which in turn is phosphorylated and activated by AMP-activated protein kinase kinase (AMPKK). Glucagon (and epinephrine), after increasing cAMP, activate this latter enzyme via cAMP-dependent protein kinase. The kinase kinase enzyme is also believed to be activated by acyl-CoA. Insulin activates acetyl-CoA carboxylase, probably through an "activator" protein and an insulin-stimulated protein kinase.

Insulin stimulates lipogenesis by several other mechanisms as well as by increasing acetyl-CoA carboxylase activity. It increases the transport of glucose into the cell (eg, in adipose tissue), increasing the availability of both pyruvate for fatty acid synthesis and glycerol 3-phosphate for esterification of the newly formed fatty acids, and also converts the inactive form of pyruvate dehydrogenase to the active form in adipose tissue but not in liver. Insulin also—by its ability to depress the level of intracellular cAMP—inhibits lipolysis in adipose tissue and thereby reduces the concentration of... 

The Fatty Acid Synthase Complex Acetyl-CoA Carboxylase Are Adaptive Enzymes... 

The synthesis of long-chain fatty acids (lipogenesis) is carried out by two enzyme systems acetyl-CoA carboxylase and fatty acid synthase. 

Acetyl-CoA carboxylase is required to convert acetyl-CoA to malonyl-CoA. In turn, fatty acid synthase, a multienzyme complex of one polypeptide chain with seven separate enzymatic activities, catalyzes the assembly of palmitate from one acetyl-CoA and seven malonyl-CoA molecules. 

Lipogenesis is regulated at the acetyl-CoA carboxylase step by allosteric modifiers, phosphorylation/de-phosphorylation, and induction and repression of enzyme synthesis. Citrate activates the enzyme, and long-chain acyl-CoA inhibits its activity. Insulin activates acetyl-CoA carboxylase whereas glucagon and epinephrine have opposite actions. 

Biotin functions to transfer carbon dioxide in a small number of carboxylation reactions. A holocarboxylase synthetase acts on a lysine residue of the apoenzymes of acetyl-CoA carboxylase, pymvate carboxylase, propi-onyl-CoA carboxylase, or methylcrotonyl-CoA carboxylase to react with free biotin to form the biocytin residue of the holoenzyme. The reactive intermediate is 1-7V-carboxybiocytin, formed from bicarbonate in an ATP-dependent reaction. The carboxyl group is then transferred to the substrate for carboxylation (Figure 21—1). 

The reaction is catalyzed by the biotin,enzyme acetyl-CoA carboxylase (E-biotin) assisted by Mg2+ ions. This enzyme is a tetramer with a molecular mass of 400 000-500000. 

Jin, M., Fischbach, M.A. and Clardy, J. (2006) A biosynthetic gene cluster for the acetyl-CoA carboxylase inhibitor andrimid. Journal of the American Chemical Society, 128, 10660-10661. 

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