Biotin carboxylase

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 ... 

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. 

FIGURE 21-1 The acetyl-CoA carboxylase reaction. Acetyl-CoA carboxylase has three functional regions biotin carrier protein (gray) biotin carboxylase, which activates C02 by attaching it to a nitrogen in the biotin ring in an ATP-dependent reaction (see Fig. 16-16) and transcarboxylase, which transfers activated C02 (shaded green) from... 

The much studied E. coli enzyme is composed of a 156-residue biotin carboxyl carrier protein,38 a 449-residue biotin carboxylase, whose three-dimensional structure in known 39/393 and a carboxyltransferase subunit consisting of 304 (a)- and 319 (P)- residue chains. These all associate as a dimer of the three subunits (eight peptide chains).40-42... 

The biotin carboxyl carrier subunit of E. coli acetyl-CoA carboxylase contains the covalently bound biotin.55a/b The larger biotin carboxylase subunit catalyzes the ATP-dependent attachment of C02 to the biotin and the carboxyltransferase subunit catalyzes the final transcarboxylation step (Eq. 14-5, step b) by which acetyl-CoA is converted into malonyl-CoA. 

Carboxyphosphate. During the initial carboxyla-tion step lsO from labeled bicarbonate enters the P that is split from ATP. This suggested transient formation of carboxyphosphate by nucleophilic attack of HC03 on ATP (Eq. 14-7). The carboxyl group of this reactive mixed anhydride56 could then be transferred to biotin. This mechanism is supported by the fact that biotin carboxylase catalyzes the transfer of a... 

Biotin 515, 516,711, 721, 723 - 730,723s biosynthesis of 718, 745 in enzymes, table 724 mechanism of action 725 - 729 nutritional requirement 756 Biotin-binding proteins 728 Biotin carboxylase 724 Biotin carboxyl carrier protein 724 Biotin holoenzyme synthetase 724... 

Reactions catalyzed by acetyl-CoA carboxylase. In E. coli, BCCP and the two enzymatic activities (biotin carboxylase and carboxyltransferase) can be separated from each other. In contrast, in the liver all three components exist on a single multifunctional polypeptide. 

Acetyl-CoA carboxylase of E. coli is a multienzyme complex that consists of three protein components that can be isolated individually Biotin carboxyl carrier protein (BCCP), biotin carboxylase, and carboxyltransferase (fig. 

The reaction sequence involves an initial carboxyla-tion of BCCP, catalyzed by biotin carboxylase. The C02 is covalently linked to one of the nitrogen atoms of biotin (fig. 

Choi-Rhee, E. and Cronan, J. E. (2003) The biotin carboxylase-biotin carboxyl carrier protein complex of Escherichia coli acetyl-CoA carboxylase. J. Biol. Chem. 278, 30,806-30,812... 

Mammalian pyruvate carboxylase has four identical subunits, and the isolated monomer will catalyze the complete reaction. By contrast, three distinct subunits can be isolated from acetyl CoA carboxylase of Escherichia coli and spinach chloroplasts a biotinyl carrier protein, biotin carboxylase, and carboxyl transferase. 

Acetyl-CoA carboxylase is a biotin-dependent enzyme. It has been purified from microorganisms, yeast, plants, and animals. In animal cells, it exists as an inactive pro-tomer (M.W. 400,000) and as an active polymer (M.W. 4-8 million). The protomer contains the activity of biotin carboxylase, biotin carboxyl carrier protein (BCCP), transcarboxylase, and a regulatory allosteric site. Each protomer contains a biotinyl group bound in amide linkage to the e-amino group of a lysyl residue. 

Kondo, S., Nakajima, Y., Sugio, S., Yong-Biao, J., Sueda, S., and Kondo, H. (2004) Structur of the biotin carboxylase subunit of pyruvate cyarboxylase from Aquifex aeolicus at 2.2 A resolution. Acta Crystallogr. D,... 

Each ACC half-reaction is catalyzed by a different protein sub-complex. The vitamin biotin is covalently coupled through an amide bond to a lysine residue on biotin carboxyl carrier protein (BCCP, a homodimer of 16.7-kDa monomers encoded by accB) by a specific enzyme, biotin-apoprotein ligase (encoded by birA), and is essential to activity. The crystal and solution structures of the biotinyl domain of BCCP have been determined, and reveal a unique thumb required for activity (J. Cronan, 2001). Carboxylation of biotin is catalyzed by biotin carboxylase (encoded by accC), a homodimeric enzyme composed of 55-kDa subunits that is copurified complexed with BCCP. The accB and accC genes form an operon. The three-dimensional structure of the biotin carboxylase subunit has been solved by X-ray diffraction revealing an ATP-grasp motif for nucleotide binding. The mechanism of biotin carboxylation involves the reaction of ATP and CO2 to form the shortlived carboxyphosphate, which then interacts with biotin on BCCP for CO2 transfer to the I -nitrogen. 

Animal and fungal ACCs are comprised of large multifunctional polypeptides containing the biotin carboxylase, biotinyl carboxyl carrier protein, and carboxyltransferase... 

Fig. 1. Reactions catalyzed by acetyl-CoA carboxylase. BCCP, biotin carboxyl carrier protein BC, biotin carboxylase CT, carboxyl transferase.

Fig. 2. Acetyl-CoA carboxylase. (A) Eukaryotic ACCs contain -2300 residues organized into three functional domains — biotin carboxylase (BC), biotin carboxyl carrier protein (BCCP), and carboxyltransferase (CT). The role of the region between the biotin carboxyl carrier and carboxyltransferase domains is unknown. The biotin carboxyl carrier protein contains a typical conserved biotin attachment-site motif, VMKMV. The sites of phosphorylation are indicated by asterisks. (B) Electron micrograph of polymerized rat acetyl-CoA carboxylase (F. Ahmad, 1978). (C) Crystal structure of the biotin carboxylase domain of the yeast enzyme. In the presence of soraphen A, the biotin carboxyl carrier protein domain forms an inactive monomer the likely position of the modeled ATP-binding site is shown (adapted from Ref. [2]). (D) Crystal structure of the dimeric carboxyltransferase domain of the yeast enzyme. Although acetyl-CoA was included in the crystallization, density was observed only for CoA at one site and adenine at the other (adapted from Ref. [2]). (E) NMR structure of the biotin carboxyl carrier apoprotein domain of the human ACC2 The lysine attachment site for biotin is shown (RIKEN Structural Genomics/Proteomics Initiative, 2006). (See color plate section, plate no. 3.)...

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