Biotin carboxylase domain

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.)...

Acetyl-CoA carboxylase (ACC) catalyzes the first committed step in long-chain fatty acid biosynthesis (see Chapter 7.11). The overall reaction is catalyzed in two sequential reactions (Scheme 3). First, the biotin carboxylase domain catalyzes the ATP-dependent carboxylation of biotin (which is attached to a carrier protein) using bicarbonate as a CO2 donor. In the second reaction, the carboxyl group is transferred from biotin to acetyl-CoA to form malonyl-CoA. In mammals, both reactions are catalyzed by a single protein, but in Escherichia coli and other bacteria, the activity is catalyzed by two separate proteins, a biotin carboxylase and a carboxytransferase. Due to its role in fatty acid synthesis, inhibitors of the overall ACC reaction are proposed to be useful as antiobesity drugs in mammals as well as novel antibiotics against bacteria. 

Figure 16.25. Domain Structure of Pyruvate Carboxylase. The ATP-grasp domain activates HCO3 and transfers CO2 to the biotin-binding domain. From there, the CO2 is transferred to pyruvate generated in the central domain.

Figure 16.26. Biotin-Binding Domain of Pyruvate Carboxylase. This likely structure is based on the structure of the homologous domain from the enzyme acetyl CoA carboxylase (Section 22.4.1). The biotin is on a flexible tether, allowing it to move between the ATP-bicarbonate site and the pyruvate site.

The core of the complex is formed hy E2. Acetyltransferase consists of eight catalytic trimers assembled to form a hollow cube. Each of the three subunits forming a trimer has three major domains (Figure 17.8). At the amino terminus is a small domain that contains a bound lipoamide cofactor attached to a lysine residue. This domain is homologous to biotin-binding domains such as that of pyruvate carboxylase (see Figure 16.26). The lipoamide domain is followed by a small domain that interacts with E3 within the complex. A larger transacetylase domain completes an E2 subunit. Ej is an... 

Biotin-binding domain of pyruvate carboxylase Figure 16.24... 

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. 

In prokaryotes and in plastids of some plants, the ACC is a multisubunit enzyme, whereas in eukaryotes the cytosolic isozyme and, in some instances also the plastid isozyme, are multidomain proteins. The latter contain three major functional domains, which account for the biotin carboxylase (BT), biotin carboxyl-carrier (BCC) and carboxyltransferase (CT) activities and, which are organized in one large polypeptide. 

Fig. 9.2. Schematic representation of a plastidic homomeric acetyl coenzyme A carboxylase (ACC) showing the three functional domains (BC, biotin carboxylase BCC, biotin carboxyl-carrier and CT, carboxyl transferase) and the transit peptide (TP) that...

In animal systems, the enzyme exists as a polyfunctional inactive protein with a MW of about 240,000 (Hardie and Cohen, 1978 Tanabe et al., 1975 MacKall and Lane, 1977), which consists of three domains—biotin carboxylase, BCCP, and transcarboxylase. These are linked by an exposed polypeptide chain peculiarly susceptible to attack by endogenous proteases. In the presence of citrate this protomer rapidly aggregates to polymers of 7-10 million daltons, and these polymers are active forms of the enzyme. These polymers rapidly depolymerize in the presence of low concentrations of palmi-toyl-CoA to the inactive protomer. In addition, there is increasing evidence that a phosphorylation-dephosphorylation cycle is involved in further modulating the activity of this important enzyme (Hardie and Cohen, 1978 Lee and Kim, 1977). 

Song, J., Wurtele, E.S., and Nikolau B.J. (1994) Molecular cloning and characterization of the cDNA coding for die biotin-containing subunit of 3-methylcrotonyl-CoA carboxylase Identification of the biotin carboxylase and ciotin-carrier domains. Proc. Nat. Acad. Sci. U.S.A. 91, 5779-5783. 

Acetyl-CoA carboxylase (ACCase) catalyzes the synthesis of malonyl-CoA, the first intermediate in fatty acid synthesis. There are two forms of ACCase a prokaryote form consisting of three protein components, biotin carboxylase, carboxyltransferase, and biotin carboxylase carrier protein, and a eukaryote form consisting of three functional domains on a single polypeptide. About 20 years ago, Kannangara and Stumpf reported the existence of the prokaryote form in spinach chloroplasts [1], the major site of fatty acid synthesis, but this finding has been dismissed because the prokaryote form has not yet been purified and the purified ACCases from various plants are all eukaryote form consisting of a subunit size of about 200 kDa, like that of the mammal enzyme. 

Repressors may have similar recognition domains but may vary greatly both in size and in the functioning of fheir other domains, which may react both with small allosteric effectors and with other proteins. The repressor BirA of the E. coli biotin synthesis operon is an enzyme. The 321-residue protein activates biohn to form biotinyl 5 -adenylafe and transfers the biotinyl group to proteins such as acetyl-CoA carboxylase ° ° and also represses transcriphon. 

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