Search for novel targets of the PII signal transduction protein in Bacteria identifies the BCCP component of acetyl‐CoA carboxylase as a PII binding partner

The P_II family comprises a group of widely distributed signal transduction proteins. The archetypal function of P_II is to regulate nitrogen metabolism in bacteria. As P_II can sense a range of metabolic signals, it has been suggested that the number of metabolic pathways regulated by P_II may be much greater than described in the literature. In order to provide experimental evidence for this hypothesis a P_II protein affinity column was used to identify P_II targets in Azospirillum brasilense. One of the P_II partners identified was the biotin carboxyl carrier protein (BCCP), a component of the acetyl‐CoA carboxylase which catalyses the committed step in fatty acid biosynthesis. As BCCP had been previously identified as a P_II target in Arabidopsis thaliana we hypothesized that the P_II–BCCP interaction would be conserved throughout Bacteria. In vitro experiments using purified proteins confirmed that the P_II–BCCP interaction is conserved in Escherichia coli. The BCCP–P_II interaction required MgATP and was dissociated by increasing 2‐oxoglutarate. The interaction was modestly affected by the post‐translational uridylylation status of P_II; however, it was completely dependent on the post‐translational biotinylation of BCCP.     

A multisubunit acetyl coenzyme A carboxylase from soybean

A multisubunit form of acetyl coenzyme A (CoA) carboxylase (ACCase) from soybean (Glycine max) was characterized. The enzyme catalyzes the formation of malonyl CoA from acetyl CoA, a rate-limiting step in fatty acid biosynthesis. The four known components that constitute plastid ACCase are biotin carboxylase (BC), biotin carboxyl carrier protein (BCCP), and the alpha- and beta-subunits of carboxyltransferase (alpha- and beta-CT). At least three different cDNAs were isolated from germinating soybean seeds that encode BC, two that encode BCCP, and four that encode alpha-CT. Whereas BC, BCCP, and alpha-CT are products of nuclear genes, the DNA that encodes soybean beta-CT is located in chloroplasts. Translation products from cDNAs for BC, BCCP, and alpha-CT were imported into isolated pea (Pisum sativum) chloroplasts and became integrated into ACCase. Edman microsequence analysis of the subunits after import permitted the identification of the amino-terminal sequence of the mature protein after removal of the transit sequences. Antibodies specific for each of the chloroplast ACCase subunits were generated against products from the cDNAs expressed in bacteria. The antibodies permitted components of ACCase to be followed during fractionation of the chloroplast stroma. Even in the presence of 0.5 M KCl, a complex that contained BC plus BCCP emerged from Sephacryl 400 with an apparent molecular mass greater than about 800 kD. A second complex, which contained alpha- and beta-CT, was also recovered from the column, and it had an apparent molecular mass of greater than about 600 kD. By mixing the two complexes together at appropriate ratios, ACCase enzymatic activity was restored. Even higher ACCase activities were recovered by mixing complexes from pea and soybean. The results demonstrate that the active form of ACCase can be reassembled and that it could form a high-molecular-mass complex.     

The biotin carboxylase-biotin carboxyl carrier protein complex of Escherichia coli acetyl-CoA carboxylase

Escherichia coli acetyl-CoA carboxylase (ACC) is composed of four different protein molecules. These proteins form a large but very unstable complex. Hints of a sub-complex between the biotin carboxylase (BC) and biotin carboxyl carrier protein (BCCP) subunits have been reported in the literature, but the complex was not isolated and thus the protein stoichiometry could not be determined. We report isolation of the BC.BCCP complex. By use of affinity chromatography using two different affinity tags it was shown that the complex consists of a two BCCP molecules per BC molecule. The molar ratio in the complex is the same as the ratio of the subunit proteins synthesized in vivo. We conclude that the complex consists of a dimer of BC plus four BCCP molecules instead of the 2BC.2BCCP complex previously assumed. This subunit ratio allows two conflicting models of the ACC mechanism to be rectified. We also report that the N-terminal 30 or so residues of BCCP are responsible for the interaction of BCCP with BC and that the BC.BCCP complex is a substrate for biotinylation in vitro.     

Co-purification,co-imniunoprecipitation,and coordinate expression of acetyl-coenzyme A carboxylase activity,biotin carboxylase,and biotin carboxyl carrier protein of higher plants

Acetyl-CoA carboxylase (ACCase; EC 6.4.1.2) is a regulatory enzyme of fatty acid synthesis, and in some higher-plant plastids is a multi-subunit complex consisting of biotin carboxylase (BC), biotin-carboxyl carrier protein (BCCP), and carboxyl transferase (CT). We recently described a Nicotiana tabacum L. (tobacco) cDNA with a deduced amino acid sequence similar to that of prokaryotic BC. We here provide further biochemical and immunological evidence that this higher-plant polypeptide is an authentic BC component of ACCase. The BC protein co-purified with ACCase activity and with BCCP during gel permeation chromatography of Pisum sativum L. (pea) chloroplast proteins. Antibodies to the Ricinus communis L. (castor) BC co-precipitated ACCase activity and BCCP. During castor seed development, ACCase activity and the levels of BC and BCCP increased and subsequently decreased in parallel, indicating their coordinate regulation. The BC protein comprised about 0.8% of the soluble protein in developing castor seed, and less than 0.05% of the protein in young leaf or root. Polypeptides cross-reacting with antibodies to castor BC were detected in several dicotyledons and in the monocotyledons Hemerocallis fulva L. (day lily), Iris L., and Allium cepa L. (onion), but not in the Gramineae species Hordeum vulgare L. (barley) and Panicum virgatum L. (switchgrass). The castor endosperm and pea chloroplast ACCases were not significantly inhibited by long-chain acyl-acyl carrier protein, free fatty acids or acyl carrier protein. The BC polypeptide was detected throughout Brassica napus L. (rapeseed) embryo development, in contrast to the multi-functional ACCase isoenzyme which was only detected early in development. These results firmly establish the identity of the BC polypeptide in plants and provide insight into the structure, regulation and roles of higherplant ACCases.Abbreviations ACCase acetyl-CoA carboxylase - ACP acyl carrier protein - BC biotin carboxylase - BCCP biotin carboxyl carrier protein - CT carboxyl transferase - MF multi-functional - MS multi-subunit We thank our colleagues Nicki Engeseth and Vicki Eccleston for advice on fatty acid analysis and Sarah Hunter for providing the developing Iris seed. This work was supported in part by grant MCB 9406466 from NSF. Acknowledgement is also made to the Michigan Agriculture Experiment Station for its support of this research.     

Structural evidence for substrate-induced synergism and half-sites reactivity in biotin carboxylase

Bacterial acetyl-CoA carboxylase is a multifunctional biotin-dependent enzyme that consists of three separate proteins: biotin carboxylase (BC), biotin carboxyl carrier protein (BCCP), and carboxyltransferase (CT). Acetyl-CoA carboxylase is a potentially attractive target for novel antibiotics because it catalyzes the first committed step in fatty acid biosynthesis. In the first half-reaction, BC catalyzes the ATP-dependent carboxylation of BCCP. In the second half-reaction, the carboxyl group is transferred from carboxybiotinylated BCCP to acetyl-CoA to produce malonyl-CoA. A series of structures of BC from several bacteria crystallized in the presence of various ATP analogs is described that addresses three major questions concerning the catalytic mechanism. The structure of BC bound to AMPPNP and the two catalytically essential magnesium ions resolves inconsistencies between the kinetics of active-site BC mutants and previously reported BC structures. Another structure of AMPPNP bound to BC shows the polyphosphate chain folded back on itself, and not in the correct (i.e., extended) conformation for catalysis. This provides the first structural evidence for the hypothesis of substrate-induced synergism, which posits that ATP binds nonproductively to BC in the absence of biotin. The BC homodimer has been proposed to exhibit half-sites reactivity where the active sites alternate or "flip-flop" their catalytic cycles. A crystal structure of BC showed the ATP analog AMPPCF(2)P bound to one subunit while the other subunit was unliganded. The liganded subunit was in the closed or catalytic conformation while the unliganded subunit was in the open conformation. This provides the first structural evidence for half-sites reactivity in BC.     

Organization and nucleotide sequences of the genes encoding the biotin carboxyl carrier protein and biotin carboxylase protein of Pseudomonas aeruginosa acetyl coenzyme A carboxylase.

The genetic organization of the Pseudomonas aeruginosa acetyl coenzyme A carboxylase (ACC) was investigated by cloning and characterizing a P. aeruginosa DNA fragment that complements an Escherichia coli strain with a conditional lethal mutation affecting the biotin carboxyl carrier protein (BCCP) subunit of ACC. DNA sequencing and RNA blot hybridization studies indicated that the P. aeruginosa accB (fabE) homolog, which encodes BCCP, is part of a 2-gene operon that includes accC (fabG), the structural gene for the biotin carboxylase subunit of ACC. P. aeruginosa homologs of the E. coli accA and accD, encoding the alpha and beta subunits of the ACC carboxyltransferase, were identified by hybridization of P. aeruginosa genomic DNA with the E. coli accA and accD. Data are presented which suggest that P. aeruginosa accA and accD homologs are not located either immediately upstream or downstream of the P. aeruginosa accBC operon. In contrast to E. coli, where BCCP is the only biotinylated protein, P. aeruginosa was found to contain at least three biotinylated proteins.     

Ligand specificity of group I biotin protein ligase of Mycobacterium tuberculosis

Background

Fatty acids are indispensable constituents of mycolic acids that impart toughness & permeability barrier to the cell envelope of M. tuberculosis. Biotin is an essential co-factor for acetyl-CoA carboxylase (ACC) the enzyme involved in the synthesis of malonyl-CoA, a committed precursor, needed for fatty acid synthesis. Biotin carboxyl carrier protein (BCCP) provides the co-factor for catalytic activity of ACC.

Methodology/Principal Findings

BPL/BirA (Biotin Protein Ligase), and its substrate, biotin carboxyl carrier protein (BCCP) of Mycobacterium tuberculosis (Mt) were cloned and expressed in E. coli BL21. In contrast to EcBirA and PhBPL, the ∼29.5 kDa MtBPL exists as a monomer in native, biotin and bio-5′AMP liganded forms. This was confirmed by molecular weight profiling by gel filtration on Superdex S-200 and Dynamic Light Scattering (DLS). Computational docking of biotin and bio-5′AMP to MtBPL show that adenylation alters the contact residues for biotin. MtBPL forms 11 H-bonds with biotin, relative to 35 with bio-5′AMP. Docking simulations also suggest that bio-5′AMP hydrogen bonds to the conserved ‘GRGRRG’ sequence but not biotin. The enzyme catalyzed transfer of biotin to BCCP was confirmed by incorporation of radioactive biotin and by Avidin blot. The K_m for BCCP was ∼5.2 µM and ∼420 nM for biotin. MtBPL has low affinity (K_b = 1.06×10⁻⁶ M) for biotin relative to EcBirA but their K_m are almost comparable suggesting that while the major function of MtBPL is biotinylation of BCCP, tight binding of biotin/bio-5′AMP by EcBirA is channeled for its repressor activity.

Conclusions/Significance

These studies thus open up avenues for understanding the unique features of MtBPL and the role it plays in biotin utilization in M. tuberculosis.     

Single‐particle analysis of urea amidolyase reveals its molecular mechanism

Urea amidolyase (UA), a bifunctional enzyme that is widely distributed in bacteria, fungi, algae, and plants, plays a pivotal role in the recycling of nitrogen in the biosphere. Its substrate urea is ultimately converted to ammonium, via successive catalysis at the C‐terminal urea carboxylase (UC) domain and followed by the N‐terminal allophanate hydrolyse (AH) domain. Although our previous studies have shown that Kluyveromyces lactis UA (KlUA) functions efficiently as a homodimer, the architecture of the full‐length enzyme remains unresolved. Thus how the biotin carboxyl carrier protein (BCCP) domain is transferred within the UC domain remains unclear. Here we report the structures of full‐length KlUA in its homodimer form in three different functional states by negatively‐stained single‐particle electron microscopy. We report here that the ADP‐bound structure with or without urea shows two possible locations of BCCP with preferred asymmetry, and that when BCCP is attached to the carboxyl transferase domain of one monomer, it is attached to the biotin carboxylase domain in the second domain. Based on this observation, we propose a BCCP‐swinging model for biotin‐dependent carboxylation mechanism of this enzyme.     

Regulation of the structure and activity of pyruvate carboxylase by acetyl CoA

In this review we examine the effects of the allosteric activator, acetyl CoA on both the structure and catalytic activities of pyruvate carboxylase. We describe how the binding of acetyl CoA produces gross changes to the quaternary and tertiary structures of the enzyme that are visible in the electron microscope. These changes serve to stabilize the tetrameric structure of the enzyme. The main locus of activation of the enzyme by acetyl CoA is the biotin carboxylation domain of the enzyme where ATP-cleavage and carboxylation of the biotin prosthetic group occur. As well as enhancing reaction rates, acetyl CoA also enhances the binding of some substrates, especially HCO3-, and there is also a complex interaction with the binding of the cofactor Mg2. The activation of pyruvate carboxylase by acetyl CoA is generally a cooperative processes, although there is a large degree of variability in the degree of cooperativity exhibited by the enzyme from different organisms. The X-ray crystallographic holoenzyme structures of pyruvate carboxylases from Rhizobium etli and Staphylococcus aureus have shown the allosteric acetyl CoA binding domain to be located at the interfaces of the biotin carboxylation and carboxyl transfer and the carboxyl transfer and biotin carboxyl carrier protein domains.     

Selectivity in post-translational biotin addition to five human carboxylases

Human holocarboxylase synthetase (HCS) catalyzes linkage of the vitamin biotin to the biotin carboxyl carrier protein (BCCP) domain of five biotin-dependent carboxylases. In the two-step reaction, the activated intermediate, bio-5'-AMP, is first synthesized from biotin and ATP, followed by covalent linkage of the biotin moiety to a specific lysine residue of each carboxylase BCCP domain. Selectivity in HCS-catalyzed biotinylation to the carboxylases was investigated in single turnover stopped flow and quench flow measurements of biotin transfer to the minimal biotin acceptor BCCP fragments of the carboxylases. The results demonstrate that biotinylation of the BCCP fragments of the mitochondrial carboxylases propionyl-CoA carboxylase, pyruvate carboxylase, and methylcrotonoyl-CoA carboxylase is fast and limited by the bimolecular association rate of the enzyme with substrate. By contrast, biotinylation of the acetyl-CoA carboxylase 1 and 2 (ACC1 and ACC2) fragments, both of which are accessible to HCS in the cytoplasm, is slow and displays a hyperbolic dependence on substrate concentration. The correlation between HCS accessibility to biotin acceptor substrates and the kinetics of biotinylation suggests that mitochondrial carboxylase sequences evolved to produce fast association rates with HCS in order to ensure biotinylation prior to mitochondrial import. In addition, the results are consistent with a role for HCS specificity in dictating biotin distribution among carboxylases.     

Biotinylation in the hyperthermophile Aquifex aeolicus.

Biotin protein ligase (BPL) catalyses the biotinylation of the biotin carboxyl carrier protein (BCCP) subunit of acetyl CoA carboxylase and this post-translational modification of a single lysine residue is exceptionally specific. The exact details of the protein-protein interactions involved are unclear as a BPL:BCCP complex has not yet been isolated. Moreover, detailed information is lacking on the composition, biosynthesis and role of fatty acids in hyperthermophilic organisms. We have cloned, overexpressed and purified recombinant BPL and the biotinyl domain of BCCP (BCCP Delta 67) from the extreme hyperthermophile Aquifex aeolicus. In vitro assays have demonstrated that BPL catalyses biotinylation of lysine 117 on BCCP Delta 67 at temperatures of up to 70 degrees C. Limited proteolysis of BPL with trypsin and chymotrypsin revealed a single protease-sensitive site located 44 residues from the N-terminus. This site is adjacent to the predicted substrate-binding site and proteolysis of BPL is significantly reduced in the presence of MgATP and biotin. Chemical crosslinking with 1-ethyl-3-(dimethylamino-propyl)-carbodiimide (EDC) allowed the isolation of a BPL:apo-BCCP Delta 67 complex. Furthermore, this complex was also formed between BPL and a BCCP Delta 67 mutant lacking the lysine residue (BCCP Delta 67 K117L) however, complex formation was considerably reduced using holo-BCCP Delta 67. These observations provide evidence that addition of the biotin prosthetic group reduces the ability of BCCP Delta 67 to heterodimerize with BPL, and emphasizes that a network of interactions between residues on both proteins mediates protein recognition.     

Biotinoyl domain of human acetyl-CoA carboxylase: Structural insights into the carboxyl transfer mechanism

Acetyl-CoA carboxylase (ACC) catalyzes the first step in fatty acid biosynthesis: the synthesis of malonyl-CoA from acetyl-CoA. As essential regulators of fatty acid biosynthesis and metabolism, ACCs are regarded as therapeutic targets for the treatment of metabolic diseases such as obesity. In ACC, the biotinoyl domain performs a critical function by transferring an activated carboxyl group from the biotin carboxylase domain to the carboxyl transferase domain, followed by carboxyl transfer to malonyl-CoA. Despite the intensive research on this enzyme, only the bacterial and yeast ACC structures are currently available. To explore the mechanism of ACC holoenzyme function, we determined the structure of the biotinoyl domain of human ACC2 and analyzed its characteristics and interaction with the biotin ligase, BirA using NMR spectroscopy. The 3D structure of the hACC2 biotinoyl domain has a similar folding topology to the earlier determined domains from E. coli and P. shermanii. However, the local structures near the biotinylation sites have notable differences that include the geometry of the consensus "Met-Lys-Met" (MKM) motif and the absence of "thumb" structure in the hACC2 biotinoyl domain. Observations of the NMR signals upon the biotinylation indicate that the biotin group of hACC2 does not affect the structure of the biotinoyl domain, while the biotin group for E. coli ACC interacts directly with the thumb residues that are not present in the hACC2 structure. These results imply that, in the E. coli ACC reaction, the biotin moiety carrying the carboxyl group from BC to CT can pause at the thumb of the BCCP domain. The human biotinoyl domain, however, lacks the thumb structure and does not have additional noncovalent interactions with the biotin moiety; thus, the flexible motion of the biotinylated lysine residue must underlie the "swinging arm" motion. The chemical shift perturbation and the cross saturation experiments of the human ACC2 holo-biotinoyl upon the addition of the biotin ligase (BirA) showed the interaction surface near the MKM motif, the two glutamic acids (Glu 926, Glu 953), and the positively charged residues (several lysine and arginine residues). This study provides insight into the mechanism of ACC holoenzyme function and supports the swinging arm model in human ACCs.     

Comparison of the backbone dynamics of the apo- and holo-carboxy-terminal domain of the biotin carboxyl carrier subunit of Escherichia coli acetyl-CoA carboxylase

The biotin carboxyl carrier protein (BCCP) is a subunit of acetyl-CoA carboxylase, a biotin-dependent enzyme that catalyzes the first committed step of fatty acid biosynthesis. In its functional cycle, this protein engages in heterologous protein-protein interactions with three distinct partners, depending on its state of post-translational modification. Apo-BCCP interacts specifically with the biotin holoenzyme synthetase, BirA, which results in the post-translational attachment of biotin to a single lysine residue on BCCP. Holo-BCCP then interacts with the biotin carboxylase subunit of acetyl-CoA carboxylase, which leads to the addition of the carboxylate group of bicarbonate to biotin. Finally, the carboxy-biotinylated form of BCCP interacts with transcarboxylase in the transfer of the carboxylate to acetyl-CoA to form malonyl-CoA. The determinants of protein-protein interaction specificity in this system are unknown. The NMR solution structure of the unbiotinylated form of an 87 residue C-terminal domain fragment (residue 70-156) of BCCP (holoBCCP87) and the crystal structure of the biotinylated form of a C-terminal fragment (residue 77-156) of BCCP from Escherichia coli acetyl-CoA carboxylase have previously been determined. Comparative analysis of these structures provided evidence for small, localized conformational changes in the biotin-binding region upon biotinylation of the protein. These structural changes may be important for regulating specific protein-protein interactions. Since the dynamic properties of proteins are correlated with local structural environments, we have determined the relaxation parameters of the backbone 15N nuclear spins of holoBCCP87, and compared these with the data obtained for the apo protein. The results indicate that upon biotinylation, the inherent mobility of the biotin-binding region and the protruding thumb, with which the biotin group interacts in the holo protein, are significantly reduced.     

Overexpression of Acetyl-CoA Carboxylase Gene of <Emphasis Type="Italic">Mucor rouxii</Emphasis> Enhanced Fatty Acid Content in <Emphasis Type="Italic">Hansenula polymorpha</Emphasis>

Malonyl-CoA is an essential precursor for fatty acid biosynthesis that is generated from the carboxylation of acetyl-CoA. In this work, a gene coding for acetyl-CoA carboxylase (ACC) was isolated from an oleaginous fungus, Mucor rouxii. According to the amino acid sequence homology and the conserved structural organization of the biotin carboxylase, biotin carboxyl carrier protein, and carboxyl transferase domains, the cloned gene was characterized as a multi-domain ACC1 protein. Interestingly, a 40% increase in the total fatty acid content of the non-oleaginous yeast Hansenula polymorpha was achieved by overexpressing the M. rouxii ACC1. This result demonstrated a significant improvement in the production of fatty acids through genetic modification in this yeast strain.     

粘红酵母乙酰辅酶A羧化酶（ACC）基因的克隆及CT功能域基因的原核表达

利用简并PCR结合染色体步移法首次克隆获得粘红酵母乙酰辅酶A羧化酶（ACC）基因的全长序列信息。序列分析表明,该基因包含2个内含子,分别位于42～147 bp和315～677 bp处,编码区域总长为6 801bp,推导的氨基酸序列进行二级结构分析具备乙酰辅酶A羧化酶典型的3个功能域：生物素羧化酶（BC）、生物素羧基载体蛋白（BCCP）和羧基转移酶（CT）。克隆该基因的CT功能域基因,连接到原核表达载体pET-28a上,在Escherichia coli BL21（DE3）中成功表达,利用Ni-NTA树脂柱纯化获得CT的可溶性重组蛋白,浓度为1.8mg/mL,为研究ACC的功能和针对CT作用的除草剂机理研究提供了有价值的材料。     

The biotinyl domain of Escherichia coli acetyl-CoA carboxylase. Evidence that the "thumb" structure id essential and that the domain functions as a dimer

Biotin carboxyl carrier protein (BCCP) is the small biotinylated subunit of Escherichia coli acetyl-CoA carboxylase (ACC), the enzyme that catalyzes the first committed step of fatty acid synthesis. Similar proteins are found in other bacteria and in chloroplasts. E. coli BCCP is a member of a large family of protein domains modified by covalent attachment of biotin to a specific lysine residue. However, the BCCP biotinyl domain differs from many of these proteins in that an eight-amino acid residue insertion is present upstream of the biotinylated lysine. X-ray crystallographic and multidimensional NMR studies show that these residues constitute a structure that has the appearance of an extended thumb that protrudes from the otherwise highly symmetrical domain structure. I report that expression of two mutant BCCPs lacking the thumb residues fails to restore growth and fatty acid synthesis to a temperature-sensitive E. coli strain that lacks BCCP when grown at nonpermissive temperature. Alignment of BCCPs from various organisms shows that only two of the eight thumb residues are strictly conserved, and amino acid substitution of either residue results in proteins giving only weak growth of the temperature-sensitive E. coli strain. Therefore, the thumb structure is essential for the function of BCCP in the ACC reaction and provides a useful motif for distinguishing the biotinylated proteins of multisubunit ACCs from those of enzymes catalyzing other biotin-dependent reactions. An unexpected result was that expression of a mutant BCCP in which the biotinylated lysine residue was substituted with cysteine was able to partially restore growth and fatty acid synthesis to the temperature-sensitive E. coli strain. This complementation was shown to be specific to BCCPs having native structure (excepting the biotinylated lysine) and is interpreted in terms of dimerization of the BCCP biotinyl domain during the ACC reaction.     

Mechanism of biotin carboxylase inhibition by ethyl 4-[[2-chloro-5-(phenylcarbamoyl)phenyl]sulphonylamino]benzoate

The rise of antibacterial-resistant bacteria is a major problem in the United States of America and around the world. Millions of patients are infected with antimicrobial resistant bacteria each year. Novel antibacterial agents are needed to combat the growing and present crisis. Acetyl-CoA carboxylase (ACC), the multi-subunit complex which catalyses the first committed step in fatty acid synthesis, is a validated target for antibacterial agents. However, there are at present, no commercially available antibiotics that target ACC. Ethyl 4-[[2-chloro-5-(phenylcarbamoyl)phenyl]sulfonylamino]benzoate (SABA1) is a compound that has been shown to have antibacterial properties against Pseudomonas aeruginosa and Escherichia coli. SABA1 inhibits biotin carboxylase (BC), the enzyme that catalyses the first half reaction of ACC. SABA1 inhibits BC via an atypical mechanism. It binds in the biotin binding site in the presence of ADP. SABA1 represents a potentially new class of antibiotics that can be used to combat the antibacterial resistance crisis.