Publications

• Crystal structures of biotin protein ligase from Pyrococcus horikoshii OT3 and its complexes: structural basis of biotin activation.

  Bagautdinov B., Kuroishi C., Sugahara M., Kunishima N.

  Biotin protein ligase (EC 6.3.4.15) catalyses the synthesis of an activated form of biotin, biotinyl-5'-AMP, from substrates biotin and ATP followed by biotinylation of the biotin carboxyl carrier protein subunit of acetyl-CoA carboxylase. The three-dimensional structure of biotin protein ligase f ... >> More

  Biotin protein ligase (EC 6.3.4.15) catalyses the synthesis of an activated form of biotin, biotinyl-5'-AMP, from substrates biotin and ATP followed by biotinylation of the biotin carboxyl carrier protein subunit of acetyl-CoA carboxylase. The three-dimensional structure of biotin protein ligase from Pyrococcus horikoshii OT3 has been determined by X-ray diffraction at 1.6A resolution. The structure reveals a homodimer as the functional unit. Each subunit contains two domains, a larger N-terminal catalytic domain and a smaller C-terminal domain. The structural feature of the active site has been studied by determination of the crystal structures of complexes of the enzyme with biotin, ADP and the reaction intermediate biotinyl-5'-AMP at atomic resolution. This is the first report of the liganded structures of biotin protein ligase with nucleotide and biotinyl-5'-AMP. The structures of the unliganded and the liganded forms are isomorphous except for an ordering of the active site loop upon ligand binding. Catalytic binding sites are suitably arranged to minimize the conformational changes required during the reaction, as the pockets for biotin and nucleotide are located spatially adjacent to each other in a cleft of the catalytic domain and the pocket for biotinyl-5'-AMP binding mimics the combination of those of the substrates. The exact locations of the ligands and the active site residues allow us to propose a general scheme for the first step of the reaction carried out by biotin protein ligase in which the positively charged epsilon-amino group of Lys111 facilitates the nucleophilic attack on the ATP alpha-phosphate group by the biotin carboxyl oxygen atom and stabilizes the negatively charged intermediates. << Less

  J Mol Biol 353:322-333(2005) [PubMed] [EuropePMC]

  This publication is cited by 1 other entry.

• Escherichia coli biotin holoenzyme synthetase/bio repressor crystal structure delineates the biotin- and DNA-binding domains.

  Wilson K.P., Shewchuk L.M., Brennan R.G., Otsuka A.J., Matthews B.W.

  The three-dimensional structure of BirA, the repressor of the Escherichia coli biotin biosynthetic operon, has been determined by x-ray crystallography and refined to a crystallographic residual of 19.0% at 2.3-A resolution. BirA is a sequence-specific DNA-binding protein that also catalyzes the f ... >> More

  The three-dimensional structure of BirA, the repressor of the Escherichia coli biotin biosynthetic operon, has been determined by x-ray crystallography and refined to a crystallographic residual of 19.0% at 2.3-A resolution. BirA is a sequence-specific DNA-binding protein that also catalyzes the formation of biotinyl-5'-adenylate from biotin and ATP and transfers the biotin moiety to other proteins. The level of biotin biosynthetic enzymes in the cell is controlled by the amount of biotinyl-5'-adenylate, which is the BirA corepressor. The structure provides an example of a transcription factor that is also an enzyme. The structure of BirA is highly asymmetric and consists of three domains. The N-terminal domain is mostly alpha-helical, contains a helix-turn-helix DNA-binding motif, and is loosely connected to the remainder of the molecule. The central domain consists of a seven-stranded mixed beta-sheet with alpha-helices covering one face. The other side of the sheet is largely solvent-exposed and contains the active site. The C-terminal domain comprises a six-stranded, antiparallel beta-sheet sandwich. The location of biotin binding is consistent with mutations that affect enzymatic activity. A nearby loop has a sequence that has been associated with phosphate binding in other proteins. It is inferred that ATP binds in this region, adjacent to the biotin. It is proposed that the binding of corepressor to monomeric BirA may promote DNA binding by facilitating the formation of a multimeric BirA-corepressor-DNA complex. The structural details of this complex remain an open question, however. << Less

  Proc. Natl. Acad. Sci. U.S.A. 89:9257-9261(1992) [PubMed] [EuropePMC]

  This publication is cited by 1 other entry.

• Kinetics of biotinyl-5'-adenylate synthesis catalyzed by the Escherichia coli repressor of biotin biosynthesis and the stability of the enzyme-product complex.

  Xu Y., Beckett D.

  The Escherichia coli repressor of biotin biosynthesis is both a biotin ligase and the repressor of transcriptional initiation at the biotin biosynthetic operon. The small molecule, biotinyl-5'-adenylate (bio-5'-AMP), is the intermediate in the biotin ligation reaction and the positive allosteric e ... >> More

  The Escherichia coli repressor of biotin biosynthesis is both a biotin ligase and the repressor of transcriptional initiation at the biotin biosynthetic operon. The small molecule, biotinyl-5'-adenylate (bio-5'-AMP), is the intermediate in the biotin ligation reaction and the positive allosteric effector for sequence-specific DNA binding by BirA. Synthesis of the adenylate from the substrates biotin and ATP is catalyzed by BirA. Although BirA and other biotin holoenzyme synthetases have been the subject of biochemical studies, no direct measurements of the bio-5'-AMP synthesis reaction have been reported. No information relating to the mechanism and kinetic parameters governing adenylate synthesis is available. In addition to this lack of kinetic information, the thermodynamic stability of the BirA-bio-5'-AMP complex is not known. Since the BirA-adenylate complex plays a pivotal role in the biotin regulatory system, both the kinetic and thermodynamic information are essential to a quantitative understanding of the system. We have developed a method for measuring the time course of bio-5'-AMP synthesis. The results of these measurements indicate that the time course is characterized by an initial burst followed by a slow linear phase. The burst corresponds to the rapid synthesis of 1 mol of product per mole of enzyme, and the rate of the slow linear phase is limited by the release of product from the enzyme.(ABSTRACT TRUNCATED AT 250 WORDS) << Less

  Biochemistry 33:7354-7360(1994) [PubMed] [EuropePMC]

  This publication is cited by 2 other entries.