研究者業績

柴田 直樹

シバタ ナオキ  (Naoki Shibata)

基本情報

所属
兵庫県立大学 大学院理学研究科生命科学専攻 助教授・准教授
学位
博士(工学)(大阪大学)

J-GLOBAL ID
200901039667162990
researchmap会員ID
1000224235

外部リンク

論文

 138
  • Satoru Hiroto, Naoki Aratani, Naoki Shibata, Yoshiki Higuchi, Takahiro Sasamori, Norihiro Tokitoh, Hiroshi Shinokubo, Atsuhiro Osuka
    Angewandte Chemie - International Edition 48(13) 2388-2390 2009年3月16日  査読有り
    Pyridine attacks: Nucleophilic addition of pyridine derivatives to a doubly linked corrole, which is a stable singlet biradical species, occurs at the bay area with high regioselectivity to provide zwitterionic dimers (see picture Ar=C6F5). Charge transfer between the anionic corrole and the pyridinium groups induces effective fluorescence quenching of the corrole dimer, which can be utilized for selective fluoride ion recognition. © 2009 Wiley-VCH Verlag GmbH &amp Co. KGaA.
  • 大須賀 久織, 寺脇 慎一, 庄村 康人, 小森 博文, 柴田 直樹, 廣田 俊, 樋口 芳樹
    生物物理 49 S7 2009年  
  • 虎谷 哲夫, 稗田 直樹, 秋田 敬太, 川口 智史, 馬場 伸之, 森 光一, 柴田 直樹, 玉垣 裕子, 樋口 芳樹
    ビタミン 83(5) 313-314 2009年  
  • Yasufumi Ueda, Naoki Shibata, Daisuke Takeuchi, Masaya Kitamura, Yoshiki Higuchi
    ACTA CRYSTALLOGRAPHICA SECTION F-STRUCTURAL BIOLOGY AND CRYSTALLIZATION COMMUNICATIONS 64(Pt 9) 851-853 2008年9月  査読有り
    Flavoredoxin from Desulfovibrio vulgaris Miyazaki F has been overexpressed, purified and crystallized using the sitting-drop vapour-diffusion method with 10%(w/v) PEG 8000, 0.2 M zinc acetate and 100 mM MES pH 6.0. The diffraction pattern of the crystal extended to 1.05 angstrom resolution under cryogenic conditions. The space group was determined to be P3(1)21, with unit-cell parameters a = b = 53.35, c = 116.22 angstrom. Phase determination was carried out by the SAD method using methylmercuric chloride.
  • Kiyohito Kihira, Shuko Numata, Masaya Kitamura, Jun Kondo, Shinichi Terawaki, Yasuhito Shomura, Hirofumi Komori, Naoki Shibata, Yoshiki Higuchi
    ACTA CRYSTALLOGRAPHICA SECTION F-STRUCTURAL BIOLOGY AND CRYSTALLIZATION COMMUNICATIONS 64(Pt 7) 622-624 2008年7月  査読有り
    Class II release factor 3 (RF3) from the sulfate-reducing bacterium Desulfovibrio vulgaris Miyazaki F, which promotes rapid dissociation of a class I release factor, has been overexpressed, purified and crystallized in complex with GDP at 293 K using the sitting-drop vapourdiffusion method. A data set was collected to 1.8 angstrom resolution from a single crystal at 100 K using synchrotron radiation. The crystal belongs to space group P1, with unit cell parameters a = 47.39, b = 82.80, c = 148.29 angstrom, alpha = 104.21, beta = 89.78, gamma = 89.63 degrees. The asymmetric unit contains four molecules of the RF3-GDP complex. The Matthews coefficient was calculated to be 2.3 angstrom(3) Da(1) and the solvent content was estimated to be 46.6%.
  • Kihira Kiyohito, Numata Shuko, Kitamura Masaya, Kondo Jun, Terawaki Shinichi, Shomura Yasuhito, Komori Hirofumi, Shibata Naoki, Higuchi Yoshiki
    生物物理 48 S82 2008年  
  • Shibata Naoki, Tamagaki Hiroko, Komori Hirofumi, Shomura Yasuhito, Hieda Naoki, Akita Keita, Mori Koichi, Toraya Tetsuo, Yasuoka Noritake, Higuchi Yoshiki
    生物物理 48 S77 2008年  
  • Osuka Hisao, Terawaki Shin-ichi, Shomura Yasuhito, Komori Hirofumi, Shibata Naoki, Hirota Shun, Higuchi Yoshiki
    生物物理 48 S80 2008年  
  • Tanaka Y, Saito S, Mori S, Aratani N, Shinokubo H, Shibata N, Higuchi Y, Yoon ZS, Kim KS, Noh SB, Park JK, Kim D, Osuka A
    Angewandte Chemie (International ed. in English) 47(4) 681-684 2008年  査読有り
  • Hideki Kajiura, Koichi Mori, Naoki Shibata, Tetsuo Toraya
    FEBS JOURNAL 274(21) 5556-5566 2007年11月  査読有り
    Adenosylcobalamin-dependent diol and glycerol dehydratases are isofunctional enzymes and undergo mechanism-based inactivation by a physiological substrate glycerol during catalysis. Inactivated holoenzymes are reactivated by their own reactivating factors that mediate the ATP-dependent exchange of an enzyme-bound, damaged cofactor for free adenosylcobalamin through intermediary formation of apoenzyme. The reactivation takes place in two steps: (a) ADP-dependent cobalamin release and (b) ATP-dependent dissociation of the resulting apoenzyme-reactivating factor complexes. The in vitro experiments with purified proteins indicated that diol dehydratase-reactivating factor (DDR) cross-reactivates the inactivated glycerol dehydratase, whereas glycerol dehydratase-reactivating factor (GDR) did not cross-reactivate the inactivated diol dehydratase. We investigated the molecular basis of their specificities in vitro by using purified preparations of cognate and noncognate enzymes and reactivating factors. DDR mediated the exchange of glycerol dehydratase-bound cyanocobalamin for free adeninylpentylcobalamin, whereas GDR cannot mediate the exchange of diol dehydratase-bound cyanocobalamin for free adeninylpentylcobalamin. As judged by denaturing PAGE, the glycerol dehydratase-DDR complex was cross-formed, although the diol dehydratase-GDR complex was not formed. There were no specificities of reactivating factors in the ATP-dependent dissociation of enzyme-reactivating factor complexes. Thus, it is very likely that the specificities of reactivating factors are determined by the capability of reactivating factors to form complexes with apoenzymes. A modeling study based on the crystal structures of enzymes and reactivating factors also suggested why DDR cross-forms a complex with glycerol dehydratase, and why GDR does not cross-form a complex with diol dehydratase.
  • Yasuhito Shomura, Hirofumi Komori, Natsuko Miyabe, Masamitsu Tomiyama, Naoki Shibata, Yoshiki Higuchi
    JOURNAL OF MOLECULAR BIOLOGY 372(4) 1045-1054 2007年9月  査読有り
    The hydrogenase maturation protein HypE serves an essential function in the biosynthesis of the nitrile group, which is subsequently coordinated to Fe as CN- ligands in [Ni-Fe] hydrogenase. Here, we present the crystal structures of HypE from Desulfovibrio vulgaris Hildenborough in the presence and in the absence of ATP at a resolution of 2.0 angstrom and 2.6 angstrom, respectively. Comparison of the apo structure with the ATP-bound structure reveals that binding ATP causes an induced-fit movement of the N-terminal portion, but does not entail an overall structural change. The residue Cys341 at the C terminus, whose thiol group is supposed to be carbamoylated before the nitrile group synthesis, is completely buried within the protein and is located in the vicinity of the gamma-phosphate group of the bound ATP. This suggests that the catalytic reaction occurs in this configuration but that a conformational change is required for the carbamoylation of Cys341. A glutamate residue is found close to the thiol group as well, which is suggestive of deprotonation of the carbamoyl group at the beginning of the reactions. (c) 2007 Elsevier Ltd. All rights reserved.
  • Naoki Shibata, Yusuke Tomimoto, Toru Hanamura, Ryo Yamamoto, Mai Ueda, Yasufumi Ueda, Nobuhiro Mizuno, Hideaki Ogata, Hirofumi Komori, Yasuhito Shomura, Michihiko Kataoka, Sakayu Shimizu, Jun Kondo, Hideki Yamamoto, Akira Kikuchi, Yoshiki Higuchi
    ACTA CRYSTALLOGRAPHICA SECTION F-STRUCTURAL BIOLOGY COMMUNICATIONS 63(Pt 6) 529-531 2007年6月  査読有り筆頭著者責任著者
    Axin is a negative regulator of the canonical Wnt signalling pathway that mediates the phosphorylation of beta-catenin by glycogen synthase kinase 3 beta. The DIX domain of rat axin, which is important for its homooligomerization and interactions with other regulators in the Wnt pathway, was purified and crystallized by the sitting-drop vapour-diffusion technique using polyethylene glycol 6000 and lithium sulfate as crystallization agents. Crystals belong to space group P6(1) or P6(5), with unit-cell parameters a = b = 91.49, c = 84.92 angstrom. An X-ray diffraction data set has been collected to a nominal resolution of 2.9 angstrom.
  • Thomas Schwarz-Romond, Marc Fiedler, Naoki Shibata, P. Jonathan G. Butler, Akira Kikuchi, Yoshiki Higuchi, Mariann Bienz
    NATURE STRUCTURAL & MOLECULAR BIOLOGY 14(6) 484-492 2007年6月  査読有り筆頭著者
    The Wnt signaling pathway controls numerous cell fates in animal development and is also a major cancer pathway. Dishevelled (Dvl) transduces the Wnt signal by interacting with the cytoplasmic Axin complex. Dvl and Axin each contain a DIX domain whose molecular properties and structure are unknown. Here, we demonstrate that the DIX domain of Dvl2 mediates dynamic polymerization, which is essential for the signaling activity of Dvl2. The purified domain polymerizes gradually, reversibly and in a concentration dependent manner, ultimately forming fibrils. The Axin DIX domain has a novel structural fold largely composed of beta-strands that engage in head-to-tail self-interaction to form filaments in the crystal. The DIX domain thus seems to mediate the formation of a dynamic interaction platform with a high local concentration of binding sites for transient Wnt signaling partners; this represents a previously uncharacterized mechanistic principle, signaling by reversible polymerization.
  • Seiji Negoro, Taku Ohki, Naoki Shibata, Kazuhiro Sasa, Haruhisa Hayashi, Hidehiko Nakano, Kengo Yasuhira, Dai-ichiro Kato, Masahiro Takeo, Yoshiki Higuchi
    JOURNAL OF MOLECULAR BIOLOGY 370(1) 142-156 2007年6月  査読有り
    We performed X-ray crystallographic analyses of 6-aminohexanoate-dimer hydrolase (Hyb-24DN), an enzyme responsible for the degradation of nylon-6, an industry by-product, and of a complex between Hyb-24DNA- 12 (S112A-mutant of Hyb-24DN) and 6-aminohexanoate-linear dimer (Ald) at 1.58 angstrom and 1.4 angstrom resolution, respectively. In Hyb-24DN, Asp181-O-delta forms hydrogen bonds with Tyr170-O-eta, -two of the catalytic and binding amino acids, and a loop between Asn167 and Val177. This state is the so-called open form, allowing its substrate to bind in the space between the loop and catalytic residues. Upon substrate binding (in Hyb-24DN-A(112)/Ald complex), the loop is shifted 4.3 angstrom at Tyr170-C-alpha, and the side-chain of Tyr170 is rotated. By the combined effect, Tyr170-O-eta, moves a total of 10.5 angstrom, resulting in the formation of hydrogen bonds with the nitrogen of amide linkage in Ald (closed form). In addition, electrostatic interaction between Asp181-O-delta and the amino group in Ald stabilizes the substrate binding. We propose here that the enzyme catalysis proceeds according to the following steps: (i) Ald-induced transition from open to closed form, (ii) nucleophilic attack of Ser112 to Ald and formation of a tetrahedral intermediate, (iii) formation of acyl enzyme and transition to open form, (iv) deacylation. Amino acid substitutions reducing the enzyme/Ald interaction at positions 181 or 170 drastically decreased the Ald-hydrolytic activity, but had very little effect on esterolytic activity, suggesting that esterolytic reaction proceeds regardless of conversion. Present models illustrate why new activity against the nylon oligomer has evolved in an esterase with beta-lactamase folds, while retaining the original esterolytic functions. (C) 2007 Elsevier Ltd. All rights reserved.
  • Masaya Kitamura, Koji Terakawa, Hideo Inoue, Takuto Hayashida, Kyoko Suto, Yukio Morimoto, Noritake Yasuoka, Naoki Shibata, Yoshiki Higuchi
    JOURNAL OF BIOCHEMISTRY 141(4) 459-468 2007年4月  査読有り
    Mutants of flavin mononucleotide-binding protein (FMN-bp) were made by site-directed mutagenesis to investigate the role of carboxyl-terminal Leu122 of the pairing subunit in controlling redox potentials, binding the prosthetic group, and forming the tertiary and quaternary structure. We compared the oxidation-reduction potentials, FMN-binding properties, and higher structures of wild-type FMN-bp and four mutant proteins (L122Y, L122E, L122K and L122-deleted). We found that the redox potentials were affected by mutations. Also, the affinities of L122E, L122K and L122 deletion mutant apoproteins for FMN were lower than for the wild-type apoprotein, whereas the affinity of L122Y for FMN was increased. Analytical ultracentrifugation showed that the dissociation constants for dimerization of L122E and L122K were larger than for wild-type FMN-bp, whereas the dissociation constants for L122Y and the deletion mutant were lower than for the wild type. Finally, we determined the higher structures of L122Y, L122E and L122K mutants by X-ray crystallography. Our results show that the mutation of Leu122 in FMN-bp changes midpoint potentials, dissociation constants for FMN, and dimer formation, indicating that this residue is important in the pairing subunit.
  • 柴田 直樹, Schwarz-Romond Thomas, Fiedler Marc, Butler P. Jonathan G., 小森 博文, 庄村 康人, 山本 英樹, 菊池 章, Bienz Mariann, 樋口 芳樹
    生物物理 47 S28 2007年  
  • Kengo Yasuhira, Yuki Uedo, Naoki Shibata, Seiji Negoro, Masahiro Takeo, Yoshiki Higuchi
    ACTA CRYSTALLOGRAPHICA SECTION F-STRUCTURAL BIOLOGY COMMUNICATIONS 62(Pt 12) 1209-1211 2006年12月  査読有り
    6-Aminohexanoate-cyclic-dimer hydrolase (EI) from Arthrobacter sp. KI72 was expressed in Escherichia coli and purified by anion-exchange chromatography. EI was crystallized by the sitting-drop vapour-diffusion method with sodium citrate as precipitant in imidazole buffer pH 8.0. The crystal is hexagonal, with unit-cell parameters a = b = 130.75, c = 58.23 angstrom. Diffraction data were collected from native and mercury(II) dichloride-derivative crystals to resolutions of 1.90 and 2.06 angstrom, respectively.
  • Hirofumi Morishita, Masataka Umitsu, Yoji Murata, Naoki Shibata, Keiko Udaka, Yoshiki Higuchi, Hideo Akutsu, Tohru Yamaguchi, Takeshi Yagi, Takahisa Ikegami
    JOURNAL OF BIOLOGICAL CHEMISTRY 281(44) 33650-33663 2006年11月  査読有り
    The recent explosion in genome sequencing has revealed the great diversity of the cadherin superfamily. Within the superfamily, protocadherins, which are expressed mainly in the nervous system, constitute the largest subgroup. Nevertheless, the structures of only the classical cadherins are known. Thus, to broaden our understanding of the adhesion repertoire of the cadherin superfamily, we determined the structure of the N-terminal first extracellular cadherin domain of the cadherin-related neuronal receptor/protocadherin-alpha 4. The hydrophobic pocket essential for homophilic adhesiveness in the classical cadherins was not found, and the functional significance of this structural domain was supported by exchanging the first extracellular cadherin domains of protocadherin and classical cadherin. Moreover, potentially crucial variations were observed mainly in the loop regions. These included the protocadherin-specific disulfide-bonded Cys-X-5-Cys motif, which showed Ca2+-induced chemical shifts, and the RGD motif, which has been suggested to be involved in heterophilic cell adhesion via the active form of beta 1 integrin. Our findings reveal that the adhesion repertoire of the cadherin superfamily is far more divergent than would be predicted by studying the classical cadherins alone.
  • Taku Ohki, Yoshiaki Wakitani, Masahiro Takeo, Kengo Yasuhira, Naoki Shibata, Yoshiki Higuchi, Seiji Negoro
    FEBS letters 580(21) 5054-8 2006年9月18日  査読有り
    Carboxylesterase (EII') from Arthrobacter sp. KI72 has 88% homology to 6-aminohexanoate-dimer hydrolase (EII) and possesses ca. 0.5% of the level of 6-aminohexanoate-linear dimer (Ald)-hydrolytic activity of EII. To study relationship between Ald-hydrolytic and esterolytic activities, random mutations were introduced into the gene for Hyb-24 (an EII/EII' hybrid with the majority of the sequence deriving for EII' and possessing an EII'-like level of Ald-hydrolytic activity). Either a G181D or a D370Y substitution in Hyb-24 increased the Ald-hydrolytic activity ca. 10-fold, and a G181D/D370Y double substitution increased activity ca. 100-fold. On the basis of kinetic studies and the three-dimensional structure of the enzyme, we suggest that binding of Ald is improved by these mutations. D370Y increased esterolytic activity for glycerylbutyrate ca. 30-50-fold, whereas G181D decreased the activity to 30% of the parental enzyme.
  • Taisuke Kamada, Naoki Aratani, Toshiaki Ikeda, Naoki Shibata, Yoshiki Higuchi, Atsushi Wakamiya, Shigehiro Yamaguchi, Kil Suk Kim, Zin Seok Yoon, Dongho Kim, Atsuhiro Osuka
    JOURNAL OF THE AMERICAN CHEMICAL SOCIETY 128(23) 7670-7678 2006年6月  査読有り
    In noncoordinating solvents, meso-cinchomeronimide appended Zn(II) porphyrin 2 forms a cyclic trimer, while diporphyrins 7 exhibit high-fidelity self-sorting assembling to form discrete cyclic trimer, tetramer, and pentamer with large association constants from 7(in-in), 7(in-out), and 7(out-out), respectively, through almost perfect discrimination of enantiomeric and conformational differences of the meso-cinchomeronimide substituents. In the latter self-sorting processes, the dihedral angles dictated by the two pyridyl nitrogen atoms control the size of the aggregates; the trimer from 7(in-in), the tetarmer from 7(in-out), and the pentamer from 7(out-out). Cyclic structures of (2) 3 and (R-7(out-out)) 5 have been determined by single-crystal X-ray diffraction analysis.
  • H Komori, K Satomoto, Y Ueda, N Shibata, S Inagaki, S Yoshioka, S Aono, Y Higuchi
    ACTA CRYSTALLOGRAPHICA SECTION F-STRUCTURAL BIOLOGY AND CRYSTALLIZATION COMMUNICATIONS 62(Pt 5) 471-473 2006年5月  査読有り
    CooA, a homodimeric haem-containing protein, is responsible for transcriptional regulation in response to carbon monoxide ( CO). It has a b-type haem as a CO sensor. Upon binding CO to the haem, CooA binds promoter DNA and activates expression of genes for CO metabolism. CooA from Carboxy-dothermus hydrogenoformans has been overexpressed in Escherichia coli, purified and crystallized by the vapour-diffusion method. The crystal belongs to space group P2(1), with unit-cell parameters a = 61.8, b = 94.7, c = 92.8 angstrom, beta = 104.8 degrees. The native and anomalous difference Patterson maps indicated that two CooA dimers are contained in the asymmetric unit and are related by a translational symmetry almost parallel to the c axis.
  • N Shibata, K Mori, N Hieda, Y Higuchi, M Yamanishi, T Toraya
    STRUCTURE 13(12) 1745-1754 2005年12月  査読有り筆頭著者
    The crystal structures of ADP bound and nucleotide-free forms of molecular chaperone-like diol dehydratase-reactivating factor (DDR) were determined at 2.0 and 3.0 angstrom, respectively. DDR exists as a dimer of heterodimer (alpha beta)(2). The alpha subunit has four domains: ATPase domain, swiveling domain, linker domain, and insert domain. The beta subunit, composed of a single domain, has a similar fold to the beta subunit of diol dehydratase (DD). The binding of an ADP molecule to the nucleotide binding site of DDR causes a marked conformational change of the ATPase domain of the alpha subunit, which would weaken the interactions between the DDR alpha and beta subunits and make the displacement of the DDR beta subunit by DD through the beta subunit possible. The binding of the DD beta subunit to the DDR alpha subunit induces steric repulsion between the DDR alpha and DD alpha subunits that would lead to the release of a damaged cofactor from inactivated holoDD.
  • M Nakabayashi, N Shibata, H Komori, Y Ueda, H Iino, A Ebihara, S Kuramitsu, Y Higuchi
    ACTA CRYSTALLOGRAPHICA SECTION F-STRUCTURAL BIOLOGY AND CRYSTALLIZATION COMMUNICATIONS 61(Pt 12) 1027-1031 2005年12月  査読有り
    The crystal structure of a conserved hypothetical protein, TTHA0849 from Thermus thermophilus HB8, has been determined at 2.4 angstrom resolution as a part of a structural and functional genomics project on T. thermophilus HB8. The main-chain folding shows a compact alpha+beta motif, forming a hydrophobic cavity in the molecule. A structural similarity search reveals that it resembles those steroidogenic acute regulatory proteins that contain the lipid-transfer (START) domain, even though TTHA0849 shows comparatively weak sequence identity to polyketide cyclases. However, the size of the ligand-binding cavity is distinctly smaller than other START domain-containing proteins, suggesting that it catalyses the transfer of smaller ligand molecules.
  • S Negoro, T Ohki, N Shibata, N Mizuno, Y Wakitani, J Tsurukame, K Matsumoto, Kawamoto, I, M Takeo, Y Higuchi
    JOURNAL OF BIOLOGICAL CHEMISTRY 280(47) 39644-39652 2005年11月  査読有り
    6-Aminohexanoate- dimer hydrolase (EII), responsible for the degradation of nylon-6 industry by-products, and its analogous enzyme (EII') that has only similar to 0.5% of the specific activity toward the 6-aminohexanoate-linear dimer, are encoded on plasmid pOAD2 of Arthrobacter sp. (formerly Flavobacterium sp.) KI72. Here, we report the three-dimensional structure of Hyb-24 (a hybrid between the EII and EII' proteins; EII'-level activity) by x-ray crystallography at 1.8 angstrom resolution and refined to an R-factor and R-free of 18.5 and 20.3%, respectively. The fold adopted by the 392-amino acid polypeptide generated a two-domain structure that is similar to the folds of the penicillin-recognizing family of serine-reactive hydrolases, especially to those of D-alanyl-D-alanine-carboxypeptidase from Streptomyces and carboxylesterase from Burkholderia. Enzyme assay using purified enzymes revealed that EII and Hyb-24 possess hydrolytic activity for carboxyl esters with short acyl chains but no detectable activity for D-alanyl-D-alanine. In addition, on the basis of the spatial location and role of amino acid residues constituting the active sites of the nylon oligomer hydrolase, carboxylesterase, D-alanyl-D-alanine-peptidase, and beta-lactamases, we conclude that the nylon oligomer hydrolase utilizes nucleophilic Ser(112) as a common active site both for nylon oligomer-hydrolytic and esterolytic activities. However, it requires at least two additional amino acid residues (Asp(181) and Asn(266)) specific for nylon oligomer-hydrolytic activity. Here, we propose that amino acid replacements in the catalytic cleft of a preexisting esterase with the beta-lactamase fold resulted in the evolution of the nylon oligomer hydrolase.
  • H Ogata, S Hirota, A Nakahara, H Komori, N Shibata, T Kato, K Kano, Y Higuchi
    STRUCTURE 13(11) 1635-1642 2005年11月  査読有り
    Hydrogenases catalyze oxidoreduction of molecular hydrogen and have potential applications for utilizing dihydrogen as an energy source. [NiFe] hydrogenase has two different oxidized states, Ni-A (unready, exhibits a lag phase in reductive activation) and Ni-B (ready). We have succeeded in converting Ni-B to Ni-A with the use of Na2S and O-2 and determining the high-resolution crystal structures of both states. Ni-B possesses a monatomic nonprotein bridging ligand at the Ni-Fe active site, whereas Ni-A has a diatomic: species. The terminal atom of the bridging species of Ni-A occupies a similar position as C of the exogenous CO in the CO complex (inhibited state). The common features of the enzyme structures at the unready (Ni-A) and inhibited (CO complex) states are proposed. These findings provide useful information on the design of new systems of biomimetic dihydrogen production and fuel cell devices.
  • T Ohki, N Mizuno, N Shibata, M Takeo, S Negoro, Y Higuchi
    ACTA CRYSTALLOGRAPHICA SECTION F-STRUCTURAL BIOLOGY AND CRYSTALLIZATION COMMUNICATIONS 61(Pt 10) 928-930 2005年10月  査読有り
  • K Mori, N Hieda, M Yamanishi, N Shibata, T Toraya
    ACTA CRYSTALLOGRAPHICA SECTION F-STRUCTURAL BIOLOGY AND CRYSTALLIZATION COMMUNICATIONS 61(Pt 6) 603-605 2005年6月  査読有り
  • Hirota, S, Ogata, H, Nakahara, A, Komori, H, Shibata, N, Funasaki, N, Higuchi, Y
    12th International Conference on Biological Chemistry 2005年4月1日  
  • Ogata, H, Hirota, S, Nakahara, A, Komori, H, Shibata, N, Higuchi, Y
    XX Congress of the International Union of Crystallography 2005年4月1日  
  • 樋口 芳樹, 宮部 奈津子, 中原 明香, 緒方 英明, 小森 博文, 柴田 直樹, 廣田 俊
    生物物理 45 S206 2005年  
  • 柴田 直樹
    日本結晶学会誌 46(3) 209-215 2004年6月30日  
    Coenzyme B12 or adenosylcobalamin, commonly known as Vitamin B12, is one of the most complicated natural organic compounds involved in heteroatom elimination and carbonskeleton rearrangement reactions. Diol dehydratase is an adenosylcobalamin-dependant enzyme that catalyzes conversion from 1, 2-diol compounds to the corresponding aldehydes. Comparison of the structures of the enzyme of substrate-bound and substrate-free forms reveals the activation mechanism of adenosylcobalamin. Structural studies of chiral substrates imply that this enzyme catalyses each enantiomer with different mechanisms in the final step of the reaction.
  • A Yamakawa, H Ogata, K Morita, N Shibata, N Andou, H Sanuki, K Yamada, T Hioki, T Ishii, Y Higuchi
    ACTA CRYSTALLOGRAPHICA SECTION D-BIOLOGICAL CRYSTALLOGRAPHY 60(Pt 4) 764-766 2004年4月  査読有り
    The catalytic domain of death-associated protein kinase (DAPK) has been overexpressed, purified and crystallized using the sitting-drop vapour-diffusion method with PEG 8000 and magnesium acetate as precipitants. Complexes with the inhibitor staurosporine and its analogue BDB402 were also crystallized in the presence of PEG 400 and PEG 8000, respectively. Diffraction data were collected to 2.4 Angstrom for the native catalytic domain, to 2.9 Angstrom for the staurosporine complex and to 2.7 Angstrom for the BDB402 complex. All three crystals belong to space group P2(1)2(1)2(1), with unit-cell parameters a=77.992, b=109.909, c=50.063 Angstrom for the catalytic domain, a=78.911, b=113.162, c=50.658 Angstrom for the staurosporine complex and a=77.337, b=108.869, c=50.186 Angstrom for the BDB402 complex. In both complexes the inhibitor molecule was clearly assigned in the difference Fourier map calculated on the basis of the phases obtained from the structure of the catalytic domain.
  • N Shibata, K Suto, E Ichimura, K Yoshimura, K Muneo, S Tomigami, Y Morimoto, M Ogata, T Yagi, Y Higuchi, N Yasuoka
    PROTEIN AND PEPTIDE LETTERS 11(1) 93-96 2004年2月  査読有り筆頭著者
    Hexadecaheme high molecular weight cytochrome c from a sulfate-reducing bacterium, Desulfovibrio vulgaris Miyazaki F has been successfully purified and crystallized. X-ray diffraction data have been collected by the multiple wavelength anomalous dispersion method. The crystal belongs to the space group P2(1)2(1)2(1) with unit-cell parameters a = 60.42, b = 84.29 and c = 144.16 Angstrom and contains one molecule per asymmetric unit.
  • 中原 明香, 緒方 英明, 小森 博文, 柴田 直樹, 菊池 晶裕, 加藤 立久, 山内 脩, 廣田 俊, 樋口 芳樹
    生物物理 44 S111 2004年  
  • 佐藤 翠, 森本 幸生, 柴田 直樹, 小森 博文, 高山 裕生, 小澤 潔, 阿久津 秀雄, 樋口 芳樹
    生物物理 44 S131 2004年  
  • Y.Morimoto, M.Sato, N.Shibata, Y.Higuchi
    SPring-8 User Experiment Report 12 176 2004年  
  • M Sato, N Shibata, Y Morimoto, Y Takayama, K Ozawa, H Akutsu, Y Higuchi, N Yasuoka
    JOURNAL OF SYNCHROTRON RADIATION 11(Pt 1) 113-116 2004年1月  査読有り
    The crystal structures of high-molecular weight cytochrome e (HMC) from Desulfovibrio vulgaris Hildenborough in the transient and reduced states have been determined at 2.8 Angstrom resolution. An absorption spectrum measured with microspectrophotometor indicated that about 86% of the hemes were reduced after 45 min irradiation of x-ray beam. Further exposure for 90 min did not significantly change the spectrum. These results suggest that HMC in crystalline state is easily reduced by illumination of x-ray beam from synchrotron radiation.
  • 中原明香, 緒方英明, 柴田直樹, 廣田 俊, 樋口芳樹
    理研構造生物ポスターセッション 2003年8月4日  
  • N Shibata, Y Nakanishi, M Fukuoka, M Yamanishi, N Yasuoka, T Toraya
    JOURNAL OF BIOLOGICAL CHEMISTRY 278(25) 22717-22725 2003年6月  査読有り筆頭著者
    Adenosylcobalamin- dependent diol dehydratase of Klebsiella oxytoca is apparently not stereospecific and catalyzes the conversion of both ( R)- and ( S)- 1,2- propanediol to propionaldehyde. To explain this unusual property of the enzyme, we analyzed the crystal structures of diol dehydratase in complexes with cyanocobalamin and ( R)- or ( S)- 1,2- propanediol. ( R)- and ( S)- isomers are bound in a symmetrical manner, although the hydrogen- bonding interactions between the substrate and the active- site residues are the same. From the position of the adenosyl radical in the modeled " distal" conformation, it is reasonable for the radical to abstract the pro- R and pro- S hydrogens from ( R)- and ( S)- isomers, respectively. The hydroxyl groups in the substrate radicals would migrates from C( 2) to C( 1) by a suprafacial shift, resulting in the stereochemical inversion at C(1). This causes 60degrees clockwise and 70degrees counterclockwise rotations of the C( 1) - C( 2) bond of the ( R)- and ( S)- isomers, respectively, if viewed from K+. A modeling study of 1,1- gem- diol intermediates indicated that new radical center C( 2) becomes close to the methyl group of 5'-deoxyadenosine. Thus, the hydrogen back- abstraction ( recombination) from 5'- deoxyadenosine by the product radical is structurally feasible. It was also predictable that the substitution of the migrating hydroxyl group by a hydrogen atom from 5'- deoxyadenosine takes place with the inversion of the configuration at C( 2) of the substrate. Stereospecific dehydration of the 1,1- gem- diol intermediates can also be rationalized by assuming that Asp-alpha335 and Glu-alpha170 function as base catalysts in the dehydration of the ( R)- and ( S)- isomers, respectively. The structure- based mechanism and stereochemical courses of the reaction are proposed.
  • 山本 亮, 上田 康史, 山本 英樹, 菊地 章, 柴田 直樹, 樋口 芳樹
    生物物理 43 S37 2003年  
  • 中原 明香, 緒方 英明, 柴田 直樹, 廣田 俊, 樋口 芳樹
    生物物理 43 S37 2003年  
  • N Shibata, J Masuda, Y Morimoto, N Yasuoka, T Toraya
    BIOCHEMISTRY 41(42) 12607-12617 2002年10月  査読有り筆頭著者
    Substrate binding triggers catalytic radical formation through the cobalt-carbon bond homolysis in coenzyme B-12-dependent enzymes. We have determined the crystal structure of the substrate-free form of Klebsiella oxytoca diol dehydratase(.)cyanocobalamin complex at 1.85 Angstrom resolution. The structure contains two units of the heterotrimer consisting of alpha, beta, and gamma subunits. As compared with the structure of its substrate-bound form, the beta subunits are tilted by similar to3degrees and cobalamin is also tilted so that pyrrole rings A and D are significantly lifted up toward the substrate-binding site, whereas pyrrole rings B and C are only slightly lifted up. The structure revealed that the potassium ion in the substrate-binding site of the substrate-free enzyme is also heptacoordinated; that is, two oxygen atoms of two water molecules coordinate to it instead of the substrate hydroxyls. A modeling study in which the structures of both the cobalamin moiety and the adenine ring of the coenzyme were superimposed onto those of the enzyme-bound cyanocobalamin and the adenine ring-binding pocket, respectively, demonstrated that the distortions of the Co-C bond in the substrate-free form are already marked but slightly smaller than those in the substrate-bound form. It was thus strongly suggested that the Co-C bond becomes largely activated (labilized) when the coenzyme binds to the apoenzyme even in the absence of substrate and undergoes homolysis through the substrate-induced conformational changes of the enzyme. Kinetic coupling of Co-C bond homolysis with hydrogen abstraction from the substrate shifts the equilibrium to dissociation.
  • M Yamanishi, M Yunoki, T Tobimatsu, H Sato, J Matsui, A Dokiya, Y Iuchi, K Oe, K Suto, N Shibata, Y Morimoto, N Yasuoka, T Toraya
    EUROPEAN JOURNAL OF BIOCHEMISTRY 269(18) 4484-4494 2002年9月  査読有り
    Recombinant glycerol dehydratase of Klebsiella pneumoniae was purified to homogeneity. The subunit composition of the enzyme was most probably alpha(2) beta(2) gamma(2) . When (R )- and (S )-propane-1,2-diols were used independently as substrates, the rate with the (R )-enantiomer was 2.5 times faster than that with the (S )-isomer. In contrast to diol dehydratase, an isofunctional enzyme, the affinity of the enzyme for the (S )-isomer was essentially the same or only slightly higher than that for the (R )-isomer (K (m(R )) /K (m(S )) = 1.5). The crystal structure of glycerol dehydratase in complex with cyanocobalamin and propane-1,2-diol was determined at 2.1 Angstrom resolution. The enzyme exists as a dimer of the alphabetagamma heterotrimer. Cobalamin is bound at the interface between the alpha and beta subunits in the so-called 'base-on' mode with 5,6-dimethylbenzimidazole of the nucleotide moiety coordinating to the cobalt atom. The electron density of the cyano group was almost unobservable, suggesting that the cyanocobalamin was reduced to cob(II)alamin by X-ray irradiation. The active site is in a (beta/alpha)(8) barrel that was formed by a central region of the alpha subunit. The substrate propane-1,2-diol and essential cofactor K+ are bound inside the (beta/alpha)(8) barrel above the corrin ring of cobalamin. K+ is hepta-coordinated by the two hydroxyls of the substrate and five oxygen atoms from the active-site residues. These structural features are quite similar to those of diol dehydratase. A closer contact between the alpha and beta subunits in glycerol dehydratase may be reminiscent of the higher affinity of the enzyme for adenosylcobalamin than that of diol dehydratase. Although racemic propane-1,2-diol was used for crystallization, the substrate bound to glycerol dehydratase was assigned to the (R )-isomer. This is in clear contrast to diol dehydratase and accounts for the difference between the two enzymes in the susceptibility of suicide inactivation by glycerol.
  • E Mizohata, H Matsumura, Y Okano, M Kumei, H Takuma, J Onodera, K Kato, N Shibata, T Inoue, A Yokota, Y Kai
    JOURNAL OF MOLECULAR BIOLOGY 316(3) 679-691 2002年2月  査読有り
    Ribulose-1,5-bisphosphate carboxylase/oxygenase (rubisco) catalyzes the initial steps of photosynthetic carbon reduction and photorespiratory carbon oxidation cycles by combining CO2 and O-2, respectively, with ribulose-1,5-bisphosphate. Many photosynthetic organisms have form 1 rubiscos comprised of eight large (L) and eight small (S) subunits. The crystal structure of the complex of activated rubisco from the green alga Chlamydomonas reinliardtii and the reaction intermediate analogue 2-carboxyarabinitol-1,5-bisphosphate (2-CABP) has been solved at 1.84 Angstrom resolution (R-cryst of 15.2% and R-free of 18.1%). The subunit arrangement of Chlamydomonas rubisco is the same as those of the previously solved form I rubiscos. Especially, the present structure is very similar to the activated spinach structure complexed with 2-CABP in the L-subunit folding and active-site conformation, but differs in S-subunit folding. The central insertion of the Chlamydomonas S-subunit forms the longer betaA-betaB loop that protrudes deeper into the solvent channel of rubisco than higher plant, cyanobacterial, and red algal (red-like) betaA-betaB loops. The C-terminal extension of the Chlamydomonas S-subunit does not protrude into the solvent channel, unlike that of the red algal S-subunit, but lies on the protein surface anchored by interactions with the N-terminal region of the S-subunit. Further, the present high-resolution structure has revealed novel post-translational modifications. Residue 1 of the S-subunit is N-alpha-methylmethionine, residues 104 and 151 of the L-subunit are 4-hydroxyproline, and residues 256 and 369 of the L-subunit are S-gamma-methylcysteine. Furthermore, the unusual electron density of residue 471 of the L-subunit, which has been deduced to be threonine from the genomic DNA sequence, suggests that the residue is isoleucine produced by RNA editing or O-gamma-methylthreonine. (C) 2002 Elsevier Science Ltd.
  • Y.Morimoto, K.Suto, M.Nakabayashi, N.Shibata, N.Yasuoka
    SPring-8 User Experiment Report 8 290 2002年  
  • Y.Morimoto, K.Suto, N.Shibata, T.Hayashida, Y.Umena, N.Yasuoka
    SPring-8 User Experiment Report 8 183 2002年  
  • J Masuda, N Shibata, Y Morimoto, T Toraya, N Yasuoka
    JOURNAL OF SYNCHROTRON RADIATION 8(6) 1182-1185 2001年11月  査読有り
    In the course of structural studies of diol dehydratase-cobalamin complexes, it was found that the electron density corresponding to the cyano group of the enzyme-bound cyanocobalamin is almost not observable at room temperature and very low even at cryogenic temperatures, suggesting its dissociation from the Co atom upon X-ray irradiation. On the contrary, the adenine moiety of the enzyme-bound adeninylpentylcobalamin was clearly located in the electron density map. When the enzyme-adeninylpentylcobalamin complex was illuminated with visible light, the electron density between the C5' and Co atoms disappeared, and the temperature factors of the atoms comprising the pentamethylene group became much larger than those in the dark. This indicates a Co-C bond cleavage and that the adenine moiety remains held by hydrogen bonds with some residues in the enzyme. Thus, the formation of an adenine-anchored radical upon illumination was demonstrated crystallographically with this complex. These observations clearly indicate that homolysis of the Co-C bond of alkylcobalamin takes place upon illumination with visible light but is not readily cleaved during X-ray irradiation.
  • 安岡 則武, 柴田 直樹, 虎谷 哲夫
    生物物理 41(4) 201-204 2001年7月25日  

MISC

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  • 柴田直樹, 樋口芳樹, 虎谷哲夫
    第468回ビタミンB研究協議会 2022年9月  筆頭著者
  • 根来誠司, 武尾正弘, 柴田直樹, 樋口芳樹, 加藤太一郎, 重田育照
    月刊バイオインダストリー 2019年6月  招待有り
  • Yamanaka Masaru, Hoshizumi Makoto, Nagao Satoshi, Nakayama Ryoko, Shibata Naoki, Higuchi Yoshiki, Hirota Shun
    2017年2月14日  
    The number of artificial protein supramolecules has been increasing; however, control of protein oligomer formation remains challenging. Cytochrome c′ from Allochromatium vinosum (AVCP) is a homodimeric protein in its native form, where its protomer exhibits a four-helix bundle structure containing a covalently bound five-coordinate heme as a gas binding site. AVCP exhibits a unique reversible dimer-monomer transition according to the absence and presence of CO. Herein, domain-swapped dimeric AVCP was constructed and utilized to form a tetramer and high-order oligomers. The X-ray crystal structure of oxidized tetrameric AVCP consisted of two monomer subunits and one domain-swapped dimer subunit, which exchanged the region containing helices αA and αB between protomers. The active site structures of the domain-swapped dimer subunit and monomer subunits in the tetramer were similar to those of the monomer subunits in the native dimer. The subunit-subunit interactions at the interfaces of the domain-swapped dimer and monomer subunits in the tetramer were also similar to the subunit-subunit interaction in the native dimer. Reduced tetrameric AVCP dissociated to a domain-swapped dimer and two monomers upon CO binding. Without monomers, the domain-swapped dimers formed tetramers, hexamers, and higher-order oligomers in the absence of CO, whereas the oligomers dissociated to domain-swapped dimers in the presence of CO, demonstrating that the domain-swapped dimer maintains the CO-induced subunit dissociation behavior of native ACVP. These results suggest that protein oligomer formation may be controlled by utilizing domain swapping for a dimer-monomer transition protein.
  • 衣笠 凌, 羽田 圭吾, 谷本 悠樹, 竹原 一起, 柴田 直樹, 加藤 太一郎, 武尾 正弘, 根来 誠司
    日本生物工学会大会講演要旨集 67 282-282 2015年  
  • Y. Higuchi, M. Akter, C. Inoue, Y. Shomura, N. Shibata, K. Inaka, K. Kataoka, T. Sakurai, K. Komori
    JOURNAL OF BIOLOGICAL INORGANIC CHEMISTRY 19 S324-S324 2014年3月  

書籍等出版物

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主要な共同研究・競争的資金等の研究課題

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