Mechanism in Regulation of Transcription Factor containing Fe-S Clusters Sensing Oxidative Stress

• Project/Area Number: 19570128
• Research Category: Grant-in-Aid for Scientific Research (C)
• Allocation Type: Single-year Grants
• Section: 一般
• Research Field: Functional biochemistry
• Research Institution: Osaka University
• Principal Investigator: KOBAYASHI Kazuo Osaka University, 産業科学研究所, 助教 (30116032)
• Project Period (FY): 2007 – 2008
• Project Status: Completed (Fiscal Year 2008)
• Budget Amount: ¥4,420,000 (Direct Cost: ¥3,400,000、Indirect Cost: ¥1,020,000); Fiscal Year 2008: ¥1,560,000 (Direct Cost: ¥1,200,000、Indirect Cost: ¥360,000); Fiscal Year 2007: ¥2,860,000 (Direct Cost: ¥2,200,000、Indirect Cost: ¥660,000)
• Keywords: 遺伝子 / ストレス / 放射線 / X線 / 粒子線 / 生物物理 / シグナル伝達 / 転写因子 / 酸化ストレス / DNA / 鉄イオウクラスター / 酸化還元 / X線結晶解析

Research Abstract

本研究ではバクテリアにおけるSoxRの多様な生理機能を分子レベルで解明することを目的として、構造生物学的手法による研究として、X線構造解析により大腸菌SoxRの酸化ストレスを感知する機構や酸化還元によりおこる構造変化と、機能発現を原子レベルで解明した。SoxR-DNAの複合体では2.8A、SoxRでは3.2 Aの分解能の結果が得られた。全体の構造はバクテリアの水銀解毒に働く転写因子であるMerR familyとよく似た構造を示し、センサー部位である鉄イオウクラスタードメイン、DNA結合ドメイン、ダイマー生成ドメインから成ることが分かった。DNAが結合すると、DNA結合ドメイン約9o, 鉄イオウクラスターが約6o外側に傾き、全体として大きく構造変化した。酸化ストレスセンサー部位である鉄イオウクラスターは完全に溶媒に露出し非対称の環境にあり、もう一方のサブユニットと相互作用し安定化している。この非対称の環境の酸化還元によりタンパク質の構造変化とプロモーターとの相互作用が変化すると考えられる
SoxRに結合したDNAはワトソンークリック相互作用を保持したまま折れ曲がり、20塩基の長さが1塩基分だけ短くなり、その結果転写が活性化される。以上のX線構造解析により大腸菌SoxRの酸化ストレスを感知する機構および酸化還元によりおこる構造変化を解明した

Research Products

• [Journal Article] Intramolecular electron transfer processes in CuB-deficient cytochome bo studied by pulse radiolysis (2009)
  • Author(s): K. Kobayashi, S. Tagawa, and T. Mogi
  • Journal Title: J. Biochem 145 Pages: 685-691
  • Peer Reviewed
• [Journal Article] Function of a bound ubiquinone in Escherichia coli quinoprotein glucose dehydrogenase (2009)
  • Author(s): G. Mustafa, Y. Ichikawa, K. Kobayashi, C. T. Migita, S. Tagawa, and M. Yamada
  • Journal Title: BioFactors 32 Pages: 23-29
• [Journal Article] Pulse Radiolysis of Hexameric Nitrite Reductase Containing the two Type 1 Cu Sites in a Monomer (2008)
  • Author(s): S. Suzuki, T. Maetani, M. Nojiri, K. Yamaguchi, K. Kobayashi, and S. Tagawa
  • Journal Title: Bull. Chem. Soc Jpn 81 Pages: 1525-1527
  • NAID: 10024446895
  • Peer Reviewed
• [Journal Article] Formation of spectral intermediate G-C and A-T anion complex in duplex DNA studied by pulse radiolysis (2008)
  • Author(s): R. Yamagami, K. Kobayashi, and S. Tagawa
  • Journal Title: J. Am. Chem. Soc 130 Pages: 14772-14777
  • Peer Reviewed
• [Journal Article] Menaquinone as well as ubiquinone as a bound quinine crucial for catalytic activity and intramolecular electron transfer in in Escherichia coli membrane-bound glucose dehydrogenase (2008)
  • Author(s): G. Mustafa, C. T. Migita,Y. Ishikawa, K. Kobayashi, S. Tagawa, and M. Yamada,
  • Journal Title: J. Biol. Chem 283 Pages: 28169-28175
  • Peer Reviewed
• [Journal Article] Carbon- and nitrogen-centered radicals produced from L-lysine by radiationinduced oxidation : Chem (2008)
  • Author(s): T. Ito, S. Morimoto, S. Fujita, K. Kobayashi, S. Tagawa, and S. Nishimoto
  • Journal Title: Phys. Lett 462 Pages: 116-120
  • Peer Reviewed
• [Journal Article] Effect of Base Sequence and Deprotonation of Guanine Cation Radical in DNA (2008)
  • Author(s): K. Kobayashi, R. Yamagami, and S. Tagawa
  • Journal Title: J. Phys. Chem 112 Pages: 10752-10757
  • Peer Reviewed
• [Journal Article] Amino Acid Residues Interacting with Both the Bound Quinone and Coenzyme, Pyrroloquinoline Quinone, in Escherichia coli Membrane-bound Glucose Dehydrogenase (2008)
  • Author(s): G. Mustafa, Y. Ishikawa, K. Kobayashi, C. T. Migita, MD Elias, S. Nakamura, S. Tagawa, and M. Yamada
  • Journal Title: J. Biol. Chem 283 Pages: 22215-22221
  • Peer Reviewed
• [Journal Article] Crystal structure of the [2Fe-2S] oxidative-stress sensor SoxR bound to DNA, Proc. Natl (2008)
  • Author(s): S. Watanabe, A. Kita, K. Kobayashi, and K. Miki
  • Journal Title: Acad. Sci. U.S.A 105 Pages: 4121-4126
  • Peer Reviewed
• [Journal Article] Crystal structure of the [2Fe-2S] oxidative-stress sensor SoxR bound to DNA (2008)
  • Author(s): S. Watanabe, A. Kita, K. Kobayashi, K. Miki
  • Journal Title: Proc. Natl. Acad. Sci. U.S.A. 105 Pages: 4121-4126
  • Peer Reviewed
• [Journal Article] Amino Acid Residues Interacting with Both the Bound Quinone and Coenzyme, Pyrroloquinoline Quinone, in Escherichia coli Membrane-bound Glucose Dehydrogenase (2008)
  • Author(s): G. Mustafa, Y. Ishikawa, K. Kobavashi. C.. Migita, MD Elias, S. akamura, S. Tagawa, M. Yamada
  • Journal Title: J. Biol. Chem. 283 Pages: 22215-22221
  • Peer Reviewed
• [Journal Article] Effect of Base Sequence and Deprotonation of Guanine Cation Radical in DNA (2008)
  • Author(s): K. Kobayashi, R. Yamagami, S. Tagawa
  • Journal Title: J. Phys. Chem. B 112 Pages: 10752-10757
  • Peer Reviewed
• [Journal Article] Menaquinone as well as ubiquinone as a bound quinine crucial for catalytic activity and intramolecular electron transfer in in Escherichia coli membrane-bound glucose dehydrogenase (2008)
  • Author(s): G. Mustafa, C. T. Migita, Y. Ishikawa, K. Kobayashi, S. Tagawa, M. Yamada
  • Journal Title: J. Biol. Chem. 283 Pages: 28169-28175
  • Peer Reviewed
• [Journal Article] Effect of Base Sequence and Deprotonation of Guanine Cation Radical in DNA (2008)
  • Author(s): R. Yamagami, K, Kobayashi, S. Tagawa
  • Journal Title: J. Am. Chem. Soc. 130 Pages: 14772-14777
  • Peer Reviewed
• [Journal Article] Pulse Radiolysis of Hexameric Nitrite Reductase Containing the two Type 1 Cu Sites in a Monomer (2008)
  • Author(s): S. Suzuki, T. Maetani, M. Nojiri, K. Yamaguchi, K. Kobayashi, S. Tagawa
  • Journal Title: Bull. Chem. Soc. Jpn 81 Pages: 1525-1527
  • NAID: 10024446895
  • Peer Reviewed
• [Journal Article] In vitro preparation of iron-substituted human manganese superoxide dismutase : Possible toxic properties for mitochondria (2007)
  • Author(s): F. Yamakura, K. Kobayashi, S. Furukawa, and Y. Suzuki
  • Journal Title: Free Radical Biology and Medicine 43 Pages: 423-430
  • Peer Reviewed
• [Journal Article] Electron transfer Processes in Subunit I Mutants of Cytochrome bo Quinol Oxidase from Escherichia coli
  • Author(s): K. Kobayashi, S. Tagawa, and T. Mogi
  • Journal Title: Biosci. Biotechnol. Biochem (in press)
  • NAID: 10027543657
  • Peer Reviewed
• [Journal Article] Crystal structure of the [2Fe-2S] oxidative-stress sensor SoxR bound to DNA
  • Author(s): S. Watanabe, A. Kita, K. Kobayashi, K. Miki
  • Journal Title: Proc. Natl. Acad. Sci. U.S.A. In print
  • Peer Reviewed
• [Presentation] Dynamics of Excess Excess Electron in Duplex DNA Studied by Pulse Radiolysis (2008)
  • Author(s): R. Yamagami, K. Kobayashi, and S. Tagawa
  • Place of Presentation: Waterville, USA
• [Presentation] Effect of Base Sequence and Deprtonation of Guanine Cation Radical in DNA (2008)
  • Author(s): K. Kobayashi, R. Yamagami, and S. Tagawa
  • Organizer: Gordon Conference
  • Place of Presentation: Waterville, USA
• [Presentation] Dynamics of Guanine Radical Cations in Duplex DNA Studied by Pulse Radiolysis (2008)
  • Author(s): K. Kobayashi, R. Yamagami, and S. Tagawa
  • Organizer: 10th International Workshop Radiation Damage to DNA
  • Place of Presentation: Urabandai Japan
• [Presentation] Dynamics of Excess Electron in Duplex DNA Studied by Pulse Radiolysis (2008)
  • Author(s): R. Yamagami, K. Kobayashi, and S. Tagawa
  • Organizer: 10th International Workshop Radiation Damage to DNA
  • Place of Presentation: Urabandai Japan 2004066R 8
• [Presentation] Dynamics of Guanine Radical Cations in Duplex DNA Studied by Pulse Radiolysis (2008)
  • Author(s): K. Kobayashi Y. Yamagami, S. Tagawa
  • Organizer: 10^<th> International Workshop on Radiation Damage to DNA
  • Place of Presentation: 裏磐梯
• [Presentation] パルスラジオリシス法を用いたDNA酸化損傷の初期過程 (2007)
  • Author(s): 小林一雄、山上隆平、田川精一
  • Organizer: 日本放射線影響学会第50回大会
  • Place of Presentation: 幕張メッセ
  • Year and Date: 2007-11-16
• [Presentation] DNA鎖中生成するラジカルカチオンの脱プロトン過程のダイナミックス (2007)
  • Author(s): 小林一雄、山上隆平、田川精一
  • Organizer: 第50回放射線化学討論会
  • Place of Presentation: 京都大学
  • Year and Date: 2007-10-10