Structure and function of two different threonyl-tRNA synthetases of crenarchaea

2010 Fiscal Year Final Research Report

• Project/Area Number: 20570104
• Research Category: Grant-in-Aid for Scientific Research (C)
• Allocation Type: Single-year Grants
• Section: 一般
• Research Field: Structural biochemistry
• Research Institution: Iwaki Meisei University
• Principal Investigator: TAKENAKA Akio Iwaki Meisei University, 薬学部・大学院・生命理工学研究科, 教授・特任教授 (80016146)
• Co-Investigator(Kenkyū-buntansha): SEKIGUCHI Takeshi いわき明星大学, 科学技術学部, 教授 (00108193)
• Co-Investigator(Renkei-kenkyūsha): MORAS Dino IGBMC-CNRS, France, Director
• Project Period (FY): 2008 – 2010
• Keywords: アミノアシルtRNA合成酵素 / タンパク質合成系 / X線結晶解析 / クレン古細菌 / トレオニルtRNA合成酵素

Research Abstract

Threonyl-tRNA synthetase (ThrRS) plays an essential role in protein synthesis by catalyzing aminoacylation of tRNA^<Thr> and by editing misacylation. ThrRS is generally composed of the three domains, an N-terminal editing domain, a catalytic domain and an anticodon-binding domain. In general, an organism possesses one kind of gene for ThrRS. However, it has been recently found that some organisms have two different genes for ThrRS in the genome, suggesting that their proteins ThrRS-1 and ThrRS-2 function separately and complement each other in threonylation of tRNA^<Thr> ; one for catalysis and the other for editing of misacylated Ser-tRNA^<Thr>. From sequence alignment of ThrRS-1 and ThrRS-2 with those of other organisms, we have found that the editing domain in ThrRS from archaea are different from those in bacteria and eukaryotes. Furthermore, in several creanarchaea including Aeropyrum pernix K1 and Sulfolobus tokodaii strain 7, each contains two genes encoding either the catalytic or the editing domains of ThrRS. In order to clarify the structural basis for the evolutionary divergence, the two types of ThrRSs from crenarchaea Aeropyrum pernix and those from Sulfolobus tokodaii (ApThrRS-1, ApThrRS-2, StThrRS-1 and StThrRS-2) have been overexpressed in Eschericha coli, purified and successfully crystallized by the hanging-drop vapor-diffusion method. Diffraction data were collected, and the structure of selenomethionine-labeled ApThrRS-1 crystal has been solved usingthe MAD method, and the atomic parameters were refined by the least-squares method at 2.3 resolution. ApThrRS-1 is a dimeric enzyme composed of two identical subunits, each containing two domains for the catalytic reaction and for anticodon binding. The essential editing domain is completely missing as expected. These structural features reveal that ThrRS-1 catalyzes only the aminoacylation of the cognate tRNA, suggesting the necessity of the second enzyme ThrRS-2 for editing. Since the N-terminal sequence of ApThrRS-2 is similar to the sequence of the editing domain of ThrRS from Pyrococcus abyssi, ApThrRS-2 has been expected to catalyze deaminoacylation of a misacylated serine moiety at the CCA terminus. The tertiary structure of ThrRS-2 was constructed based on the sequence alignment with similar protein, based on which the dimer structure formed between the two editing domains. Gel shift assay experiments shows that ThrRS-1 and ThrRS-2 do not interact to form a dimer and that each forms a self dimer. Furthermore, we have found that tRNA^<Thr> interacts to both, but ThrRS-1 is strongly bound to tRNA^<Thr>. Finally we succeeded to crystallize the complexes between tRNA^<Thr> and ThrRS-1 and between tRNA and ThrRS-2, respectively. X-Ray analyses of these crystals will reveal the interaction geometry from which the specificities of ThrRS-1 and ThrRS-2 to tRNA^<Thr> will be clarified.

Research Products

• [Journal Article] Actinohivin: specific amino acid residues essential foranti-HIV activity (2010)
  • Author(s): Masaru Tsunoda, Kaoru Suzuki, Akio Takenaka, Takeshi Sekiguchi, Hideaki Umeyama, Jun Hirabayashi, Satoshi Omura, Haruo Tanaka
  • Journal Title: J.Antibiot. 106 Pages: doi:10.1038/ja.2010
• [Journal Article] Insights into the DNA stabilizing contributions of a bicyclic cytosine analogue: crystal structures of DNA duplex containing 7,8-dihydropyrido[2,3-d]pyrimidin-2-on (2010)
  • Author(s): Ella CzarinaMagat Juan, Satoru Shimizu, Xiao Ma, Taizo Kurose, Tsuyoshi Haraguchi1, Fang Zhang, Masaru Tsunoda, Akihiro Ohkubo, Mitsuo Sekine, Takayuki Shibata, Christopher L. Millington, David M. Williams, Akio Takenaka
  • Journal Title: Nucl.Acids Res. 38 Pages: 6737-6745
• [Journal Article] Insights into the mutagenic structures of DNA damaged by hydroxyl radical, XI : crystal structures of DNA duplexes containing 5-formyluracil (2010)
  • Author(s): Masaru Tsunoda, Takeshi Sakaue, Satoko Naito, Tomoko Sunami, Naoko Abe, Yoshihito Ueno, Akira Matsuda, Akio Takenaka
  • Journal Title: J.Nucl.Acids Pages: doi:10.4061/2010/107289
• [Journal Article] Crystallo-graphic study of wild type carbonic anhydrase αCA1 from Chlamydomonas reinhardtii (2010)
  • Author(s): Kaoru Suzuki, Satoru Shimizu, Ella Czarina Magat Juan, Takahiro Miyamoto, Fang Zhang, Md. Mominul Hoque, Yoshiteru Sato, Masaru Tsunoda, Takeshi Sekiguchi, Akio Takenaka
  • Journal Title: Acta Crystallogr. F66 Pages: 1082-1085
• [Journal Article] Mechanism by which the lectin actinohivin blocks HIV infection of target cells (2009)
  • Author(s): Haruo Tanaka, Harumi Chiba, Junji Inokoshi, Atsushi Kuno, Takahiro Sugai, Atsushi Takahashi, Yukishige Ito, Masaru Tsunoda, Kaoru Suzuki, Akio Takenaka, Takeshi Sekiguchi, Hideaki Umeyama, Jun Hirabayashi, Satoshi Omura
  • Journal Title: Proc.Natl.Acad.Sci.USA 106 Pages: 15633-15638
• [Journal Article] Two complementary enzymes for threonylation of tRNA in crenarchaeota; crystal structure of Aeropyrum pernix threonyl-tRNA synthetase lacking a cis-editing domain (2009)
  • Author(s): Satoru Shimizu, Ella Czarina Magat Juan, Yu-ichiro Miyashita, Yoshiteru Sato, Md. Mominul Hoque, Kaoru Suzuki, Tsubasa Sagara, Masaru Tsunoda, Takeshi Sekiguchi, Anne-Catherine Dock-Bregeon, Dino Moras, Akio Takenaka
  • Journal Title: J.Mol.Biol. 394 Pages: 286-296
• [Journal Article] Crystallization preliminary crystallographic studies of putative RNA 3'-terminal phosphate cyclase from crenarchaeon Sulfolobus tokodaii (2009)
  • Author(s): Satoru Shimizu, Masanori Ohki, Nami Okubo, Kaoru Suzuki, Masaru Tsunoda, Takeshi Sekiguchi, Akio Takenaka
  • Journal Title: Acta Crystallogr. F65 Pages: 565-570
• [Journal Article] Crystal structure of D-3-hydroxybutyrate dehydrogenase, prepared in the presence of the substrate D-3-hydroxybutyrate NAD^+ (2009)
  • Author(s): Md. Mominul Hoque, Satoru Shimizu, Ella Czarina Magat Juan, Yoshiteru Sato, Md. Tofazzal Hossain, Tamotsu Yamamoto, Shigeyuki Imamura, Kaoru Suzuki, Hitoshi Amano, Takeshi Sekiguchi, Masaru Tsunoda, Akio Takenaka
  • Journal Title: Acta Crystallogr. F65 Pages: 331-335
• [Journal Article] Crystallographic study on two types of putative threonyl-tRNA synthetases from crenarchaea Aeropyrum pernix those from Sulfolobus tokodaii (2008)
  • Author(s): Satoru Shimizu, Ella Czarina Magat Juan, Yu-ichiro Miyashita, Yoshiteru Sato, Kaoru Suzuki, Masataka Yogiashi, Masaru Tsunoda, Anne-Catherine Dock-Bregeon, Dino Moras, Takeshi Sekiguchi, Akio Takenaka
  • Journal Title: Acta Crystallogr. F64 Pages: 903-910
• [Journal Article] Structures of Arthrobacter globiformis urate oxidase-lig complexes (2008)
  • Author(s): Ella Czarina Magat Juan, Md Mominul Hoque, Satoru Shimizu, Md Tofazzal Hossain, Tamotsu Yamamoto, Shigeyuki Imamura, Kaoru Suzuki, Masaru Tsunoda, Hitoshi Amano, Takeshi Sekiguchi, Akio Takenaka
  • Journal Title: Acta Crystallogr. D64 Pages: 815-822
• [Journal Article] The structures of Alcaligenes faecalis D-3-hydroxybutyrate dehydrogenase before after NAD^+ acetate binding suggest a dynamical reaction mechanism as a member of the SDR family (2008)
  • Author(s): Md Mominul Hoque, Satoru Shimizu, Md Tofazzal Hossain, Tamotsu Yamamoto, Shigeyuki Imamura, Kaoru Suzuki, Masaru Tsunoda, Hitoshi Amano, Takeshi Sekiguchi, Akio Takenaka
  • Journal Title: Acta Crystallogr. D64 Pages: 496-505
• [Presentation] Crystal structure of anti-HIV actinohivin in complexwith 1, 2-mannobiose (2010)
  • Author(s): M.M.Hoque, K.Suzuki, M.Tsunoda, A.Takahashi, T.Sekiguchi, H.Tanaka, A.Takenaka
  • Organizer: 日本結晶学会年会講演要旨集,126
  • Place of Presentation: 大阪
  • Year and Date: 20101203-20101205
• [Presentation] Crystal structures of DNA duplexes containing O^6-carboxymethyl-2'-deoxy guanosine as an analogueof damaged residue, relevant to gastrointestinal cancer (2010)
  • Author(s): F.Zhang, K.Suzuki, M.M.Hoque, M.Tsunoda, C.L.Millington, D.M.Williams, A.Takenaka
  • Organizer: 日本結晶学会年会講演要旨集, 127
  • Place of Presentation: 大阪
  • Year and Date: 20101203-20101205
• [Presentation] An insight into the pairing geometry of DNA duplexes containing O^6-carboxymethylguanine, a damaged base analogue relevant to gastrointestinal cancer (2010)
  • Author(s): Zhang Fang, Kaoru Suzuki, Md.Mominul Hoque, Masaru Tsunoda, Christopher L.Millington, David M.Williams, Akio Takenaka
  • Organizer: AsCA2010 (The 10^<th> Conference of the Asian Crystallographic Association), Abstract 169
  • Place of Presentation: Busan(Korea)
  • Year and Date: 20101031-20101103
• [Presentation] An insight into the pairing geometry of DNA duplexes containing O^6-carboxymethylguanine, an analogue damaged base relevant to gastrointestinal cancer (2010)
  • Author(s): Fang Zhang, Kaoru Suzuki, Md. Mominul Hoque, Masaru Tsunoda, Christopher L. Millington, David M.Williams, Akio Takenaka
  • Organizer: ISNAC2010 (The 37^<th> International Symposium in Nucleic Acids Chemistry 2010), Abstract, 74-75, Nov.10-12
  • Place of Presentation: Yokohama
  • Year and Date: 20100000
• [Presentation] Two complementary enzymes for threonylation of tRNA in crenarchaeota ; crystal structure of Aeropyrum pernix threonyl-tRNA synthetase lacking a cis-editing domain (2010)
  • Author(s): A.Takenaka, S.Shimizu, Y.Sato, E.C.M.Juan, Y.Miyashita, T.Sagara, K.Suzuki, M.Tsunoda, T.Sekiguchi,, A.C.D.Bregeon, D.Moras
  • Organizer: IGBMC(フランス国立遺伝学分子細胞生物学研究所)セミナー
  • Year and Date: 2010-10-23
• [Presentation] X-Ray analyses of two evolutionarily different threonyl-tRNA synthetases which perform a function by supplementing their defects to each other in crenarchaea (2010)
  • Author(s): A.Takenaka, S.Shimizu, Y.Sato, E.C.M.Juan, Y.Miyashita, T.Sagara, K.Suzuki, M.Tsunoda, T.Sekiguchi, A.C.D.Bregeon, D.Moras
  • Organizer: Centre for Chemical Biologyセミナー
  • Place of Presentation: Sheffield大学
  • Year and Date: 2010-10-18
• [Presentation] 古細菌Sulfolobus tokodaii由来トレオニルtRNA合成酵素の精製と結晶化構 (2010)
  • Author(s): 斉藤彰浩, 鈴木薫, 関口武司, 角田大, 竹中章郎
  • Organizer: 平成22年度化学系学協会東北大会 講演予稿集186
  • Place of Presentation: 盛岡
  • Year and Date: 2010-09-25
• [Presentation] X-Ray analyses of two evolutionarily different threonyl-tRNA synthetases which perform a function by supplementing their defects to each other in crenarchaea (2009)
  • Author(s): Satoru Shimizu, Yoshiteru Sato, Ella Czarina Magat Juan, Yu-ichiro Miyashita, Tsubasa Sagara, Kaoru Suzuki, Masaru Tsunoda, Takeshi Sekiguchi, Anne-Catherine Dock-Bregeon, Dino Moras, Akio Takenaka
  • Organizer: Acta Crystallogr., A65, s17-s18
  • Year and Date: 20090000
• [Presentation] Crystal structures of actinohivin, an anti-HIV proteinfrom an actinomycete, its complex with mannobiose (2009)
  • Author(s): 角田大, 鈴木薫, 相良翼, 高橋淳, 猪腰淳嗣, 大村智, 関口武司, 田中晴雄, 竹中章郎
  • Organizer: Acta Crystallogr., A65, s149-s150
  • Year and Date: 20090000
• [Presentation] 新規抗HIVタンパク質アクチノヒビンのマンノビオースとの共結晶化 (2009)
  • Author(s): 相良翼, 斉藤彰浩, 高橋淳, 鈴木薫, 関口武司, 角田大, 田中晴雄, 竹中章郎
  • Organizer: 日本結晶学会2009年度年会,講演要旨集116
  • Year and Date: 20090000
• [Presentation] 放線菌より単離した新規anti-HIV蛋白質アクチノヒビンならびにそのマンノビオースとの複合体のX線構造 (2009)
  • Author(s): 角田大, 鈴木薫, 相良翼, 高橋淳, 猪腰淳嗣, 大村智, 関口武司, 田中晴雄, 竹中章郎
  • Organizer: 第82回日本生化学会大会,要旨集4T9p-124P-629
  • Place of Presentation: 神戸
  • Year and Date: 20090000
• [Presentation] RNAプロセシングに関与するタンパク質,RNA3'-末端リン酸基環状化酵素の反応機構 (2009)
  • Author(s): 清水了, 大木正則, 大久保奈弥, 鈴木薫, 角田大, 関口武司, 竹中章郎
  • Organizer: 第11回RNA ミーティング,要旨集6
  • Place of Presentation: 新潟
  • Year and Date: 20090000
• [Presentation] 国際結晶学連合大会IUCr2008組織委員会の外務委員会報告 (2009)
  • Author(s): 坂田誠, 佐々木聡, 竹中章郎, 三木邦夫, 高田昌樹, 吉朝朗
  • Organizer: 日本結晶学会誌51,22-28
  • Year and Date: 20090000
• [Presentation] Structure reaction mechanism of an enzyme RNA 3'-terminal phosphate cyclase in tRNA splicing (2009)
  • Author(s): 竹中章郎, 大木正則, 大久保奈弥, 鈴木薫, 角田大, 関口武司, 清水了
  • Organizer: tRNA Workshop, Aveiro, Portugal
  • Year and Date: 20090000
• [Presentation] X-Ray Analyses of Two Evolutionary Different Threonyl-tRNA Synthetases Which Prefer a Functionby Supplementing Their Defects to Each Other in Crenarchaea (2009)
  • Author(s): S.Shimizu, Y.Sato, E.C.M.Juan, Y.Miyashita, T.Sagara, K.Suzuki, M.Tsunoda, T.Sekiguchi, A.C.D.Bregeon, D.Moras, A.Takenaka
  • Organizer: 25th European Crystallographic Meeting, ECM25 (Istanbul) Abstract
  • Year and Date: 20090000
• [Presentation] Crystal Structures of Actinohivin, an Anti-HIV Protein from an Actinomycete, its Complex (2009)
  • Author(s): 角田大, 鈴木薫, 相良翼, 高橋淳, 猪腰淳嗣, 大村智, 関口武司, 田中晴雄, 竹中章郎
  • Organizer: 25th European Crystallographic Meeting, ECM25 (Istanbul) Abstract
  • Year and Date: 20090000
• [Presentation] Crystal structures of DNA duplexes stabilized by bicyclic-C residues (2008)
  • Author(s): Tsuyoshi Haraguchi, Satoru Shimizu, Xiao Ma, Taizo Kurose, Ella Czarina Magat Juan, Akihiro Ohkubo, Mitsuo Sekine, Takayuki Shibata, Christopher L. Millington, David M. Williams, Akio Takenaka
  • Organizer: Nucleic Acids Symp Ser.52, 127-128
  • Year and Date: 20080000
• [Presentation] Crystal structures of RNA 3'-terminal phosphate cyclase its complexes with Mg^<2+>+ATP, ATP or Mn^<2+> (2008)
  • Author(s): Satoru Shimizu, Masanori Ohki, Nami Ohkubo, Kaoru Suzuki, Masaru Tsunoda, Takeshi Sekiguchi, Akio Takenaka
  • Organizer: Nucleic Acids Symp Ser.52, 221-222
  • Year and Date: 20080000
• [Presentation] Crystal structure of α-carbonic anhydrase from Chlamydomonas reinhardtii (2008)
  • Author(s): Kaoru Suzuki, Satoru Shimizu, Risa Ohbayashi, Yoshiteru Sato, Akio Takenaka, Takeshi Sekiguchi, Shi-Yuan Yang
  • Organizer: Acta Cryst., A64, C263
  • Year and Date: 20080000
• [Presentation] Two threonyl-tRNA synthetases with complementaryfunctions ; Crystal structure of ThrRS-1 fromAeropyrum pernix (2008)
  • Author(s): Akio Takenaka, Satoru Shimizu, Yu-ichiro Miyashita, Yoshiteru Sato, Ella Czarina Magat Juan, Kaoru Suzuki, Masaru Tsunoda, Anne-Catherine Dock-Bregeon, Dino Moras, Takeshi Sekiguchi
  • Organizer: Acta Cryst., A64, C311
  • Year and Date: 20080000
• [Presentation] RNA splicing related proteins; Crystal structure of RNA 3'-terminal phosphate cyclase (2008)
  • Author(s): Satoru Shimizu, Masanori Ohki, Nami Ohkubo, Kaoru Suzuki, Masaru Tsunoda, Takeshi Sekiguchi, Akio Takenaka
  • Organizer: Acta Cryst., A64, C307
  • Year and Date: 20080000
• [Presentation] The substrate recognition the catalytic reaction mechanisms of D-3hydroxybutyrate dehydrogenase (2008)
  • Author(s): Md Mominul Hoque, Satoru Shimizu, Md Tofazzal Hossain, Tamotsu Yamamoto, Shigeyuki Imamura, Kaoru Suzuki, Masaru Tsunoda, Hitoshi Amano, Takeshi Sekiguchi, Akio Takenaka
  • Organizer: Acta Cryst., A64, C252
  • Year and Date: 20080000
• [Presentation] X-Ray analyses of DNA duplexes stabilized by bicyclic-C residues (2008)
  • Author(s): Tsuyoshi Haraguchi, Satoru Shimizu, Xiao Ma, Taizo Kurose, Ella Czarina Magat Juan, Akihiro Ohkubo, Mitsuo Sekine, Takayuki Shibata, Christopher L. Millington, David M. Williams, Akio Takenaka
  • Organizer: Acta Crystallogr., A64, C298
  • Year and Date: 20080000
• [Presentation] Crystal structure of actinohivin ; A novel anti-humanimmunodeficiency virus protein (2008)
  • Author(s): 角田大, 鈴木薫, 相良翼, 高橋淳, 猪腰淳嗣, 大村智, 関口武司, 竹中章郎, 田中晴雄
  • Organizer: Acta Crystallogr., A64, C237
  • Year and Date: 20080000
• [Book] 耐熱酵素のさらなる安定性向上ISBN978-4-86043-271-3, 518-522(酵素利用技術大系-基礎・解析から改変・高機能化・産業利用まで-)(小宮山眞監修)東京 (2010)
  • Author(s): 竹中章郎・関口武司著
  • Publisher: 株式会社エヌ・ティー・エス
• [Book] 分子の個性を理解する(研究の目指すところ-夢を語る-,75-86 (2010)
  • Author(s): 竹中章郎
  • Publisher: いわき明星大学大学院理工学研究科
• [Book] キトサナーゼの基質特異性と構造(キチン・キトサン開発技術) (2009)
  • Author(s): 安達渉, 中村聡, 竹中章郎
• [Book] タンパク質のX線結晶解析法((J. Drenth著))(第3版) (2008)
  • Author(s): 竹中章郎, 勝部幸輝, 笹田義夫, 若槻壮一共訳
  • Publisher: シュプリンガー・ジャパン
• [Remarks] ホームページ等
  • URL: http://www.iwakimu.ac.jp/~BioSC/NewScience/Kakenhi-1/Kakenhi-2008-2010