Development of protein crystallization tag using crystalline lattice

• Project/Area Number: 15K13747
• Research Category: Grant-in-Aid for Challenging Exploratory Research
• Allocation Type: Multi-year Fund
• Research Field: Bio-related chemistry
• Research Institution: The University of Tokushima
• Principal Investigator: MAITA Nobuo 徳島大学, 先端酵素学研究所(次世代), 准教授 (00404046)
• Project Period (FY): 2015-04-01 – 2018-03-31
• Project Status: Completed (Fiscal Year 2017)
• Budget Amount: ¥3,900,000 (Direct Cost: ¥3,000,000、Indirect Cost: ¥900,000); Fiscal Year 2016: ¥2,340,000 (Direct Cost: ¥1,800,000、Indirect Cost: ¥540,000); Fiscal Year 2015: ¥1,560,000 (Direct Cost: ¥1,200,000、Indirect Cost: ¥360,000)
• Keywords: タンパク質結晶構造解析 / 結晶工学 / タンパク質結晶構造 / タンパク質結晶化 / Ｘ線結晶構造解析 / 結晶格子

Outline of Final Research Achievements

Crystal structure analysis of protein requires several hundreds of crystallization conditions screening to obtain high quality crystals, which is the most important bottleneck. I focused on proteins that make a porous crystal lattice and developed a method of genetic engineering fusion of guest proteins to the crystal lattice as it is to obtain its crystal structure. Eight proteins were selected as fusion guests to R1EN, which assembles a honeycomb lattice with an inner diameter of 11 nm. I prepared ubiquitin fusions (R1EN-Ub) with different linker lengths, and successfully crystallized under the same conditions as R1EN alone. I succeeded in structural analysis of three ubiquitin constructs. I could demonstrated the effectiveness of this R1EN fusion method.

Research Products

• [Journal Article] Protease-resistant modified human β-hexosaminidase B ameliorates symptoms in GM2 gangliosidosis model. (2016)
  • Author(s): Kitakaze K, Mizutani Y, Sugiyama E, Tasaki C, Tsuji D, Maita N, Hirokawa T, Asanuma D, Kamiya M, Sato K, Setou M, Urano Y, Togawa T, Otaka A, Sakuraba H, Itoh K
  • Journal Title: J Clin Invest Volume: in press Issue: 5 Pages: 1691-1703
  • DOI: 10.1172/jci85300
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Journal Article] Structural analysis of the TKB domain of ubiquitin ligase Cbl-b complexed with its small inhibitory peptide, Cblin (2016)
  • Author(s): Ohno A, Ochi A, Maita N, Ueji T, Bando A, Nakao R, Hirasaka K, Abe T, Teshima-Kondo S, Nemoto H, Okumura Y, Higashibata A, Yano S, Tochio H, Nikawa T
  • Journal Title: Arch Biochem Biophys Volume: 594 Pages: 1-7
  • DOI: 10.1016/j.abb.2016.02.014
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Journal Article] Creation of Customized Bioactivity within a 14-Membered Macrolide Scaffold: Design, Synthesis, and Biological Evaluation Using a Family-18 Chitinase (2015)
  • Author(s): Akihiro Sugawara, Nobuo Maita, Hiroaki Gouda, Tsuyoshi Yamamoto, Tomoyasu Hirose, Saori Kimura, Yoshifumi Saito, Hayato Nakano, Takako Kasai, Hirofumi Nakano, Kazuro Shiomi, Shuichi Hirono, Takeshi Watanabe, Hisaaki Taniguchi, Satoshi Omura, Toshiaki Sunazuka
  • Journal Title: Journal of Medicinal Chemistry Volume: 58 Issue: 12 Pages: 4984-4997
  • DOI: 10.1021/acs.jmedchem.5b00175
  • Peer Reviewed
• [Presentation] 高病原性インフルエンザ感染に関わる宿主酵素MSPLと阻害ペプチドとの複合体構造 (2017)
  • Author(s): 真板 綾子, 真板 宣夫, 奥村 裕司, 永野 ひかる, 有田 恭平, 平坂 勝也, 安倍 知紀, 近藤 茂忠, 二川 健
  • Organizer: 第71回日本栄養・食糧学会大会
• [Presentation] 高病原性インフルエンザウィルス感染に関わる宿主酵素MSPLとペプチド性阻害剤の複合体結晶構造 (2016)
  • Author(s): 43.真板綾子, 真板宣夫, 奥村裕司, 永野ひかる, 次田早希, 有田恭平, 坂東亜紀, 田畑考統, 平坂勝也, 安倍知紀, 近藤茂忠, 二川健
  • Organizer: 第16回蛋白質科学会年会
  • Place of Presentation: 福岡国際会議場、福岡県福岡市
• [Presentation] グルコースイソメラーゼの沈殿剤フリー結晶化と結晶構造解析 (2016)
  • Author(s): 鈴木良尚, 真板宣夫
  • Organizer: 日本物理学会第71回年次大会
  • Place of Presentation: 東北学院大学、宮城県仙台市
• [Presentation] α-L-イズロニダーゼの結晶構造より明らかになった新たなＮ型糖鎖の機能 (2015)
  • Author(s): 真板宣夫、月村考宏、谷口貴子、斎藤誠司、大野一樹、谷口寿章、櫻庭均
  • Organizer: 第15回日本蛋白質科学会年会
  • Place of Presentation: あわぎんホール、徳島県徳島市
• [Presentation] 新規組換えリソソーム酵素の創製とリソソーム病治療への応用 (2015)
  • Author(s): 伊藤孝司, 西岡宗一郎, 辻大輔, 東哲也, 真板宣夫, 小林功, 瀬筒秀樹, 湯本史明, 原囿景, 石明子, 川崎ナナ
  • Organizer: 第15回日本蛋白質科学会年会
  • Place of Presentation: あわぎんホール、徳島県徳島市
• [Presentation] 改変型ヒトβ- Hexosaminidase のGM2 蓄積症モデルに対する治療効果の評価 (2015)
  • Author(s): 41.北風圭介, 水谷安通, 杉山栄二, 真板宣夫, 広川貴次, 瀬藤光利, 櫻庭均, 伊藤孝司
  • Organizer: 第38回日本分子生物学会年会・第88回日本生化学会大会 合同大会
  • Place of Presentation: 神戸国際会議場、兵庫県神戸市
• [Remarks] 徳島大学 教育研究者総覧
  • URL: http://pub2.db.tokushima-u.ac.jp/ERD/person/166156/profile-ja.html
• [Remarks] 徳島大学 教育・研究者情報データベース
  • URL: http://pub2.db.tokushima-u.ac.jp/ERD/person/166156/profile-ja.html
• [Patent(Industrial Property Rights)] 多孔性タンパク質結晶を利用したタンパク質結晶化 (2018)
  • Inventor(s): 真板 宣夫
  • Industrial Property Rights Holder: 徳島大学
  • Industrial Property Rights Type: 特許
  • Filing Date: 2018
  • Overseas
• [Patent(Industrial Property Rights)] 1.多孔性タンパク質結晶を利用したタンパク質結晶化 (2017)
  • Inventor(s): 真板 宣夫
  • Industrial Property Rights Holder: 徳島大学
  • Industrial Property Rights Type: 特許
  • Industrial Property Number: 2017-042456
  • Filing Date: 2017-03-07