Elucidation of the activation mechanism of an unfolded protein response sensor protein PERK

• Project/Area Number: 24710245
• Research Category: Grant-in-Aid for Young Scientists (B)
• Allocation Type: Multi-year Fund
• Research Field: Living organism molecular science
• Research Institution: The University of Tokyo
• Principal Investigator: NAKAMURA AKIRA 東京大学, 農学生命科学研究科, 特任助教 (40432356)
• Project Period (FY): 2012-04-01 – 2014-03-31
• Project Status: Completed (Fiscal Year 2013)
• Budget Amount: ¥4,680,000 (Direct Cost: ¥3,600,000、Indirect Cost: ¥1,080,000); Fiscal Year 2013: ¥1,690,000 (Direct Cost: ¥1,300,000、Indirect Cost: ¥390,000); Fiscal Year 2012: ¥2,990,000 (Direct Cost: ¥2,300,000、Indirect Cost: ¥690,000)
• Keywords: 活性発現の分子機構 / タンパク質 / X線結晶構造解析 / 小胞体ストレス応答 / 結晶構造解析

Research Abstract

To elucidate the activation mechanism of an unfolded protein response (ER stress response) sensor protein, PERK, the luminal domain of human PERK and human BiP, which is the regulator of PERK also known as an Hsp70 molecular chaperone, were expressed in Escherichia coli cells and purified with high purity. The interaction of these two proteins were examined by GST pull-down assays. The crystallization trials for the target proteins have not succeeded to date.

Research Products

• [Journal Article] Crystal Structure of a novel N-substituted L-amino acid dioxygenase from Burkholderia ambifaria AMMD (2013)
  • Author(s): Qin, H.-M., Miyakawa, T., Jia, M.-Z., Nakamura, A., Ohtsuka, J., Xue, Y.-L., Kawashima, T., Kasahara, T., Hibi, M., Ogawa, J., Tanokura, M.
  • Volume: 8(5)
• [Journal Article] Structure analysis of archaeal AMP phosphorylase reveals two unique modes of dimerization (2013)
  • Author(s): Nishitani, Y., Aono, R., Nakamura, A., Sato, T., Atomi, H., Imanaka, T., Miki K.
  • Journal Title: J. Mol. Biol Volume: 425(15) Pages: 2709-2721
• [Journal Article] Expression, purification, crystallization and preliminary X-ray analysis of a novel N-substituted branched-chain L-amino-acid dioxygenase from Burkholderia ambifaria AMMD (2012)
  • Author(s): Qin, H.-M., Miyakawa, T., Nakamura, A., Xue, Y.-L., Kawashima, T., Kasahara, T., Hibi, M., Ogawa, J., Tanokura, M.
  • Journal Title: Acta Crystallogr. Sect. F Struct. Biol. Cryst. Commun Volume: 68(Pt 9) Pages: 1067-1069
• [Journal Article] High pressure refolding, purification, and crystallization of flavin reductase from Sulfolobus tokodaii strain 7 (2012)
  • Author(s): Okai, M., Ohtsuka, J., Asano, A., Guo, L., Miyakawa, T., Miyazono, K., Nakamura, A., Okada, A., Zheng, H., Kimura, K., Nagata, K., Tanokura, M.
  • Journal Title: Protein Expr. Purif Volume: 84(2) Pages: 214-248
• [Journal Article] Dynamic, ligand-dependent conformational change triggers reaction of ribose-1,5-bisphosphate isomerase from Thermococcus kodakarensis KOD1 (2012)
  • Author(s): Nakamura, A., Fujihashi, M., Aono, R., Sato, T., Nishiba, Y., Yoshida, S., Yano, A., Atomi, H., Imanaka, T., Miki, K.
  • Journal Title: J. Biol. Chem Volume: 287(25) Pages: 20784-20796
• [Presentation] 磁気力場を利用したタンパク質結晶の高品質化と結晶成長のリアルタイム観察 (2014)
  • Author(s): 中村顕, 大塚淳, 廣田憲之, 和田仁, 田之倉優
  • Organizer: 日本農芸化学会2014年度会
  • Place of Presentation: 明治大学、神奈川
  • Year and Date: 2014-03-28
• [Presentation] 磁気力場利用による高品質タンパク質結晶の取得とリアルタイム観察 (2013)
  • Author(s): 中村顕, 大塚淳, 廣田憲之, 和田仁, 田之倉優
  • Organizer: 日本分子生物学会第36回年会
  • Place of Presentation: 神戸ポートアイランド、兵庫
  • Year and Date: 2013-12-03
• [Presentation] 高磁気力を利用した高効率・高品位タンパク質結晶生成システムの開発 (2013)
  • Author(s): 廣田憲之, 和田仁, 清原元輔, 田之倉優, 鈴木榮一郎, 喜田昭子, 岡田秀彦, 大出孝博, 中村顕, 大塚淳, 柏木立己, 沼本修孝
  • Organizer: 日本磁気科学会第8回年会
  • Place of Presentation: 東北大学,宮城
  • Year and Date: 2013-11-22
• [Presentation] 磁気力場利用によるタンパク質結晶化のその場観察と高品質結晶の取得 (2013)
  • Author(s): 中村顕, 大塚淳, 廣田憲之, 和田仁, 田之倉優
  • Organizer: 日本結晶学会平成25年度年会
  • Place of Presentation: 熊本大学,熊本
  • Year and Date: 2013-10-12
• [Presentation] Protein crystallization and observation of crystal growth in a magnetic field (2013)
  • Author(s): Nakamura, A., Ohtsuka, J., Tanokura, M.
  • Organizer: 4^<th> International Symposium on Diffraction Structural Biology (ISDSB2013)
  • Place of Presentation: Nagoya, Japan
  • Year and Date: 2013-05-28
• [Presentation] シロイヌナズナ由来乾燥・高温ストレス応答転写因子DREB2Aの結晶構造解析 (2013)
  • Author(s): 中村顕, 浅野敦子, 宮川拓也, 澤野頼子, 佐久間洋, 溝井順哉, 篠崎和子, 田之倉優
  • Organizer: 日本農芸化学会2013年度大会
  • Place of Presentation: 東北大学,宮城
  • Year and Date: 2013-03-25
• [Presentation] Crystal structure of a novel branched-chain amino acid dioxygenase from Burkholderia ambifaria AMMD (2012)
  • Author(s): Qin, H.-M., Miyakawa, T., Nakamura, A., Kawashima, T., Hibi, M., Ogawa, J., Tanokura, M.
  • Organizer: American Crystallographic Association, Inc. Annual Meeting
  • Place of Presentation: Boston, USA
  • Year and Date: 2012-07-29
• [Presentation] Protein Crystallization in a high gradient magnetic field (2012)
  • Author(s): Nakamura, A., Ohtsuka, J., Miyazono, K., Yamamura, A., Kubota, K., Hirose, R., Kashiwagi, T., Suzuki, E., Hirota, N., Wada, H., Ataka, M., Sawano, Y., Tanokura, M.
  • Organizer: 5^<th> International Workshop on Materials Analysis and Processing in Magnetic Fields (MAP5)
  • Place of Presentation: Autrans, France
  • Year and Date: 2012-05-15
• [Book] 「超伝導現象と高温超伝導体」(第2編材料と応用第7章計測診断機器への応用第2節超伝導磁石の強力磁場を用いたタンパク質結晶の高品質化機器の開発) (2013)
  • Author(s): 中村顕, 田之倉優
  • Total Pages: 542
  • Publisher: 株式会社エヌ・ティー・エス
• [Patent(Industrial Property Rights)] 磁気力場発生装置 (2012)
  • Inventor(s): 岡田 秀彦, 廣田 憲之, 和田 仁, 松 本 真治, 清原 元輔, 大出 孝博, 清原 耕輔, 田之倉優, 中村 顕, 大塚 淳
  • Industrial Property Rights Holder: 独立行政法人物質・材料研究機構, 株式会社清原光学, 国立大学法人東京大学
  • Industrial Property Rights Type: 特許
  • Industrial Property Number: 2012-109943
  • Filing Date: 2012-05-11