Basic technology development for Shark nanobody using antigen protein produced by wheat cell-free system

• Project/Area Number: 26640130
• Research Category: Grant-in-Aid for Challenging Exploratory Research
• Allocation Type: Multi-year Fund
• Research Field: System genome science
• Research Institution: Ehime University
• Principal Investigator: SAWASAKI TATSUYA 愛媛大学, プロテオサイエンスセンター, 教授 (50314969)
• Project Period (FY): 2014-04-01 – 2016-03-31
• Project Status: Completed (Fiscal Year 2015)
• Budget Amount: ¥3,900,000 (Direct Cost: ¥3,000,000、Indirect Cost: ¥900,000); Fiscal Year 2015: ¥1,950,000 (Direct Cost: ¥1,500,000、Indirect Cost: ¥450,000); Fiscal Year 2014: ¥1,950,000 (Direct Cost: ¥1,500,000、Indirect Cost: ¥450,000)
• Keywords: ナノボディ / 無細胞 / サメ / 抗体 / ドチザメ / 免疫 / 蛋白質 / トランスクリプトーム / 鮫 / 無細胞タンパク質合成

Outline of Final Research Achievements

We were successful to make an environment for the immunological experimental using three kinds of banded dogfishes that are harvested from off the coast of Matsuyama, Ehime Prefecture, calling Iyonada. Furthermore, we established good methods for handling, feeding, blood sampling and labeling of these sharks. Next we isolated a pure total RNA from bloods of these sharks and it is used for analysis of whole transcriptome by RNA seq. These results showed sequences of IgNAR coding shark nanobody.

Research Products

• [Journal Article] Characterization of RyDEN (C19orf66) as an interferon-stimulated cellular inhibitor against dengue virus replication. (2016)
  • Author(s): Suzuki Y, Chin WX, Han Q, Ichiyama K, Lee CH, Eyo ZW, Ebina H, Takahashi H, Takahashi C, Tan BH, Hishiki T, Ohba K, Matsuyama T, Koyanagi Y, Tan YJ, Sawasaki T, Chu1 JJ, Vasudevan SG, Sano K, Yamamoto N.
  • Journal Title: PLoS Pathogen Volume: 12 Issue: 1 Pages: 275-275
  • DOI: 10.1186/s12870-015-0660-9
  • NAID: 120005763386
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Journal Article] 無細胞技術を基盤とした抗GPCRモノクローナル抗体作製技術の開発 (2016)
  • Author(s): 竹田浩之、澤崎達也
  • Journal Title: BioIndustry Volume: 33 Pages: 56-62
• [Journal Article] Production of monoclonal antibodies against GPCR using cell-free synthesized GPCR antigen and biotinylated liposome-based interaction assay (2015)
  • Author(s): Takeda H, Ogasawara T, Ozawa T, Muraguchi A, Jih PJ, Morishita R, Uchigashima M, Watanabe M, Fujimoto T, Iwasaki T, Endo Y, Sawasaki T
  • Journal Title: Sci Rep Volume: 5 Issue: 1 Pages: 11333-11333
  • DOI: 10.1038/srep11333
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Journal Article] Members of the Plant CRK-superfamily are Capable of trans-/auto-Phosphorylation of Tyrosine Residues. (2015)
  • Author(s): Nemoto K, Takemori N, Seki M, Shinozaki K, Sawasaki T.
  • Journal Title: The Journal of Biological Chemistry Volume: 290 Issue: 27 Pages: 16665-16677
  • DOI: 10.1074/jbc.m114.617274
  • Peer Reviewed / Acknowledgement Compliant
• [Journal Article] High-throughput synthesis of stable isotope-labeled transmembrane proteins for targeted transmembrane proteomics using a wheat germ cell-free protein synthesis system. (2015)
  • Author(s): Takemori N, Takemori A, Matsuoka K, Morishita R, Matsushita N, Aoshima M, Takeda H, Sawasaki T, Endo Y, Higashiyama S.
  • Journal Title: Mol Biosyst. Volume: 11 Issue: 2 Pages: 361-365
  • DOI: 10.1039/c4mb00556b
  • Peer Reviewed / Open Access / Acknowledgement Compliant
• [Journal Article] MAPK Phosphatase 5 Expression Induced by Influenza and Other RNA Virus Infection Negatively Regulates IRF3 Activation and Type I Interferon Response. (2015)
  • Author(s): James SJ, Jiao H, Teh HY, Takahashi H, Png CW, Phoon MC, Suzuki Y, Sawasaki T, Xiao H, Chow VT, Yamamoto N, Reynolds JM, Flavell RA, Dong C, Zhang Y.
  • Journal Title: Cell Rep. Volume: 10 Issue: 10 Pages: 1722-1734
  • DOI: 10.1016/j.celrep.2015.02.030
  • Peer Reviewed / Open Access
• [Journal Article] In situ visualization of plasma cells producing antibodies reactive to Porphyromonas gingivalis in periodontitis: The application of the enzyme-labeled antigen method. (2014)
  • Author(s): Mizutani Y, Tsuge S, Takeda H, Hasegawa Y, Shiogama K, Onouchi T, Inada KI, Sawasaki T, Tsutsumi Y.
  • Journal Title: Molecular Oral Microbiology Volume: In press Issue: 4 Pages: 156-173
  • DOI: 10.1111/omi.12052
  • Peer Reviewed / Open Access
• [Journal Article] Involvement of Hepatitis C Virus NS5A Hyperphosphorylation Mediated by Casein Kinase I-α in Infectious Virus Production. (2014)
  • Author(s): Masaki T, Matsunaga S, Takahashi H, Nakashima K, Kimura Y, Ito M, Matsuda M, Murayama A, Kato T, Hirano H, Endo Y, Lemon SM, Wakita T, Sawasaki T, Suzuki T
  • Journal Title: J Virol Volume: 88(13) Issue: 13 Pages: 7541-55
  • DOI: 10.1128/jvi.03170-13
  • Peer Reviewed / Open Access / Acknowledgement Compliant
• [Journal Article] Pctaire1/Cdk16 promotes skeletal myogenesis by inducing myoblast migration and fusion. (2014)
  • Author(s): Shimizu K, Uematsu A, Imai Y, Sawasaki T.
  • Journal Title: FEBS Lett. Volume: 588 Issue: 17 Pages: 3030-3037
  • DOI: 10.1016/j.febslet.2014.05.060
  • Peer Reviewed / Acknowledgement Compliant
• [Journal Article] creenCap3: Improving prediction of caspase-3 cleavage sites using experimentally verified noncleavage sites. (2014)
  • Author(s): Fu SC, Imai K, Sawasaki T, Tomii K.
  • Journal Title: Proteomics. Volume: 14 Issue: 17-18 Pages: 2042-2046
  • DOI: 10.1002/pmic.201400002
  • Peer Reviewed / Open Access
• [Journal Article] Identification of RFPL3 protein as a novel E3 ubiquitin ligase modulating the integration activity of human immunodeficiency virus, type 1 preintegration complex using a microtiter plate-based assay. (2014)
  • Author(s): Tan BH, Suzuki Y, Takahashi H, Ying PH, Takahashi C, Han Q, Chin WX, Chao SH, Sawasaki T, Yamamoto N, Suzuki Y.
  • Journal Title: J Biol Chem. Volume: 289 Issue: 38 Pages: 26368-26382
  • DOI: 10.1074/jbc.m114.561662
  • Peer Reviewed
• [Journal Article] The apoptotic initiator caspase-8: its functional ubiquity and genetic diversity during animal evolution. (2014)
  • Author(s): Sakamaki K, Shimizu K, Iwata H, Imai K, Satou Y, Funayama N, Nozaki M, Yajima M, Nishimura O, Higuchi M, Chiba K, Yoshimoto M, Kimura H, Gracey AY, Shimizu T, Tomii K, Gotoh O, Akasaka K, Sawasaki T, Miller DJ.
  • Journal Title: Mol Biol Evol. Volume: 31 Issue: 12 Pages: 3282-3301
  • DOI: 10.1093/molbev/msu260
  • Peer Reviewed
• [Journal Article] Overexpression of the PAP1 transcription factor reveals a complex regulation of flavonoid and phenylpropanoid metabolism in Nicotiana tabacum plants attacked by Spodoptera litura. (2014)
  • Author(s): Mitsunami T., Nishihara M., Galis I., Alamgir K.M., Hojo Y., Fujita K., Sasaki N., Nemoto K., Sawasaki T., Arimura G.
  • Journal Title: PLoS One Volume: 9 Issue: 9 Pages: e108849-e108849
  • DOI: 10.1371/journal.pone.0108849
  • Peer Reviewed / Open Access
• [Journal Article] The ligand binding ability of dopamine D1 receptors synthesized using a wheat germ cell-free protein synthesis system with liposomes. (2014)
  • Author(s): Arimitsu E, Ogasawara T, Takeda H, Sawasaki T, Ikeda Y, Hiasa Y, Maeyama K.
  • Journal Title: Eur J Pharmacol. Volume: 745 Pages: 117-122
  • DOI: 10.1016/j.ejphar.2014.10.011
  • Peer Reviewed
• [Presentation] 抗膜タンパク質ウサギ高親和性モノクローナル抗体のエピトープ配列を由来とする新規アフィニティタグの開発 (2015)
  • Author(s): 矢野 智也, 竹田 浩之, 小澤 龍彦, 村口 篤, 澤崎 達也
  • Organizer: BMB2015（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: 神戸ポートアイランド（兵庫県・神戸市）
  • Year and Date: 2015-12-01
• [Presentation] 患者血清中の自己抗体を指標とした新規乳がん診断マーカーの開発 (2015)
  • Author(s): 松本 彩香, 竹田 浩之, 宮城 洋平, 澤崎 達也
  • Organizer: BMB2015（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: 神戸ポートアイランド（兵庫県・神戸市）
  • Year and Date: 2015-12-01
• [Presentation] 大規模生化学的解析はバイオインフォマティクス解析を凌駕するか？ (2014)
  • Author(s): 澤崎 達也
  • Organizer: 産総研・東京大学合同 創薬等支援技術基盤プラットフォーム(PDIS)ワークショップ ～実験とバイオインフォマティクスの協奏による構造生命科学～
  • Place of Presentation: 産総研臨海副都心センター別館（東京都江東区）
  • Year and Date: 2014-12-18
  • Invited
• [Presentation] コムギ無細胞技術を基盤とした化合物スクリーニング (2014)
  • Author(s): 澤崎 達也
  • Organizer: 第37回日本分子生物学会大会
  • Place of Presentation: パシフィコ横浜（神奈川県横浜市）
  • Year and Date: 2014-11-25 – 2014-11-27
  • Invited
• [Presentation] コムギ無細胞HTSを用いた創薬基盤技術 (2014)
  • Author(s): 澤崎 達也
  • Organizer: 第87回日本生化学会大会
  • Place of Presentation: 京都国際会議場（京都府京都市）
  • Year and Date: 2014-10-15 – 2014-10-18
  • Invited
• [Presentation] 無細胞技術を基盤とした自己抗体探索によるアンチエイジング医学のブレークスルーをめざして (2014)
  • Author(s): 澤崎 達也
  • Organizer: 第14回 日本抗加齢医学会総会
  • Place of Presentation: 大阪国際会議場（大阪府大阪市）
  • Year and Date: 2014-06-06 – 2014-06-08
  • Invited
• [Presentation] GPCR研究のためのコムギ無細胞系プロテオリポソーム技術 (2014)
  • Author(s): 澤崎 達也
  • Organizer: 第１１回GPCR研究会
  • Place of Presentation: 日本科学未来館 みらいCANホール（東京都江東区）
  • Year and Date: 2014-05-09 – 2014-05-10
  • Invited
• [Remarks] CELL-FREE Technology
  • URL: http://www.pros.ehime-u.ac.jp/cell-free/