Development of an efficient protein production system of various virus antigens for diagnostic and vaccine reagents in yeast

• Project/Area Number: 24658096
• Research Category: Grant-in-Aid for Challenging Exploratory Research
• Allocation Type: Multi-year Fund
• Research Field: Applied biochemistry
• Research Institution: Yamaguchi University
• Principal Investigator: AKADA Rinji 山口大学, 医学(系)研究科(研究院), 教授 (20201882)
• Project Period (FY): 2012-04-01 – 2015-03-31
• Project Status: Completed (Fiscal Year 2014)
• Budget Amount: ¥4,030,000 (Direct Cost: ¥3,100,000、Indirect Cost: ¥930,000); Fiscal Year 2014: ¥1,300,000 (Direct Cost: ¥1,000,000、Indirect Cost: ¥300,000); Fiscal Year 2013: ¥1,560,000 (Direct Cost: ¥1,200,000、Indirect Cost: ¥360,000); Fiscal Year 2012: ¥1,170,000 (Direct Cost: ¥900,000、Indirect Cost: ¥270,000)
• Keywords: ウイルス表面タンパク質 / 抗原 / HBsL / Saccharomyces cerevisiae / Kluyveromyces marxianus / 非相同末端結合 / ワクチン / 診断薬 / B型肝炎ウイルス / 酵母 / 耐熱性酵母 / 表面抗原タンパク質 / Ｂ型肝炎ウイルス

Outline of Final Research Achievements

The recombinant virus proteins are used for vaccinations and diagnostic reagent kits and their low cost productions are required. In this study, efficient virus protein production system was developed in yeast. When hepatitis B virus surface protein HBsL was expressed in yeast, it was degraded rapidly. We constructed highly stable HBsL mutants and also an expression system that enable stable virus protein production in yeast. These results provide strategies for the problems of virus infection that will emerge in the future.

Research Products

• [Journal Article] Screening of accurate clones for gene synthesis in yeast (2015)
  • Author(s): Tohru Yarimizu, Mikiko Nakamura, Hisashi Hoshida, and Rinji Akada
  • Journal Title: Journal of Bioscience and Bioengineering Volume: 119 Issue: 3 Pages: 251-259
  • DOI: 10.1016/j.jbiosc.2014.08.006
  • NAID: 110009923563
  • Peer Reviewed / Open Access / Acknowledgement Compliant
• [Journal Article] Synthetic signal sequences that enable efficient secretory protein production in the yeast Kluyveromyces marxianus. (2015)
  • Author(s): Tohru Yarimizu, Mikiko Nakamura, Hisashi Hoshida and Rinji Akada
  • Journal Title: Microbial Cell Factories Volume: 14 Issue: 1 Pages: 20-20
  • DOI: 10.1186/s12934-015-0203-y
  • Peer Reviewed / Open Access / Acknowledgement Compliant
• [Journal Article] Non-homologous end joining-mediated functional marker selection for DNA cloning in the yeast Kluyveromyces marxianus. (2014)
  • Author(s): Hoshida H, Murakami N, Suzuki A, Tamura R, Asakawa J, Abdel-Banat BM, Nonklang S, Nakamura M, Akada R.
  • Journal Title: Yeast Volume: 31 Issue: 1 Pages: 29-46
  • DOI: 10.1002/yea.2993
  • Peer Reviewed
• [Journal Article] Designed construction of recombinant DNA at the ura3Δ0 locus in the yeast Saccharomyces cerevisiae. (2013)
  • Author(s): Fukunaga T, Cha-Aim K, Hirakawa Y, Sakai R, Kitagawa T, Nakamura M, Nonklang S, Hoshida H, Akada R.
  • Journal Title: Yeast Volume: 30 Issue: 6 Pages: 243-253
  • DOI: 10.1002/yea.2957
  • Peer Reviewed
• [Journal Article] Identification of auxotrophic mutants of the yeast Kluyveromyces marxianus by non-homologous end joining-mediated integrative transformation with genes from Saccharomyces cerevisiae. (2013)
  • Author(s): Yarimizu T, Nonklang S, Nakamura J, Tokuda S, Nakagawa T, Lorreungsil S, Sutthikhumpha S, Pukahuta C, Kitagawa T, Nakamura M, Cha-Aim K, Limtong S, Hoshida H, Akada R.
  • Journal Title: Yeast Volume: 30 Issue: 12 Pages: 485-500
  • DOI: 10.1002/yea.2985
  • Peer Reviewed
• [Presentation] 酵母によるウイルスワクチン生産とウイルス粒子形成メカニズムの解析 (2014)
  • Author(s): 山本祐司，鈴木絢子，中村美紀子，星田尚司，赤田倫治
  • Organizer: 第３２回Yeast Workshop
  • Place of Presentation: ビューポートくれ、広島
  • Year and Date: 2014-11-14 – 2014-11-15
• [Presentation] 酵母Kluyveromyces marxianusの非相同末端結合の機構解析とそれを用いたプラスミドの構築 (2013)
  • Author(s): 星田尚司，飯泉広葉，土屋元靖，村上允唯，鎗水透，鈴木絢子，中村美紀子，赤田倫治
  • Organizer: 日本生物工学会大会
  • Place of Presentation: 広島国際会議場
• [Presentation] NHEJ-mediated recombinant DNA construction and its application for gene cloning and genetic analysis in the yeast Kluyveromyces marxianus (2013)
  • Author(s): Hisashi Hoshida, Nobutada Murakami, Ryoko Tamura, Ayako Suzuki, Rinji Akada
  • Organizer: 26th International Conference on Yeast Genetics and Molecular Biology
  • Place of Presentation: Frankfurt/Main, Germany
• [Presentation] B 型肝炎ワクチン開発に向けた酵母におけるウイルス表面タンパク質の発現解析 (2012)
  • Author(s): 冨吉啓介，中村美紀子，星田尚司，赤田倫治
  • Organizer: 日本農芸化学会中四国支部大会
  • Place of Presentation: 宇部工学部キャンパス
• [Book] Minimum GC-rich sequences for overlap extension PCR and primer annealing. (2014)
  • Author(s): Nakamura M, Suzuki A, Hoshida H, Akada R.
  • Total Pages: 17
  • Publisher: Humana Press