Fujiwara, Kei

写真a

Affiliation

Faculty of Science and Technology, Department of Biosciences and Informatics （Yagami）

Position

Associate Professor

Related Websites

External Links
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Profile 【 Display / hide 】

  • Disrupted cells won't go back into its living states. We are trying to clarify why this process is irreversible and developing a method to reconstruct living cells from biomolecules mixtures using artificial cells engineering.

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Career 【 Display / hide 】

  • 2009.04
    -
    2010.03

    The university of Tokyo, Graduate School of Frontier Sciences, Department of Medical Genome Sciences, JSPS research associate

  • 2010.04
    -
    2011.03

    Kyoto university, Institute for Integrated Cell-Material Sciences, Research Associate

  • 2011.04
    -
    2014.03

    Tohoku University, Department of Bioengineering and Robotics, JSPS research associate

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Academic Background 【 Display / hide 】

  • 2004.04
    -
    2006.03

    The University of Tokyo, Graduate School of Agricultural and Life Sciences, Department of Applied Biological Chemistry

    Graduate School, Completed, Master's course

  • 2006.04
    -
    2009.03

    The University of Tokyo, Graduate School of Frontier Sciences, Department of Medical Genome Sciences

    Graduate School, Completed, Doctoral course

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Academic Degrees 【 Display / hide 】

  • 博士(生命科学), The University of Tokyo, Coursework, 2009.03

    シャペロニンGroEL/ESの細胞内における役割の解明

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Research Areas 【 Display / hide 】

  • Life Science / Molecular biology

  • Life Science / Biophysics

  • Life Science / System genome science

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Research Keywords 【 Display / hide 】

  • Artificial-Cell engineering

  • Synthetic Biology

  • Bacteriology

  • Cell-free Life Sciences

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Books 【 Display / hide 】

  • Synthetic Biology: A Very Short Introduction

    藤原 慶,徳永 美恵, ニュートンプレス, 2021.04

    Original author： Jamie A. Davies,

  • ペプチド医薬品のスクリーニング・安定化・製剤化技術

    SUDO Kei, FUJIWARA Kei, DOI Nobuhide, 技術情報協会, 2017.12

    Scope: 1-4. mRNAディスプレイ法によるペプチドのスクリーニング

  • 人工細胞の創製とその応用

    FUJIWARA Kei, CMC出版, 2017.01

    Scope: 「3-4. 無細胞システムによる生命システムの理解」の執筆

  • DNA分子デザインのすべて ～BIOMOD虎の巻

    FUJIWARA Kei, CBT学会eBOOK, 2016.04

    Scope: p. 15-20， p. 33-34の執筆を担当

    　View Summary

    分子ロボティクス研究会編

  • 「自然世界の高分子」

    TANAKA Motohiko, TOKITA masayuki, YANAGISAWA Miho, SAKAUE Takahiro, FUJIWARA Kei, 吉岡書店, 2016.03

    Scope: 9章から11章の翻訳を担当

    　View Summary

    「Giant molecules」Alexander Y. Grosberg and Alexei R. Khokhlovの邦訳版(翻訳)

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Papers 【 Display / hide 】

  • Cytoplasmic delivery of siRNA using human-derived membrane penetration-enhancing peptide

    Nakamura M., Fujiwara K., Doi N.

    Journal of Nanobiotechnology （Journal of Nanobiotechnology)  20 （ 1 ）  2022.12

    　View Summary

    Background: Although protein-based methods using cell-penetrating peptides such as TAT have been expected to provide an alternative approach to siRNA delivery, the low efficiency of endosomal escape of siRNA/protein complexes taken up into cells by endocytosis remains a problem. Here, to overcome this problem, we adopted the membrane penetration-enhancing peptide S19 from human syncytin 1 previously identified in our laboratory. Results: We prepared fusion proteins in which the S19 and TAT peptides were fused to the viral RNA-binding domains (RBDs) as carrier proteins, added the RBD-S19-TAT/siRNA complex to human cultured cells, and investigated the cytoplasmic delivery of the complex and the knockdown efficiency of target genes. We found that the intracellular uptake of the RBD-S19-TAT/siRNA complex was increased compared to that of the RBD-TAT/siRNA complex, and the expression level of the target mRNA was decreased. Because siRNA must dissociate from RBD and bind to Argonaute 2 (Ago2) to form the RNA-induced silencing complex (RISC) after the protein/siRNA complex is delivered into the cytoplasm, a dilemma arises: stronger binding between RBD and siRNA increases intracellular uptake but makes RISC formation more difficult. Thus, we next prepared fusion proteins in which the S19 and TAT peptides were fused with Ago2 instead of RBD and found that the efficiencies of siRNA delivery and knockdown obtained using TAT-S19-Ago2 were higher than those using TAT-Ago2. In addition, we found that the smallest RISC delivery induced faster knockdown than traditional siRNA lipofection, probably due to the decreased time required for RISC formation in the cytoplasm. Conclusion: These results indicated that S19 and TAT-fused siRNA-binding proteins, especially Ago2, should be useful for the rapid and efficient delivery of siRNA without the addition of any endosome-disrupting agent.

  • Controlling the Periodicity of a Reaction-Diffusion Wave in Artificial Cells by a Two-Way Energy Supplier

    Takada S., Yoshinaga N., Doi N., Fujiwara K.

    ACS Nano （ACS Nano)  16 （ 10 ） 16853 - 16861 2022.10

    ISSN  19360851

    　View Summary

    Reaction-diffusion (RD) waves, which are dynamic self-organization structures generated by nanosize molecules, are a fundamental mechanism from patterning in nano- and micromaterials to spatiotemporal regulations in living cells, such as cell division and motility. Although the periods of RD waves are the critical element for these functions, the development of a system to control their period is challenging because RD waves result from nonlinear physical dynamics under far-from-equilibrium conditions. Here, we developed an artificial cell system with tunable period of an RD-driven wave (Min protein wave), which determines a cell division site plane in living bacterial cells. The developed system is based on our finding that Min waves are generated by energy consumption of either ATP or dATP, and the period of the wave is different between these two energy suppliers. We showed that the Min-wave period was modulated linearly by the mixing ratio of ATP and dATP and that it was also possible to estimate the mixing ratio of ATP and dATP from the period. Our findings illuminated a previously unidentified principle to control the dissipative dynamics of biomolecules and, simultaneously, built an important framework to construct molecular robots with spatiotemporal units.

  • Cell-Size Space Regulates the Behavior of Confined Polymers: From Nano- and Micromaterials Science to Biology

    Yanagisawa M., Watanabe C., Yoshinaga N., Fujiwara K.

    Langmuir （Langmuir)  38 （ 39 ） 11811 - 11827 2022.10

    ISSN  07437463

    　View Summary

    Polymer micromaterials in a liquid or gel phase covered with a surfactant membrane are widely used materials in pharmaceuticals, cosmetics, and foods. In particular, cell-sized micromaterials of biopolymer solutions covered with a lipid membrane have been studied as artificial cells to understand cells from a physicochemical perspective. The characteristics and phase transitions of polymers confined to a microscopic space often differ from those in bulk systems. The effect that causes this difference is referred to as the cell-size space effect (CSE), but the specific physicochemical factors remain unclear. This study introduces the analysis of CSE on molecular diffusion, nanostructure transition, and phase separation and presents their main factors, i.e., short- and long-range interactions with the membrane surface and small volume (finite element nature). This serves as a guide for determining the dominant factors of CSE. Furthermore, we also introduce other factors of CSE such as spatial closure and the relationships among space size, the characteristic length of periodicity, the structure size, and many others produced by biomolecular assemblies through the analysis of protein reaction-diffusion systems and biochemical reactions.

  • Mode selection mechanism in traveling and standing waves revealed by Min wave reconstituted in artificial cells

    Sakura Takada, Natsuhiko Yoshinaga, Nobuhide Doi, Kei Fujiwara

    Science Advances （Science Advances)  8 （ 23 ） eabm8460 2022.06

    Research paper (scientific journal), Joint Work, Last author, Corresponding author, Accepted

    　View Summary

    Reaction-diffusion coupling (RDc) generates spatiotemporal patterns, including two dynamic wave modes: traveling and standing waves. Although mode selection plays a substantial role in the spatiotemporal organization of living cell molecules, the mechanism for selecting each wave mode remains elusive. Here, we investigated a wave mode selection mechanism using Min waves reconstituted in artificial cells, emerged by the RDc of MinD and MinE. Our experiments and theoretical analysis revealed that the balance of membrane binding and dissociation from the membrane of MinD determines the mode selection of the Min wave. We successfully demonstrated that the transition of the wave modes can be regulated by controlling this balance and found hysteresis characteristics in the wave mode transition. These findings highlight a previously unidentified role of the balance between activators and inhibitors as a determinant of the mode selection of waves by RDc and depict an unexplored mechanism in intracellular spatiotemporal pattern formations.

  • Characterization of the membrane penetration-enhancing peptide S19 derived from human syncytin-1 for the intracellular delivery of TAT-fused proteins

    Suzuki M., Iwaki K., Kikuchi M., Fujiwara K., Doi N.

    Biochemical and Biophysical Research Communications （Biochemical and Biophysical Research Communications)  586   63 - 67 2022.01

    ISSN  0006291X

    　View Summary

    Although cell-penetrating peptides such as the HIV-derived TAT peptide have been used as tools for the intracellular delivery of therapeutic peptides and proteins, a problem persists: the endosomal escape efficiency is low. Previously, we found that the fusogenic peptide S19, derived from the human protein syncytin-1, enhance the endosomal escape efficiency of proteins that incorporated by endocytosis via TAT. In this study, we first performed Ala-scanning mutagenesis of S19, and found that all Ile, Val, Leu and Phe with high β-sheet forming propensities in S19 are important for the intracellular uptake of S19-TAT-fused proteins. In a secondary structure analysis of the mutated S19-TAT peptides in the presence of liposomes mimicking late endosomes (LEs), the CD spectra of V3A and I4A mutants with low uptake activity showed the appearance of an α-helix structure, whereas the mutant G5A retained both the uptake activity and the β-structure. In addition, we investigated the appropriate linking position and order of the S19 and TAT peptides to a cargo protein including an apoptosis-induced peptide and found that both the previous C-terminal S19-TAT tag and the N-terminal TAT-S19 tag promote the cytoplasmic delivery of the fusion protein. These results and previous results suggest that the interaction of TAT with the LE membrane causes a structural change in S19 from a random coil to a β-strand and that the subsequent parallel β-sheet formation between two S19 peptides may promote adjacent TAT dimerization, resulting in endosomal escape from the LE membrane.

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Papers, etc., Registered in KOARA 【 Display / hide 】

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Reviews, Commentaries, etc. 【 Display / hide 】

  • Min波の人工細胞内再構成とそこから見えた細胞サイズ空間効果

    光山 隼史, 義永 那津人, 藤原 慶

    生物物理学会誌 (生物物理学会)   62 ( 1 ) 19 - 23 2022.03

    Article, review, commentary, editorial, etc. (scientific journal), Last author, Corresponding author

  • 酵素を内包した人工細胞が拓く新しい醸造業や医薬品の開発

    藤原慶

    MDB 技術予測レポート    項目36 2019

  • Artificial cell fermentation as a next platform for biosynthesis

    FUJIWARA Kei

    バイオサイエンスとインダストリー ((一財)バイオインダストリー協会)  76 ( 4 ) 302 - 303 2018.07

    Article, review, commentary, editorial, etc. (trade magazine, newspaper, online media), Single Work

  • DNA ナノテクノロジー コラム ”BIOMODへ参加しよう！”

    FUJIWARA Kei, Tadakuma Hisashi

    現代化学 (東京化学同人)   ( 11 ) 36 - 37 2016.10

    Article, review, commentary, editorial, etc. (trade magazine, newspaper, online media), Joint Work

  • Microdroplets as a Model System for the Study of Macromolecular Crowding in Cells

    YANAGISAWA Miho, FUJIWARA Kei

    生物物理 (日本生物物理学会)  55 ( 5 ) 246 - 249 2015.09

    Article, review, commentary, editorial, etc. (scientific journal), Joint Work

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Presentations 【 Display / hide 】

  • 細胞内反応拡散波の人工細胞再構成系で探る細胞空間の時空間パターン形成原理

    藤原慶

    定量生物学の会第十回年会, 

    2022.12

  • 人工細胞内再構成系で探る生化学システムの細胞サイズ効果

    藤原慶

    広島大学健康長寿研究拠点（HiHA）WS「サイズ生物学の探究」, 

    2022.10

  • 細胞を創る研究 (ボトムアップ合成生物学) への誘い

    藤原慶

    TARA Seminar理論合成インシリコ生物学セミナー, 

    2022.07

  • 細胞を創る研究から考える微生物の生

    藤原慶

    第44回日本分子生物学会, 

    2021.12

  • 人工細胞創成から見えたきた新しい生化学とその将来像

    藤原慶

    隅基礎科学創成財団 微生物CS グループ1 第5回定例会, 

    2021.05

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Research Projects of Competitive Funds, etc. 【 Display / hide 】

  • 創るトランスクリプトームにより迫る生命の設計原理

    2023.04
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    2027.03

    創発的研究支援事業, Principal investigator

  • 細胞内にチューリングパターンは形成可能か？

    2022.06
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    2024.03

    MEXT,JSPS, Grant-in-Aid for Scientific Research, 挑戦的研究（萌芽）, Principal investigator

  • Establishment of artificial cell fermentation technology as a practical material for industrial usage

    2022.06
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    2024.03

    MEXT,JSPS, Grant-in-Aid for Scientific Research, 学術変革領域研究(A), Principal investigator

  • Conversion between mechanical power and information of intracellular reaction-diffusion wave

    2022.04
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    2024.03

    MEXT,JSPS, Grant-in-Aid for Scientific Research, 新学術領域研究（研究領域提案型）, Principal investigator

  • Elucidation of physics underlying spatiotemporal pattern formation in cell size spaces

    2020.04
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    2023.03

    MEXT,JSPS, Grant-in-Aid for Scientific Research, Grant-in-Aid for Scientific Research (B), Principal investigator

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Courses Taught 【 Display / hide 】

  • TOPICS IN BIOSCIENCES AND INFORMATICS 2

    2024

  • SEMINAR IN BIOSCIENCES AND INFORMATICS

    2024

  • OFF-CAMPUS STUDY PROGRAM 3

    2024

  • OFF-CAMPUS STUDY PROGRAM 2

    2024

  • OFF-CAMPUS STUDY PROGRAM 1

    2024

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Courses Previously Taught 【 Display / hide 】

  • 微生物学

    慶應義塾大学

    2018.04
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    2019.03

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