CLS

Laboratory for Chemistry and Life Science, Institute of Innovative Research, Tokyo Institute of Technology

東京工業大学
科学技術創成研究院
化学生命科学研究所

LAST UPDATE 2020/06/08

  • 研究者氏名
    Researcher Name

    上田宏　Hiroshi UEDA
    教授　Professor

  • 所属
    Professional Affiliation

    東京工業大学科学技術創成研究院化学生命科学研究所
    Laboratory for Chemistry and Life Science, Institute of Innovative Research, Tokyo Institute of Technology

    分子生命化学領域
    Division of Molecular Bioscience

  • 研究キーワード
    Research Keywords

    蛋白質工学
    免疫測定
    酵素工学

    protein engineering
    immunoassay
    enzyme engineering
研究テーマ
Research Subject
タンパク質工学的手法に基づく新規バイオプロセスの創成
Development of novel bioprocesses based on protein engineering methodologies

研究の背景 Background

我々生命の主体であるタンパク質の中には，依然人工的に模倣しがたい高い性能（分子認識能や触媒能など）を持つものが数多くあります。しかし天然タンパク質は人類社会の今後の持続的発展への応用の観点から見て，必ずしも最適な性質を持つとは限りません。我々はここにタンパク質工学の醍醐味があると考え，さまざまなタンパク質の，合目的デザインと分子進化的技術，さらにケミカルバイオロジー的技術をも用いた創製を試みています。目的にかなった方法を開発し，それを駆使することで，天然ではできない芸当ができるタンパク質が創製できるものと期待しています。

In nature, many proteins exhibit high performances such as molecular recognition ability and catalytic activity, which are still difficult to emulate artificially. However, natural protein does not always have ideal property from the viewpoint of application to the continuous development of human society. That is why we aim at creating novel proteins with superior performances and the assays using them, through the rational design and molecular evolution, as well as chemical biological techniques.

研究の目標 Outcome

我々が現在特に力を入れているのは，第一に組み換え抗体を用いた新規検出素子・測定法の開発であり、低分子を高感度かつ容易に測定可能な免疫診断法（オープンサンドイッチ法､OS法）と，その発展型として「抗原結合により光る」蛍光標識抗体Quenchbodyの開発と応用に力を入れています。第２に，例えばホタル発光酵素変異体を用いた新規タンパク質間相互作用検出系の開発など，酵素のエンジニアリングによる新規プロセスの構築を目標としています。

Our primary target is the development and application of novel detection probes and assays based on recombinant antibodies, such as ‘Open sandwich immunoassay’ which can detect small molecules easily and sensitively, as well as ‘Quenchbodies’ which can detect various antigens by the increase of fluorescence. Secondly, we aims at constructing various novel processes based on enzyme engineering, such as development of novel protein-protein interaction assay FlimPIA, based on firefly luciferase mutants.

研究図Research Figure

Fig.1. Principle of Open-sandwich Immunoassay (Upper left). The two antigen-binding fragments VH and VL associate stronger in the presence of antigen, whose association is detected by the immobilized enzyme activityFig. 2. Working model of Quenchbody (Lower left). When the antibody VH and/or VL fragments are N-terminally labeled with fluorescent dye, it is quenched by the Trp residues in the antibody. Upon adding antigen, the V region is stabilized and the dye no longer interacts with Trp, resulting in increased fluorescence. Fig. 3. Principle of FlimPIA. Fluc mutant whose oxidative luminescent reaction is slow (donor) and another mutant whose adenylation reaction is slow (acceptor) are used. When the donor and acceptor come close at the time of interaction, luminescence activity is enhanced by the relay of reaction intermediate LH2-AMP.

文献 / Publications

ACS Sens. 5, 24 (2020). Biotechnol. Bioeng. 117, 1259 (2020). Sci. Rep. 9, 18189 (2019). Sci. Rep. 9, 10188 (2019). Biochemistry 58, 2695 (2019). Anal. Chem. 90, 3001 (2018). Chem. Commun. 53, 10200 (2017). WIREs Nanomed. Nanobiotechnol., e1457 (2017).

研究者HP