Reversible utilization of enzyme by grafting and refolding

2020 Fiscal Year Final Research Report

• Project/Area Number: 18K04821
• Research Category: Grant-in-Aid for Scientific Research (C)
• Allocation Type: Multi-year Fund
• Section: 一般
• Review Section: Basic Section 27020:Chemical reaction and process system engineering-related
• Research Institution: Tokyo University of Agriculture and Technology
• Principal Investigator: Ohashi Hidenori 東京農工大学, 工学(系)研究科(研究院), 准教授 (00541179)
• Project Period (FY): 2018-04-01 – 2021-03-31
• Keywords: 固定化酵素 / タンパク質 / リフォールディング / 大気圧プラズマ / グラフト重合

Outline of Final Research Achievements

Conventional immobilized enzyme is hard to recover its function once denatured. On the other hand, in the case of free enzyme as protein, after unfolded to a linear chain of amino acid sequences with denaturant, it can recover its function by removing denaturant to refold the active 3D structure. In the conventional design, immobilized enzyme is hard to refold, because the structure was stiffly anchored to mitigate the inactivation of protein. In the present study, novel type of immobilized enzyme was designed, which can recover its function reversibly by unfolding/refolding by temporary and subsequent covalent immobilization with terminal groups. The specific functional groups for both fixation were introduced to a model protein and to a material surface respectively, and they were combined in order. Further, atmospheric-pressure plasma graft polymerization technique was thoroughly investigated to facilitate the functionalization of material surface.

Free Research Field

化学工学

Academic Significance and Societal Importance of the Research Achievements

酵素は高い基質特異性を持ち、副生成物の少ない高選択的な反応を可能とする優れた生体触媒であり、分離操作を必要としない化学プロセスへの応用が見込まれる。一方で、酵素は化成品と比べると高価な上、失活しやすいため、失活を防ぐ目的で酵素の固定化法が開発されてきた。しかし従来の固定化法では一旦失活すると機能回復が極めて困難である。本課題では、失活しても容易に機能回復を可能とする固定化酵素設計により、高価故に見送られてきた機能性酵素を再度実用化に近づけることを可能とする。また、グラフト鎖導入を簡便に行う手法を開発したことで、様々な材料の表面に、種々の高機能を付与することが可能になった。