Search for enzymes that introduce a triple bond between two carbon atoms and elucidation of their reaction mechanisms

• Project/Area Number: 16K14898
• Research Category: Grant-in-Aid for Challenging Exploratory Research
• Allocation Type: Multi-year Fund
• Research Field: Applied biochemistry
• Research Institution: University of Tsukuba
• Principal Investigator: TANAKA Toshiyuki 筑波大学, 生命環境系, 教授 (10217052)
• Project Period (FY): 2016-04-01 – 2020-03-31
• Project Status: Completed (Fiscal Year 2019)
• Budget Amount: ¥3,640,000 (Direct Cost: ¥2,800,000、Indirect Cost: ¥840,000); Fiscal Year 2018: ¥1,170,000 (Direct Cost: ¥900,000、Indirect Cost: ¥270,000); Fiscal Year 2017: ¥1,170,000 (Direct Cost: ¥900,000、Indirect Cost: ¥270,000); Fiscal Year 2016: ¥1,300,000 (Direct Cost: ¥1,000,000、Indirect Cost: ¥300,000)
• Keywords: アセチレナーゼ様酵素 / クローニング / タンパク質発現 / 酵素 / 酵素化学

Outline of Final Research Achievements

We tried to clone the genes of proteins that is likely to be involved in forming a carbon-carbon triple bond from mainly plants and microorganisms, and to establish their expression systems and purification methods. As a result, 18 kinds of proteins were successfully cloned, and for 9 of them we were able to obtain highly pure proteins. Crystallization was performed using these purified proteins, but crystals suitable for structural analysis have not yet been obtained. To analyze their three-dimensional structures and elucidate their reaction mechanisms, we will continue experiments such as crystallization.

Academic Significance and Societal Importance of the Research Achievements

炭素間三重結合を有する天然有機化合物は反応性が高く、強力な抗菌活性を示すものも多い。しかし、この炭素間三重結合を形成する酵素（アセチレナーゼ）は数種類しか同定されておらず、反応機構についても全くわかっていない。今回の研究で、18種類のアセチレナーゼと推測されるあるいは関連するタンパク質のクローニングに成功した。継続して研究を続け、反応機構が原子レベルで解明出来れば、酵素の工業的利用、酵素の改変や微生物利用により、新しい抗菌剤などの生産が可能になると考えられる。