Application of gene editing by Clostron technology for biorefinery by hydrogen producing Clostridium

2023 Fiscal Year Final Research Report

• Project/Area Number: 21K12307
• Research Category: Grant-in-Aid for Scientific Research (C)
• Allocation Type: Multi-year Fund
• Section: 一般
• Review Section: Basic Section 64030:Environmental materials and recycle technology-related
• Research Institution: Mie University
• Principal Investigator: Kimura Tetsuya 三重大学, 生物資源学研究科, 教授 (00281080)
• Project Period (FY): 2021-04-01 – 2024-03-31
• Keywords: Clostridium / biomass / hydrogen gas / pyruvate / acetyl-CoA / ethanol / chitin

Outline of Final Research Achievements

C. paraputrificum decomposes chitin and produces hydrogen gas and ethanol. Pathway from pyruvate to acetyl-CoA is a critical step for hydrogen gas production. Pyruvate ferredoxin oxidoreductase (PFO) catalyzes this important step. However, Pyruvate formate lyase (PFL), which converts pyruvate to formate and acetyl-CoA, bypasses the PFO pathway. Therefore, we attempted to stop PFL pathway and to concentrate metabolic flow from pyruvate to acetyl-CoA in PFO pathway. The PFL gene and the PFL activase (PFLA) gene were encoded in polycistronic and we tried to disrupt PFL and PFLA gene. Only the PFLA gene was disrupted by the genome editing ClosTron method. The production of hydrogen gas increased in glucose as a carbon source. In addition, we succeeded in disrupting the production pathway of butyric acid and the strain increased the ethanol production. Furthermore, we have succeeded in developing a high-expression vector for cellulolytic bacterium R. josui.

Free Research Field

応用微生物学

Academic Significance and Societal Importance of the Research Achievements

地球温暖化による気候変動は人類が直面する重大な課題であり世界的に注目が集まっている。必要なエネルギーの多くを輸入化石燃料に頼っている我が国にとって、バイオマスを有効利用することは重要である。本研究で対象とする嫌気性細菌は、キチンやセルロースを分解して水素ガスやアルコールを生産する。本研究では、キチンを分解する細菌の遺伝子編集技術を利用して、代謝経路を改変することで水素ガス生産やアルコール生産の向上を行った。本研究で扱う嫌気性細菌は、研究室で単離した固有の細菌であり、我が国独自の技術としてオリジナリティーが高い。