| Summary: | 碩士 === 國立臺灣大學 === 分子與細胞生物學研究所 === 99 === 1-butanol has been considered as a promising biofuel because of many advantages over ethanol, such as high energy content, low hygroscopicity, low vapor pressure, and allowing for a complete replacement of gasoline without modifications to the existing vehicle engines. Metabolic engineering of user-friendly host, Escherichia coli, for 1-butanol production has been accomplished. However, the pivotal problem associate with the industrial production of 1-butanol is that this product is toxic towards the host, resulting in a low 1-butanol yield. To comprehend the complex basis for its toxicity and obtain a global conception of 1-butanol stress response, we applied array-based, quantitative screening procedures for monitoring growth of bacterial in different 1-butanol concentration at genome-wide scale. After screening and analysis, we identified a group of 1-butanol sensitive strains and a group of tolerant strains. In 1% 1-butanol condition, growth rate, cell viability, cell number and integrity of cell membrane of these 1-butanol tolerant strains, acrA and acrB mutants, were significantly higher than wild type. The cluster of orthologous genes (COG) can provide the framework for functional genome analysis. Therefore, we used COG to categorize these 1-butanol sensitive strains, and found that COG C (energy conversion and production) was the major group. The sensitive and tolerant candidates and network analysis were integrated to identify the 1-butanol stress response networks and their enriched biological functions. The 1-butanol stress response network uncovered that oxidative phosphorylation might play important roles in 1-butanol stress response. Furthermore, mutants had increased sensitivity to 1-butanol because they had no these transcription factors, including Crp and FruR which activated complexes in oxidative phosphorylation. Overexpression of Crp and FruR in certain mutants could increase their 1-butanol tolerance. Taken together, these results reveal a key relationship between oxidative phosphorylation and 1-butanol stress response. Our studies also provide a valuable in-depth insight into development of a 1-butanol tolerant strain, even high-titer, 1-butanol-producing host.
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