Department of Biochemistry, Faculty of Science, Chulalongkorn University, Bangkok 10330, Thailand.
AMB Express. 2011 May 30;1(1):10. doi: 10.1186/2191-0855-1-10.
As alternative microbial hosts for butanol production with organic-solvent tolerant trait are in high demands, a butanol-tolerant bacterium, Bacillus subtilis GRSW2-B1, was thus isolated. Its tolerance covered a range of organic solvents at high concentration (5%v/v), with remarkable tolerance in particular to butanol and alcohol groups. It was susceptible for butanol acclimatization, which resulted in significant tolerance improvement. It has versatility for application in a variety of fermentation process because it has superior tolerance when cells were exposed to butanol either as high-density, late-exponential grown cells (up to 5%v/v) or under growing conditions (up to 2.25%v/v). Genetic transformation procedure was optimized, yielding the highest efficiency at 5.17 × 103 colony forming unit (μg DNA)-1. Gene expression could be effectively driven by several promoters with different levels, where as the highest expression was observed with a xylose promoter. The constructed vector was stably maintained in the transformants, in the presence or absence of butanol stress. Adverse effect of efflux-mediated tetracycline resistance determinant (TetL) to bacterial organic-solvent tolerance property was unexpectedly observed and thus discussed. Overall results indicate that B. subtilis GRSW2-B1 has potential to be engineered and further established as a genetic host for bioproduction of butanol.
由于对具有有机溶剂耐受性的丁醇生产替代微生物宿主的需求很高，因此分离到了一株耐丁醇细菌枯草芽孢杆菌 GRSW2-B1。它对多种有机溶剂（5%v/v）具有耐受性，尤其对丁醇和醇类具有显著的耐受性。它对丁醇驯化具有敏感性，这导致其耐受性显著提高。由于在高细胞密度、迟滞期生长的细胞（高达 5%v/v）或在生长条件下（高达 2.25%v/v）暴露于丁醇时，细胞具有卓越的耐受性，因此它具有在各种发酵过程中应用的多功能性。遗传转化程序得到了优化，在 5.17×103 个集落形成单位（μg DNA）-1 时达到了最高效率。几种不同水平的启动子可以有效地驱动基因表达，其中木糖启动子的表达最高。构建的载体在转化体中能够稳定维持，无论是否存在丁醇胁迫。出人意料地观察到外排介导的四环素抗性决定簇（TetL）对细菌有机溶剂耐受性特性的不良影响，并进行了讨论。总体结果表明，枯草芽孢杆菌 GRSW2-B1 具有被工程改造并进一步确立为丁醇生物生产遗传宿主的潜力。
