William G. Lowrie Department of Chemical and Biomolecular Engineering, The Ohio State University, 140 West 19th Ave., Columbus, OH 43210, USA.
Appl Microbiol Biotechnol. 2012 Jan;93(2):881-9. doi: 10.1007/s00253-011-3736-y. Epub 2011 Dec 4.
Clostridium tyrobutyricum ATCC 25755 can produce butyric acid, acetic acid, and hydrogen as the main products from various carbon sources. In this study, C. tyrobutyricum was used as a host to produce n-butanol by expressing adhE2 gene under the control of a native thiolase promoter using four different conjugative plasmids (pMTL82151, 83151, 84151, and 85151) each with a different replicon (pBP1 from C. botulinum NCTC2916, pCB102 from C. butyricum, pCD6 from Clostridium difficile, and pIM13 from Bacillus subtilis). The effects of different replicons on transformation efficiency, plasmid stability, adhE2 expression and aldehyde/alcohol dehydrogenase activities, and butanol production by different mutants of C. tyrobutyricum were investigated. Among the four plasmids and replicons studied, pMTL82151 with pBP1 gave the highest transformation efficiency, plasmid stability, gene expression, and butanol biosynthesis. Butanol production from various substrates, including glucose, xylose, mannose, and mannitol were then investigated with the best mutant strain harboring adhE2 in pMTL82151. A high butanol titer of 20.5 g/L with 0.33 g/g yield and 0.32 g/L h productivity was obtained with mannitol as the substrate in batch fermentation with pH controlled at ~6.0.
凝结芽孢杆菌 ATCC 25755 可以利用各种碳源作为主要产物产生丁酸、乙酸和氢气。在这项研究中，凝结芽孢杆菌被用作宿主，通过在天然硫解酶启动子的控制下表达 adhE2 基因，使用四种不同的可接合质粒（pMTL82151、83151、84151 和 85151）来生产正丁醇，每种质粒都有不同的复制子（来自 C. botulinum NCTC2916 的 pBP1、来自 C. butyricum 的 pCB102、来自 Clostridium difficile 的 pCD6 和来自 Bacillus subtilis 的 pIM13）。研究了不同复制子对转化效率、质粒稳定性、adhE2 表达和醛/醇脱氢酶活性的影响，以及不同突变体凝结芽孢杆菌的丁醇生产。在所研究的四种质粒和复制子中，带有 pBP1 的 pMTL82151 具有最高的转化效率、质粒稳定性、基因表达和丁醇生物合成。然后，用带有 pMTL82151 中的 adhE2 的最佳突变菌株研究了葡萄糖、木糖、甘露糖和甘露醇等各种底物的丁醇生产。在 pH 控制在~6.0 的分批发酵中，以甘露醇为底物时，获得了 20.5 g/L 的高丁醇浓度，产率为 0.33 g/g，生产强度为 0.32 g/L·h。
