Kataoka Naoya, Vangnai Alisa S, Pongtharangkul Thunyarat, Yakushi Toshiharu, Matsushita Kazunobu
Division of Agricultural Sciences, Graduate School of Sciences and Technology for Innovation, Yamaguchi University, Yamaguchi 753-8515, Japan; Research Center for Thermotolerant Microbial Resources, Yamaguchi University, Yamaguchi 753-8515, Japan.
Department of Biochemistry, Faculty of Science, Chulalongkorn University, Bangkok 10330, Thailand; Center of Excellence in Hazardous Substance Management (HSM), Chulalongkorn University, Bangkok 10330, Thailand.
J Biosci Bioeng. 2017 May;123(5):562-568. doi: 10.1016/j.jbiosc.2016.12.008. Epub 2017 Jan 11.
Butyrate is an important industrial platform chemical. Although several groups have reported butyrate production under oxygen-limited conditions by a native producer, Clostridium tyrobutylicum, and by a metabolically engineered Escherichia coli, efforts to produce butyrate under aerobic growth conditions have met limited success. Here, we constructed a novel butyrate synthetic pathway that functions under aerobic growth conditions in E. coli, by modifying the 1-butanol synthetic pathway reported previously. The pathway consists of phaA (acetyltransferase) and phaB (NADPH-dependent acetoacetyl-CoA reductase) from Ralstonia eutropha, phaJ ((R)-specific enoyl-CoA hydratase) from Aeromonas caviae, ter (trans-enoyl-CoA reductase) from Treponema denticola, and endogenous thioesterase(s) of E. coli. To evaluate the potential of this pathway for butyrate production, culture conditions, including pH, oxygen supply, and concentration of inorganic nitrogen sources, were optimized in a mini-jar fermentor. Under the optimal conditions, butyrate was produced at a concentration of up to 140 mM (12.3 g/L in terms of butyric acid) after 54 h of fed-batch culture.
丁酸盐是一种重要的工业平台化学品。尽管有几个研究小组报道了在氧气受限条件下，天然生产菌酪丁酸梭状芽孢杆菌以及经过代谢工程改造的大肠杆菌能够生产丁酸盐，但在有氧生长条件下生产丁酸盐的努力取得的成功有限。在此，我们通过修改先前报道的1-丁醇合成途径，构建了一条在大肠杆菌有氧生长条件下起作用的新型丁酸盐合成途径。该途径由来自真养产碱菌的phaA（乙酰转移酶）和phaB（NADPH依赖性乙酰乙酰辅酶A还原酶）、来自豚鼠气单胞菌的phaJ（(R)-特异性烯酰辅酶A水合酶）、来自龋齿密螺旋体的ter（反式烯酰辅酶A还原酶）以及大肠杆菌的内源性硫酯酶组成。为了评估该途径生产丁酸盐的潜力，在小型罐式发酵罐中对培养条件进行了优化，包括pH值、氧气供应和无机氮源浓度。在最佳条件下，经过54小时的补料分批培养后，丁酸盐的产量高达140 mM（以丁酸计为12.3 g/L）。
