Metabolic engineering of Enterobacter cloacae for high-yield production of enantiopure (2R,3R)-2,3-butanediol from lignocellulose-derived sugars
Metabolic engineering of Enterobacter cloacae for high-yield production of enantiopure (2R,3R)-2,3-butanediol from lignocellulose-derived sugars
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阴沟肠杆菌的代谢工程，用于从木质纤维素衍生的糖中高产生产对映体纯 (2R,3R)-2,3-丁二醇
DOI：
10.1016/j.ymben.2014.11.010
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发表时间：
2015-03-01
影响因子：
8.4
通讯作者：
Xu, Ping
中科院分区：
文献类型：
--
作者：
Li, Lixiang;Li, Kun;Xu, Ping
生物燃料的生物技术生产受到产品如乙醇和丁醇的毒性的限制。2，3-丁二醇（2，3-BD）作为一种生物燃料和平台化学品，对微生物的毒性较低，有望成为利用可再生生物资源生产生物燃料的一种替代品。此外，没有报道细菌菌株使用木质纤维素水解产物生产对映体纯的2，3-BD。本研究对阴沟肠杆菌SDM菌株进行了系统的代谢工程改造，构建了一种高效的生物催化剂，用于生产燃料和对映体纯的生物化学品（2 R，3R）-2，3-BD。首先，将各种（2 R，3R）-2，3-BD脱氢酶编码基因在内消旋-2，3-BD脱氢酶编码基因被破坏的E.在大肠杆菌2，3-BD生物合成基因簇的天然启动子P-b下，clothing.然后，通过葡萄糖转运蛋白编码基因ptsG的失活和半乳糖通透酶编码基因galP的过表达来消除碳分解代谢物阻遏。该菌株能同时利用葡萄糖和木糖。为了提高（2 R，3R）-2，3-BD的生产效率，敲除了副产物产生基因（ldh和frdA），从而使（2 R，3R）-2，3-BD的产量在500-mL锥形瓶中提高了16.5%。在5L的生物反应器中采用分批补料发酵，以木质纤维素水解液中的两种主要碳水化合物葡萄糖和木糖为原料，在44 h内得到152.0g/L（2 R，3R）-2，3-BD（纯度> 97.5%），比生产率为3.5g/[L·h]，产率为97.7%。以木质纤维素水解液为底物，在51 h内可得到119.4 g/L（2 R，3R）-2，3-BD（纯度> 96.0%），产率为2.3 g/[L·h]，收率为95.0%。这些结果表明，通过天然或工程菌株从生物质衍生的糖生产对映体纯（2 R，3R）-2，3-BD获得了最高记录。除了生产2，3-BD之外，我们的系统方法还可以用于通过使用木质纤维素衍生的糖来生产其他重要的化学品。(C)2014年国际代谢工程学会。爱思唯尔公司出版All rights reserved.
Biotechnological production of biofuels is restricted by toxicity of the products such as ethanol and butanol. As its low toxicity to microbes, 2,3-butanediol (2,3-BD), a fuel and platform bio-chemical, could be a promising alternative for biofuel production from renewable bioresources. In addition, no bacterial strains have been reported to produce enantiopure 2,3-BD using lignocellulosic hydrolysates. In this study, Enterobacter cloacae strain SDM was systematically and metabolically engineered to construct an efficient biocatalyst for production of the fuel and enantiopure bio-chemical (2R,3R)-2,3-BD. First, the various (2R,3R)-2,3-BD dehydrogenase encoding genes were expressed in a meso-2,3-BD dehydrogenase encoding gene disrupted E. cloacae strain under native promoter P-b of the 2,3-BD biosynthetic gene cluster of E. cloacae. Then, carbon catabolite repression was eliminated via inactivation of the glucose transporter encoding gene ptsG and overexpression of a galactose permease encoding gene galP. The resultant strain could utilize glucose and xylose simultaneously. To improve the efficiency of (2R,3R)-2,3-BD production, the byproduct-producing genes (ldh and frdA) were knocked out, thereby enhancing the yield of (2R,3R)-2,3-BD by 16.5% in 500-mL Erlenmeyer flasks. By using fed-batch fermentation in a 5-L bioreactor, 152.0 g/L (2R,3R)-2,3-BD (purity > 97.5%) was produced within 44 h with a specific productivity of 3.5 g/[L h] and a yield of 97.7% from a mixture of glucose and xylose, two major carbohydrate components in lignocellulosic hydrolysates. In addition, when a lignocellulosic hydrolysate was used as the substrate, 119.4 g/L (2R,3R)-2,3-BD (purity > 96.0%) was produced within 51 h with a productivity of 2.3 g/[L h] and a yield of 95.0%. These results show that the highest records have been acquired for enantiopure (2R,3R)-2,3-BD production by a native or engineered strain from biomass-derived sugars. In addition to producing the 2,3-BD, our systematic approach might also be used in the production of other important chemicals by using lignocellulose-derived sugars. (C) 2014 International Metabolic Engineering Society. Published by Elsevier Inc. All rights reserved.