Zhejiang Provincial Key Laboratory of Biometrology and Inspection and Quarantine, College of Life Sciences, China Jiliang University, Hangzhou, Zhejiang Province, China.
J Basic Microbiol. 2013 Apr;53(4):348-54. doi: 10.1002/jobm.201100580. Epub 2012 Jun 26.
It is generally known that genes dhaB and dhaT are responsible for 1,3-propanediol (1,3-PD) production in the presence of glycerol in Klebsiella pneumoniae and these genes are organized in one operon. In the present study, a genetic means of increasing the enzyme activities of 1,3-PD formation pathway through the over-expression of 1,3-PD opeorn was performed in K. pneumoniae S6. The recombinant strain S6-PD showed 27- and 15-fold increase in enzymatic activities of DhaB and DhaT, respectively with respect to wild-type strain while failed to improve the 1,3-PD yield due to the inadequacy of cofactor NADH. Therefore, in order to increase NADH availability, a NADH regeneration system was constructed by heterologous expression of NAD(+) -dependent formate dehydrogenase gene (fdh1) from Candida boidinii and introduced into S6-PD to investigate its effects on the glycerol utilization and 1,3-PD formation. The results demonstrated that the increase of NADH availability could efficiently improve glycerol metabolism and promote 1,3-PD yield.
众所周知，在肺炎克雷伯氏菌中，基因 dhaB 和 dhaT 负责在存在甘油的情况下生产 1,3-丙二醇（1,3-PD），这些基因组织在一个操纵子中。在本研究中，通过过表达 1,3-PD 操纵子，在肺炎克雷伯氏菌 S6 中采用遗传手段增加 1,3-PD 形成途径的酶活性。重组菌株 S6-PD 中 DhaB 和 DhaT 的酶活性分别比野生型菌株提高了 27 倍和 15 倍，而由于辅因子 NADH 的不足，1,3-PD 的产量并没有提高。因此，为了增加 NADH 的可用性，通过异源表达 Candida boidinii 中的 NAD(+) -依赖型甲酸脱氢酶基因（fdh1）构建了 NADH 再生系统，并将其引入 S6-PD 中，以研究其对甘油利用和 1,3-PD 形成的影响。结果表明，增加 NADH 的可用性可以有效地改善甘油代谢并促进 1,3-PD 的产量。
