一株在有氧条件下转化丙酮醇为1，2-丙二醇的大肠杆菌菌株的研究

生物技术通报 ›› 2016, Vol. 32 ›› Issue (12): 160-165.

一株在有氧条件下转化丙酮醇为1，2-丙二醇的大肠杆菌菌株的研究

邓静静,胡洪波,张雪洪

上海交通大学生命科学技术学院微生物代谢国家重点实验室，上海 200240

收稿日期:2016-03-01 出版日期:2016-12-25 发布日期:2016-12-07
作者简介:邓静静，女，硕士研究生，研究方向：大肠杆菌利用甘油生产高价值产物的研究；E-mail：977613938@qq.com
基金资助:
“973”项目（2012CB721005），国家自然科学基金项目（J1210047）

Research on an Escherichia coli Strain Capable of Transferring Acetol to 1，2-Propanediol Aerobically

DENG Jing-jing,HU Hong-bo,ZHANG Xue-hong

Key Laboratory of Microbial Metabolism，School of Life Sciences and Biotechnology，Shanghai Jiao Tong University，Shanghai 200240

Received:2016-03-01 Published:2016-12-25 Online:2016-12-07

摘要/Abstract

摘要： 本实验室在前期构建了一株能高效利用甘油合成丙酮醇的大肠杆菌工程菌株，如果能在有氧条件下将丙酮醇转化为1，2-丙二醇，则可以大幅提高1，2-丙二醇的合成速率。以大肠杆菌BW25113ΔglpK为出发菌株，筛选得到一株可以在有氧条件下代谢甘油的突变菌株PDO1。结果表明，该菌株可以在有氧条件下将丙酮醇转化为1，2-丙二醇，1，2-丙二醇的产量达到0.73 g/L，转化率达到0.493 g/g丙酮醇。突变株PDO1的甘油脱氢酶酶活和胞内的NADH百分比分别是对照株的75.9倍和1.64倍，由此推测，甘油脱氢酶酶活的提高以及NADH的相对提高，使得突变株可以在有氧条件下通过甘油脱氢酶途径代谢甘油，并转化丙酮醇为1，2-丙酮醇。

关键词: 大肠杆菌, 1, 2-丙二醇, 甘油, 丙酮醇, 静息细胞发酵

Abstract: We constructed an engineered Escherichia coli strain that can efficiently convert glycerol to acetol in our lab，the productivity of 1，2- Propanediol（1，2-PDO）may increase significantly if the strain can convert acetol to 1，2-PDO under aerobic condition. A mutant strain named PDO1 was screened from original strain E. coli BW25113ΔglpK，which metabolized glycerol aerobically. The further experiments showed that the strain PDO1 converted acetol to 1，2-PDO with the yield of 0.73 g/L and conversion rate of 0.493 g/g acetol under aerobic condition. The activity of glycerol dehydrogenase and the proportion of NADH were 75.6 and 1.64 times higher than that of control respectively. It is inferred that the increase of glycerol dehydrogenase activity and NADH level resulted in the mutant strain PDO1 metabolizing glycerol by glycerol dehydrogenase under aerobic condition and the conversion of acetol to 1，2-PDO .

Key words: Escherichia coli, 1, 2-Propanediol（1, 2-PDO）, glycerol, acetol, resting cells fermentation

中图分类号:

10.13560/j.cnki.biotech.bull.1985.2016.12.025

邓静静,胡洪波,张雪洪. 一株在有氧条件下转化丙酮醇为1，2-丙二醇的大肠杆菌菌株的研究[J]. 生物技术通报, 2016, 32(12): 160-165.

DENG Jing-jing,HU Hong-bo,ZHANG Xue-hong. Research on an Escherichia coli Strain Capable of Transferring Acetol to 1，2-Propanediol Aerobically[J]. Biotechnology Bulletin, 2016, 32(12): 160-165.

参考文献

[1] Clomburg JM, Gonzalez R. Anaerobic fermentation of glycerol：a platform for renewable fuels and chemicals[J] . Trends in Biotechnology, 2013, 31（1）：21-28.
[2] Zhu HL, Yi XY, Liu Y, et al. Production of acetol from glycerol using engineered Escherichia coli[J] . Bioresource Technology, 2013, 149（4）：238-243.
[3] Yang FX, Hanna MA, Sun RC. Value-added uses for crude glycerol-a byproduct of biodiesel production[J] . Biotechnology for Biofuels, 2012, 5（6）：1-10.
[4] 郑军. 1, 2 丙二醇国内外生产现状及发展前景[J] . 热固性树脂, 2009, 24（1）：58-62.
[5] Bennett GN, San KY. Microbial formation, bitechnological production and application of 1, 2-Propanediol[J] . Applied Microbiology & Biotechnology, 2001, 55（1）：1-9.
[6] Ramon G, Abhishek M, Yandi D, et al. A new model for the anaerobic fermentation of glycerol in enteric bacteria：Trunk and auxiliary pathways in Escherichia coli[J] . Metabolic Engineering, 2008, 10（5）：234-245.
[7] Liu Q, Xiong DW, Hu HB, et al. Hydrogen production from glycerol using a genetically engineered Escherichia coli HW2 strain[J] . Journal of Chemical Engineering of Chinese Universities, 2015, 29（5）：1133-1137.
[8] 曲璟秋, 刘翠花. 应用基因敲除快速构建大肠杆菌突变体改造脂肪酸代谢途径[J] . 微生物学报, 2013, 53（6）：608-614.
[9] Madyagol M, Al AH, Levarski Z, et al. Gene replacement techniques for Escherichia coli genome modification[J] . Folia Microbiologica, 2011, 56（3）：253-263.
[10] Tomoya B, Takeshi A, Miki H, et al. Construction of Escherichia coli K-12 in-frame, single gene knockout mutants：the keio collection[J] . Molecular Systems Biology, 2006, 2（1）：8-11.
[11] Jin RZ, Tang J, Lin E, et al. Experimental evolution of a novel pathway for glycerol dissimilation in Escherichia coli[J] . Journal of Molecular Evolution, 1983, 19（6）：429-436.
[12] Piattoni CV, Figueroa CM, Diez MDA, et al. Production and characterization of Escherichia coli glycerol dehydrogenase as a tool for glycerol recycling[J] . Process Biochemistry, 2013, 48（3）：406-412.
[13] Bekers KM, Heijnen JJ, Gulik WM. Determination of the in vivo NAD：NADH ratio in Saccharomyces cerevisiae under anaerobic conditions, using alcohol dehydrogenase as sensor reaction[J] . Yeast, 2015, 32（8）：541-557.
[14] Bakker BM, Overkamp KM, Van AJ, et al. Stoichiometry and compartmentation of NADH metabolism in Saccharomyces cerevisiae[J] . FEMS Microbiol, 2001, 25（1）：15-37.
[15] Rachit J, Jin H, Yuan QP, et al. Engineering microaerobic metabolism of E. coli for 1, 2-propanediol production[J] . Journal of Industrial Microbiology & Biotechnology, 2015, 42（3）：1-7.
[16] Zhang YP, Huang ZH, Du C, et al. Introduction of an NADH regeneration system into Klebsiella oxytoca leads to an enhanced oxidative and reductive metabolism of glycerol[J] . Metabolic Engineering, 2008, 11（2）：101-106.
[17] Sun FF, Dai CY, Xie J, et al. Biochemical issues in estimation of cytosolic free NAD/NADH Ratio[J] . PLoS One, 2012, 7（5）：1-10.