title: Efficient L-Alanine Production by a Thermo-Regulated Switch in Escherichia coli.
abstract: L-Alanine has important applications in food, pharmaceutical and veterinary and is used as a substrate for production of engineered thermoplastics. Microbial fermentation could reduce the production cost and promote the application of L-alanine. However, the presence of L-alanine significantly inhibit cell growth rate and cause a decrease in the ultimate L-alanine productivity. For efficient L-alanine production, a thermo-regulated genetic switch was designed to dynamically control the expression of L-alanine dehydrogenase (alaD) from Geobacillus stearothermophilus on the Escherichia coli B0016-060BC chromosome. The optimal cultivation conditions for the genetically switched alanine production using B0016-060BC were the following: an aerobic growth phase at 33 °C with a 1-h thermo-induction at 42 °C followed by an oxygen-limited phase at 42 °C. In a bioreactor experiment using the scaled-up conditions optimized in a shake flask, B0016-060BC accumulated 50.3 g biomass/100 g glucose during the aerobic growth phase and 96 g alanine/100 g glucose during the oxygen-limited phase, respectively. The L-alanine titer reached 120.8 g/l with higher overall and oxygen-limited volumetric productivities of 3.09 and 4.18 g/l h, respectively, using glucose as the sole carbon source. Efficient cell growth and L-alanine production were reached separately, by switching cultivation temperature. The results revealed the application of a thermo-regulated strategy for heterologous metabolic production and pointed to strategies for improving L-alanine production.
authors.affiliation: The Key Laboratory of Industrial Biotechnology of Ministry of Education, Jiangnan University, 1800 Lihu Avenue, Wuxi, 214122, People's Republic of China.|Center for Bioresource & Bioenergy, School of Biotechnology, Jiangnan University, 1800 Lihu Avenue, Wuxi, 214122, People's Republic of China.
authors.forename: Li
authors.initials: L
authors.lastname: Zhou
authors.affiliation: Center for Bioresource & Bioenergy, School of Biotechnology, Jiangnan University, 1800 Lihu Avenue, Wuxi, 214122, People's Republic of China.
authors.forename: Can
authors.initials: C
authors.lastname: Deng
authors.affiliation: The Key Laboratory of Industrial Biotechnology of Ministry of Education, Jiangnan University, 1800 Lihu Avenue, Wuxi, 214122, People's Republic of China.|Center for Bioresource & Bioenergy, School of Biotechnology, Jiangnan University, 1800 Lihu Avenue, Wuxi, 214122, People's Republic of China.
authors.forename: Wen-Jing
authors.initials: WJ
authors.lastname: Cui
authors.affiliation: The Key Laboratory of Industrial Biotechnology of Ministry of Education, Jiangnan University, 1800 Lihu Avenue, Wuxi, 214122, People's Republic of China.|Center for Bioresource & Bioenergy, School of Biotechnology, Jiangnan University, 1800 Lihu Avenue, Wuxi, 214122, People's Republic of China.
authors.forename: Zhong-Mei
authors.initials: ZM
authors.lastname: Liu
authors.affiliation: The Key Laboratory of Industrial Biotechnology of Ministry of Education, Jiangnan University, 1800 Lihu Avenue, Wuxi, 214122, People's Republic of China. zhmzhou@jiangnan.edu.cn.|Center for Bioresource & Bioenergy, School of Biotechnology, Jiangnan University, 1800 Lihu Avenue, Wuxi, 214122, People's Republic of China. zhmzhou@jiangnan.edu.cn.
authors.forename: Zhe-Min
authors.initials: ZM
authors.lastname: Zhou
mesh_terms: D000332:Aerobiosis; D000409:Alanine / Q000096:biosynthesis*; D050788:Alanine Dehydrogenase / Q000235:genetics / Q000378:metabolism; D019149:Bioreactors; D004926:Escherichia coli / Q000235:genetics*; D005285:Fermentation; D001411:Geobacillus stearothermophilus / Q000201:enzymology; D013696:Temperature
chemical_list: D050788:Alanine Dehydrogenase; D000409:Alanine
keywords: Chromosomal integration; Escherichia coli; L-alanine production; Metabolic engineering