title: Limitation of thiamine pyrophosphate supply to growing Escherichia coli switches metabolism to efficient D-lactate formation.
abstract: Efficient production of D-lactate by engineered Escherichia coli entails balancing cell growth and product synthesis. To develop a metabolic switch to implement a desirable transition from cell growth to product fermentation, a thiamine auxotroph B0013-080A was constructed in a highly efficient D-lactate producer E. coli strain B0013-070. This was achieved by inactivation of thiE, a gene encoding a thiamine phosphate synthase for biosynthesis of thiamine monophosphate. The resultant mutant B0013-080A failed to grow on the medium in the absence of thiamine yet growth was restored when exogenous thiamine was provided. A linear relationship between cell mass formation and amount of thiamine supplemented was mathematically determined in a shake flask experiment and confirmed in a 7-L bioreactor system. This calculation revealed that ∼ 95-96 thiamine molecules per cell were required to satisfy cell growth. This relationship was employed to develop a novel fermentation process for D-lactate production by using thiamine as a limiting condition. A D-lactate productivity of 4.11 g · L(-1) · h(-1) from glycerol under microaerobic condition and 3.66 g · L(-1) · h(-1) from glucose under anaerobic condition was achieved which is 19.1% and 10.2% higher respectively than the parental strain. These results revealed a convenient and reliable method to control cell growth and improve D-lactate fermentation. This control strategy could be applied to other biotechnological processes that require optimal allocation of carbon between cell growth and product formation.
authors.affiliation: Key Laboratory of Industrial Fermentation Microbiology, Ministry of Education, The College of Biotechnology, Tianjin University of Science and Technology, Tianjin 300457, P. R. China.
authors.forename: Kangming
authors.initials: K
authors.lastname: Tian
authors.affiliation: College of Biological Science and Engineering, Fuzhou University, Fuzhou 350108, P. R. China. ddniu0529@fzu.edu.cn.
authors.forename: Dandan
authors.initials: D
authors.lastname: Niu
authors.affiliation: Key Laboratory of Industrial Fermentation Microbiology, Ministry of Education, The College of Biotechnology, Tianjin University of Science and Technology, Tianjin 300457, P. R. China.
authors.forename: Xiaoguang
authors.initials: X
authors.lastname: Liu
authors.affiliation: Department of Microbiology, Stellenbosch University, Matieland, South Africa.
authors.forename: Bernard A
authors.initials: BA
authors.lastname: Prior
authors.affiliation: Center for Bioresource and Bioenergy, School of Biotechnology, Jiangnan University, Wuxi, P. R. China.
authors.forename: Li
authors.initials: L
authors.lastname: Zhou
authors.affiliation: Key Laboratory of Industrial Fermentation Microbiology, Ministry of Education, The College of Biotechnology, Tianjin University of Science and Technology, Tianjin 300457, P. R. China.
authors.forename: Fuping
authors.initials: F
authors.lastname: Lu
authors.affiliation: Department of Biotechnology and Food Technology, Faculty of Applied Sciences, Durban University of Technology, Durban, South Africa.
authors.forename: Suren
authors.initials: S
authors.lastname: Singh
authors.affiliation: Key Laboratory of Industrial Fermentation Microbiology, Ministry of Education, The College of Biotechnology, Tianjin University of Science and Technology, Tianjin 300457, P. R. China. zx.wang@tust.edu.cn.
authors.forename: Zhengxiang
authors.initials: Z
authors.lastname: Wang
mesh_terms: D003470:Culture Media / Q000737:chemistry; D004926:Escherichia coli / Q000235:genetics / Q000254:growth & development / Q000378:metabolism*; D019344:Lactic Acid / Q000378:metabolism*; D060847:Metabolic Engineering / Q000379:methods; D013835:Thiamine Pyrophosphate / Q000378:metabolism*
chemical_list: D003470:Culture Media; D019344:Lactic Acid; D013835:Thiamine Pyrophosphate
keywords: cell replication; d-lactate fermentation; thiamine auxotroph