title: High-level extracellular production of D-Psicose-3-epimerase with recombinant Escherichia coli by a two-stage glycerol feeding approach.
abstract: The aim of this study is to achieve high-level extracellular production of D-Psicose-3-epimerase (DPE) with recombinant Escherichia coli. High-level production of DPE is one of the key factors in D-Psicose production. In the present study, the gene AAL45544.1 from Agrobacterium tumefaciens str. C58 was modified by artificial synthesis for overexpression in E. coli. The total DPE activity reached 3.96 U mL(-1) after optimization of the media composition, induction temperature, and concentration of inducer. Furthermore, it was found that addition of glycine had a positive effect on the extracellular production of DPE, which reached 3.5 U mL(-1). Finally, a two-stage glycerol feeding strategy based on both the specific growth rate before induction and the amount of glycerol residues after induction was applied in a 3-L fermenter. After a series of optimal strategies in the 3-L fermenter, the total and extracellular DPE activity were 5.08- and 3.11-fold higher than that noted in the shake flask. The extracellular and intracellular DPE activity reached 10.9 and 13.2 U mL(-1), achieving 25.5 and 31.1 % conversion of D-fructose to D-psicose, respectively. The systemic strategies presented in this study provide valuable novel information for the industrial application of DPE.
authors.affiliation: Key Laboratory of Industrial Biotechnology, Ministry of Education, Jiangnan University, Wuxi, 214122, China, gouki_gl@hotmail.com.
authors.forename: Lei
authors.initials: L
authors.lastname: Gu
authors.forename: Juan
authors.initials: J
authors.lastname: Zhang
authors.forename: Baihong
authors.initials: B
authors.lastname: Liu
authors.forename: Chongde
authors.initials: C
authors.lastname: Wu
authors.forename: Guocheng
authors.initials: G
authors.lastname: Du
authors.forename: Jian
authors.initials: J
authors.lastname: Chen
mesh_terms: D001483:Base Sequence; D019149:Bioreactors; D003001:Cloning, Molecular; D003470:Culture Media; D017931:DNA Primers; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000235:genetics*; D005285:Fermentation; D005990:Glycerol / Q000378:metabolism*; D019998:Racemases and Epimerases / Q000096:biosynthesis* / Q000235:genetics; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics; D011995:Recombination, Genetic
chemical_list: D003470:Culture Media; D017931:DNA Primers; D011994:Recombinant Proteins; D019998:Racemases and Epimerases; D005990:Glycerol