title: Overexpression of ppc and lysC to improve the production of 4-hydroxyisoleucine and its precursor l-isoleucine in recombinant Corynebacterium glutamicum ssp. lactofermentum.
abstract: 4-hydroxyisoleucine (4-HIL) exhibits unique insulinotropic and insulin-sensitizing activities and is an attractive candidate for the treatment of type II and type I diabetes. In our previous study, l-isoleucine dioxygenase gene (ido) was cloned and overexpressed in an l-isoleucine-producing strain, Corynebacterium glutamicum ssp. lactofermentum SN01, and 4-HIL was produced from the endogenous l-isoleucine (Ile). In this study, ppc and lysC were co-expressed with ido to increase the supply of Ile, the direct precursor of 4-HIL, and to further improve the 4-HIL yield. After 144h of fermentation, the ido-ppc-expressing strain produced 95.72±1.52mM 4-HIL, 29% higher than the ido-expressing strain. The co-expression of lysC and ppc with ido resulted in a further 35% increment of carbon flux to l-aspartate family amino acids biosynthesis pathway. However, the conversion ratio of Ile to 4-HIL and the 4-HIL yield decreased to 0.31mol/mol and 30.16±2.01mM, respectively, likely due to the decreased IDO activity caused by lower pH and higher intracellular Ile concentration. Therefore, co-expression of ido and ppc was benefit for 4-HIL de novo biosynthesis, while co-expression of lysC with ido and ppc decreased the conversion ratio of Ile to 4-HIL.
authors.affiliation: State Key Laboratory of Food Science and Technology, Jiangnan University, Wuxi 214122, China; Key Laboratory of Industrial Biotechnology, Ministry of Education, School of Biotechnology, Jiangnan University, Wuxi 214122, China; Synergetic Innovation Center of Food Safety and Nutrition, Jiangnan University, Wuxi 214122, China. Electronic address: shifeng@jiangnan.edu.cn.
authors.forename: Feng
authors.initials: F
authors.lastname: Shi
authors.affiliation: State Key Laboratory of Food Science and Technology, Jiangnan University, Wuxi 214122, China; Key Laboratory of Industrial Biotechnology, Ministry of Education, School of Biotechnology, Jiangnan University, Wuxi 214122, China.
authors.forename: Huimin
authors.initials: H
authors.lastname: Fang
authors.affiliation: State Key Laboratory of Food Science and Technology, Jiangnan University, Wuxi 214122, China; Key Laboratory of Industrial Biotechnology, Ministry of Education, School of Biotechnology, Jiangnan University, Wuxi 214122, China.
authors.forename: Tengfei
authors.initials: T
authors.lastname: Niu
authors.affiliation: State Key Laboratory of Food Science and Technology, Jiangnan University, Wuxi 214122, China; Key Laboratory of Industrial Biotechnology, Ministry of Education, School of Biotechnology, Jiangnan University, Wuxi 214122, China.
authors.forename: Zhengke
authors.initials: Z
authors.lastname: Lu
mesh_terms: D001222:Aspartate Kinase / Q000235:genetics / Q000378:metabolism; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism; D053898:Biosynthetic Pathways; D057486:Carbon Cycle; D048230:Corynebacterium glutamicum / Q000235:genetics* / Q000378:metabolism*; D049308:Dioxygenases / Q000235:genetics / Q000378:metabolism; D005285:Fermentation; D005798:Genes, Bacterial; D007532:Isoleucine / Q000031:analogs & derivatives* / Q000096:biosynthesis*; D010730:Phosphoenolpyruvate Carboxylase / Q000235:genetics / Q000378:metabolism; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D015854:Up-Regulation
chemical_list: C104672:4-hydroxyisoleucine; D001426:Bacterial Proteins; D011994:Recombinant Proteins; D007532:Isoleucine; D049308:Dioxygenases; D001222:Aspartate Kinase; D010730:Phosphoenolpyruvate Carboxylase
keywords: 4-Hydroxyisoleucine; Corynebacterium glutamicum ssp. lactofermentum; l-isoleucine dioxygenase; lysC; ppc