title: Improvement of the intracellular environment for enhancing l-arginine production of Corynebacterium glutamicum by inactivation of H2O2-forming flavin reductases and optimization of ATP supply.
abstract: l-arginine, a semi essential amino acid, is an important amino acid in food flavoring and pharmaceutical industries. Its production by microbial fermentation is gaining more and more attention. In previous work, we obtained a new l-arginine producing Corynebacterium crenatum (subspecies of Corynebacterium glutamicum) through mutation breeding. In this work, we enhanced l-arginine production through improvement of the intracellular environment. First, two NAD(P)H-dependent H2O2-forming flavin reductases Frd181 (encoded by frd1 gene) and Frd188 (encoded by frd2) in C. glutamicum were identified for the first time. Next, the roles of Frd181 and Frd188 in C. glutamicum were studied by overexpression and deletion of the encoding genes, and the results showed that the inactivation of Frd181 and Frd188 was beneficial for cell growth and l-arginine production, owing to the decreased H2O2synthesis and intracellular reactive oxygen species (ROS) level, and increased intracellular NADH and ATP levels. Then, the ATP level was further increased by deletion of noxA (encoding NADH oxidase) and amn (encoding AMP nucleosidase), and overexpression of pgk (encoding 3-phosphoglycerate kinase) and pyk (encoding pyruvate kinase), and the l-arginine production and yield from glucose were significantly increased. In fed-batch fermentation, the l-arginine production and yield from glucose of the final strain reached 57.3g/L and 0.326g/g, respectively, which were 49.2% and 34.2% higher than those of the parent strain, respectively. ROS and ATP are important elements of the intracellular environment, and l-arginine biosynthesis requires a large amount of ATP. For the first time, we enhanced l-arginine production and yield from glucose through reducing the H2O2synthesis and increasing the ATP supply.
authors.affiliation: The Key Laboratory of Industrial Biotechnology of Ministry of Education, School of Biotechnology, Jiangnan University, Wuxi, Jiangsu 214122, PR China.
authors.forename: Zaiwei
authors.initials: Z
authors.lastname: Man
authors.affiliation: The Key Laboratory of Industrial Biotechnology of Ministry of Education, School of Biotechnology, Jiangnan University, Wuxi, Jiangsu 214122, PR China; State Key Laboratory of Food Science and Technology, Jiangnan University, Wuxi, Jiangsu 214122, PR China. Electronic address: raozhm@jiangnan.edu.cn.
authors.forename: Zhiming
authors.initials: Z
authors.lastname: Rao
authors.affiliation: The Key Laboratory of Industrial Biotechnology of Ministry of Education, School of Biotechnology, Jiangnan University, Wuxi, Jiangsu 214122, PR China.
authors.forename: Meijuan
authors.initials: M
authors.lastname: Xu
authors.affiliation: The Key Laboratory of Industrial Biotechnology of Ministry of Education, School of Biotechnology, Jiangnan University, Wuxi, Jiangsu 214122, PR China.
authors.forename: Jing
authors.initials: J
authors.lastname: Guo
authors.affiliation: The Key Laboratory of Industrial Biotechnology of Ministry of Education, School of Biotechnology, Jiangnan University, Wuxi, Jiangsu 214122, PR China.
authors.forename: Taowei
authors.initials: T
authors.lastname: Yang
authors.affiliation: The Key Laboratory of Industrial Biotechnology of Ministry of Education, School of Biotechnology, Jiangnan University, Wuxi, Jiangsu 214122, PR China.
authors.forename: Xian
authors.initials: X
authors.lastname: Zhang
authors.affiliation: School of Medicine and Pharmaceuticals, Jiangnan University, Wuxi, Jiangsu 214122, PR China. Electronic address: zhenghxu@jiangnan.edu.cn.
authors.forename: Zhenghong
authors.initials: Z
authors.lastname: Xu
mesh_terms: D000255:Adenosine Triphosphate / Q000378:metabolism*; D001120:Arginine / Q000096:biosynthesis* / Q000235:genetics; D053898:Biosynthetic Pathways / Q000235:genetics; D048230:Corynebacterium glutamicum / Q000502:physiology*; D038181:FMN Reductase / Q000235:genetics* / Q000378:metabolism; D024861:Genetic Enhancement / Q000379:methods; D006861:Hydrogen Peroxide / Q000378:metabolism*; D008658:Inactivation, Metabolic / Q000235:genetics; D007424:Intracellular Fluid / Q000378:metabolism; D060847:Metabolic Engineering / Q000379:methods*; D053858:Metabolic Networks and Pathways / Q000235:genetics*
chemical_list: D000255:Adenosine Triphosphate; D001120:Arginine; D006861:Hydrogen Peroxide; D038181:FMN Reductase
keywords: ATP supply; Corynebacterium glutamicum; Flavin reductase; H(2)O(2); Intracellular environment; L-arginine