title: High-level production of Arthrobacter aurescens CYC705 nitrilase in Escherichia coli for biosynthesis of iminodiacetic acid.
abstract: Nitrilase from Arthrobacter aurescens CYC705 can hydrolyze the iminodiacetonitrile to iminodiacetic acid (IDA) efficiently, and its high-level production in Escherichia coli has not been established. In the present work, the production of this nitrilase expressed in E. coli BL21(DE3) with a recombinant plasmid pET28a-cyc705 was optimized. Various culture conditions and process parameters including medium components and concentrations, inducer types and concentrations, inducing temperature and time were systematically examined in a shake flask. After optimization, the OD600 , nitrilase activity, and productivity were obviously improved and achieved to 40.91 ± 1.341, 98.12 ± 1.248 U/mL, and 2,230 ± 28.36 U L(-1)  H(-1) , respectively, about 2.1-, 30-, and 33-fold increases as compared with those in the primary medium. Furthermore, four different fermentation strategies were adopted to scale up cultivation of the recombinant E. coli BL21(DE3)/pET28a-cyc705 in a 3.7-L fermenter. Substituting the peanut powder with fish peptone and accompanying with 1.0% glycerol feeding could significantly reduce the bubble production and shorten the fermentation time, which resulted in a nitrilase productivity of 4,653 ± 38.16 U L(-1) H(-1) that was about two times higher than that in a shake flask. The high-level production of A. aurescens CYC705 nitrilase established in this study will meet the need of industrial biosynthesis of IDA.
authors.affiliation: Enzyme and Fermentation Technology Laboratory, College of Light Industry Science and Engineering, Nanjing Forestry University, Nanjing, People's Republic of China.
authors.forename: Erzheng
authors.initials: E
authors.lastname: Su
authors.affiliation: Enzyme and Fermentation Technology Laboratory, College of Light Industry Science and Engineering, Nanjing Forestry University, Nanjing, People's Republic of China.
authors.forename: Chao
authors.initials: C
authors.lastname: Lu
authors.affiliation: State Key Laboratory of Bioreactor Engineering, New World Institute of Biotechnology, East China University of Science and Technology, Shanghai, People's Republic of China.
authors.forename: Xiaoqiang
authors.initials: X
authors.lastname: Ma
authors.affiliation: State Key Laboratory of Bioreactor Engineering, New World Institute of Biotechnology, East China University of Science and Technology, Shanghai, People's Republic of China.
authors.forename: Wenwen
authors.initials: W
authors.lastname: Cai
authors.affiliation: State Key Laboratory of Bioreactor Engineering, New World Institute of Biotechnology, East China University of Science and Technology, Shanghai, People's Republic of China.
authors.forename: Shujing
authors.initials: S
authors.lastname: Zhu
mesh_terms: D000619:Aminohydrolases / Q000096:biosynthesis* / Q000378:metabolism; D001173:Arthrobacter / Q000201:enzymology*; D019149:Bioreactors / Q000382:microbiology; D001709:Biotechnology / Q000379:methods*; D002244:Carbon / Q000494:pharmacology; D003470:Culture Media / Q000737:chemistry; D004305:Dose-Response Relationship, Drug; D004926:Escherichia coli / Q000187:drug effects / Q000254:growth & development / Q000378:metabolism*; D005285:Fermentation; D007098:Imino Acids / Q000378:metabolism*; D009584:Nitrogen / Q000494:pharmacology; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000378:metabolism; D012492:Salts / Q000494:pharmacology; D013696:Temperature
chemical_list: D003470:Culture Media; D007098:Imino Acids; D011994:Recombinant Proteins; D012492:Salts; D002244:Carbon; D000619:Aminohydrolases; C021867:nitrilase; D009584:Nitrogen; C008109:iminodiacetic acid
keywords: arthrobacter aurescens CYC705; culturing medium; iminodiacetic acid; inducing conditions; nitrilase; optimization