title: Addition of Co(2+) to culture medium decides the functional expression of a recombinant nitrile hydratase in Escherichia coli.
abstract: A nitrile hydratase (NHase) gene from Aurantimonas manganoxydans, cloned and expressed in Escherichia coli, gave an enzyme that efficiently hydrated 3-cyanopyridine to nicotinamide with high thermal stability. We have now found that adding Co(2+) at 0.1 mM to LB medium was essential for production of an active enzyme. However, ≥0.3 mM Co(2+) inhibited the growth of host cells in LB medium and decreased the production of the recombinant NHase. Furthermore, β-mercaptoethanol promoted regeneration of the Co(2+)-defective apoenzyme in vitro possibly by breaking a key disulfide bond thereby promoting the incorporation of Co(2+) into the apoenzyme.
authors.affiliation: Center for Biomedicine and Health, College of Life and Environmental Sciences, Hangzhou Normal University, Hangzhou, 311121, China. pxl@hznu.edu.cn
authors.forename: Xiaolin
authors.initials: X
authors.lastname: Pei
authors.forename: Qiuyan
authors.initials: Q
authors.lastname: Wang
authors.forename: Chenglu
authors.initials: C
authors.lastname: Li
authors.forename: Xiaopu
authors.initials: X
authors.lastname: Yin
authors.forename: Rong
authors.initials: R
authors.lastname: Chen
authors.forename: Tian
authors.initials: T
authors.lastname: Xie
mesh_terms: D020561:Alphaproteobacteria / Q000201:enzymology / Q000235:genetics; D002413:Cations, Divalent / Q000378:metabolism*; D003001:Cloning, Molecular; D003035:Cobalt / Q000378:metabolism*; D003470:Culture Media / Q000737:chemistry*; D004926:Escherichia coli / Q000187:drug effects* / Q000201:enzymology* / Q000254:growth & development; D015870:Gene Expression / Q000187:drug effects; D006836:Hydro-Lyases / Q000096:biosynthesis*; D009536:Niacinamide / Q000378:metabolism; D011725:Pyridines / Q000378:metabolism; D011994:Recombinant Proteins / Q000096:biosynthesis
chemical_list: D002413:Cations, Divalent; D003470:Culture Media; D011725:Pyridines; D011994:Recombinant Proteins; D009536:Niacinamide; D003035:Cobalt; D006836:Hydro-Lyases; C048831:nitrile hydratase; C020729:3-cyanopyridine