title: Chaperones-assisted soluble expression and maturation of recombinant Co-type nitrile hydratase in Escherichia coli to avoid the need for a low induction temperature.
abstract: Nitrile hydratase (NHase) is an important industrial enzyme that biosynthesizes high-value amides. However, most of NHases expressed in Escherichia coli easily aggregate to inactive inclusion bodies unless the induction temperature is reduced to approximately 20°C. The NHase from Aurantimonas manganoxydans has been functionally expressed in E. coli, and exhibits considerable potential for the production of nicotinamide in industrial application. In this study, the effects of chaperones including GroEL/ES, Dnak/J-GrpE and trigger factor on the expression of the recombinant Co-type NHase were investigated. The results indicate that three chaperones can significantly promote the active expression of the recombinant NHase at 30°C. The total NHase activities reached to 263 and 155U/ml in shake flasks when the NHase was co-expressed with GroEL/ES and DnaK/J-GrpE, which were 52- and 31-fold higher than the observed activities without chaperones, respectively. This increase is possibly due to the soluble expression of the recombinant NHase assisted by molecular chaperones. Furthermore, GroEL/ES and DnaK/J-GrpE were determined to promote the maturation of the Co-type NHase in E. coli under the absence of the parental activator gene. These knowledge regarding the chaperones effect on the NHase expression are useful for understanding the biosynthesis of Co-type NHase.
authors: College of Material, Chemistry and Chemical Engineering, Hangzhou Normal University, Hangzhou, 310012, PR China.
authors: Xiaolin
authors: X
authors: Pei
authors: College of Material, Chemistry and Chemical Engineering, Hangzhou Normal University, Hangzhou, 310012, PR China.
authors: Qiuyan
authors: Q
authors: Wang
authors: Institute of Bioengineering, College of Chemical and Biological Engineering, Zhejiang University, Hangzhou, 310028, PR China.
authors: Lijun
authors: L
authors: Meng
authors: Institute of Bioengineering, College of Chemical and Biological Engineering, Zhejiang University, Hangzhou, 310028, PR China.
authors: Jing
authors: J
authors: Li
authors: Institute of Bioengineering, College of Chemical and Biological Engineering, Zhejiang University, Hangzhou, 310028, PR China.
authors: Zhengfen
authors: Z
authors: Yang
authors: College of Material, Chemistry and Chemical Engineering, Hangzhou Normal University, Hangzhou, 310012, PR China.
authors: Xiaopu
authors: X
authors: Yin
authors: Institute of Bioengineering, College of Chemical and Biological Engineering, Zhejiang University, Hangzhou, 310028, PR China.
authors: Lirong
authors: L
authors: Yang
authors: College of Life Science, Zhejiang Chinese Medical University, Hangzhou, 310053, PR China. Electronic address: csy003@163.com.
authors: Shaoyun
authors: S
authors: Chen
authors: Institute of Bioengineering, College of Chemical and Biological Engineering, Zhejiang University, Hangzhou, 310028, PR China. Electronic address: wjp@zju.edu.cn.
authors: Jianping
authors: J
authors: Wu
mesh_terms: D020561:Alphaproteobacteria / Q000201:enzymology / Q000235:genetics; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D006836:Hydro-Lyases / Q000235:genetics / Q000378:metabolism; D018832:Molecular Chaperones; D009536:Niacinamide / Q000378:metabolism; D010957:Plasmids; D013696:Temperature
chemical_list: D001426:Bacterial Proteins; D018832:Molecular Chaperones; D009536:Niacinamide; D006836:Hydro-Lyases; C048831:nitrile hydratase
keywords: Active expression; Induction temperature; Molecular chaperones; Multimeric metalloenzyme; Nitrile hydratase