title: Screening and Improving the Recombinant Nitrilases and Application in Biotransformation of Iminodiacetonitrile to Iminodiacetic Acid.
abstract: In this study, several nitrilase genes from phylogenetically distinct organisms were expressed and purified in E. coli in order to study their ability to mediate the biotransformation of nitriles. We identified three nitrilases: Acidovorax facilis nitrilase (AcN); Alcaligenes fecalis nitrilase (AkN); and Rhodococcus rhodochrous nitrilase (RkN), which catalyzed iminodiacetonitrile (IDAN) to iminodiacetic acid (IDA). AcN demonstrated 8.8-fold higher activity for IDAN degradation as compared to AkN and RkN. Based on homology modeling and previously described 'hot spot' mutations, several AcN mutants were screened for improved activity. One mutant M3 (F168V/L201N/S192F) was identified, which demonstrates a 41% enhancement in the conversion as well as a 2.4-fold higher catalytic efficiency towards IDAN as compared to wild-type AcN.
authors.affiliation: Institute of Bioengineering, Zhejiang University of Technology, Hangzhou, People's Republic of China.
authors.forename: Zhi-Qiang
authors.initials: ZQ
authors.lastname: Liu
authors.forename: Peter James
authors.initials: PJ
authors.lastname: Baker
authors.forename: Feng
authors.initials: F
authors.lastname: Cheng
authors.forename: Ya-Ping
authors.initials: YP
authors.lastname: Xue
authors.forename: Yu-Guo
authors.initials: YG
authors.lastname: Zheng
authors.forename: Yin-Chu
authors.initials: YC
authors.lastname: Shen
mesh_terms: D000097:Acetonitriles / Q000378:metabolism*; D042481:Alcaligenes faecalis / Q000201:enzymology; D000619:Aminohydrolases / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D001711:Biotransformation; D002851:Chromatography, High Pressure Liquid; D002942:Circular Dichroism; D042621:Comamonadaceae / Q000201:enzymology; D004926:Escherichia coli; D006868:Hydrolysis; D007098:Imino Acids / Q000378:metabolism*; D007700:Kinetics; D062105:Molecular Docking Simulation; D009154:Mutation; D017433:Protein Structure, Secondary; D011994:Recombinant Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism; D012240:Rhodococcus / Q000201:enzymology; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid
chemical_list: D000097:Acetonitriles; D001426:Bacterial Proteins; D007098:Imino Acids; D011994:Recombinant Proteins; D000619:Aminohydrolases; C021867:nitrilase; C008109:iminodiacetic acid