Purification and characterization of the enantioselective nitrile hydratase from Rhodococcus sp. AJ270.
Nitrile hydratase (NHase, EC 4.2.1.84) from Rhodococcus sp. AJ270 was purified with 23.96% yield after sonication, ammonium sulfate fractionation, ion exchange, hydrophobic and gel-filtration column chromatography. The enzyme showed intriguing characteristics: it hydrated not only aliphatic and heterocyclic nitriles but also aromatic ones. Some substrates were also hydrated enantioselectively to the corresponding amides. The enantiomeric excess (ee) value of the enzyme hydrating trans-2,2-dimethyl-3-phenylcyclopropanecarbonitrile was 84.7. The enzyme is composed of two subunits: an alpha subunit and beta subunit of 22 975 Da and 23 493 Da, respectively. The optimal temperature and pH for the catalytic reaction of the enzyme was 25 degrees C and pH 7.6. The enzyme activity of the purified NHase was strongly inhibited by some oxidizing agents and heavy metals.
Song
Liya
L
State Key Laboratories of Transducer Technology, Institute of Microbiology, Chinese Academy of Sciences, Beijing, China.
Wang
Meixiang
M
Yang
Xiuqing
X
Qian
Shijun
S
D000595:Amino Acid Sequence; D004789:Enzyme Activation; D004795:Enzyme Stability; D006836:Hydro-Lyases / Q000737:chemistry* / Q000302:isolation & purification*; D008969:Molecular Sequence Data; D012240:Rhodococcus / Q000201:enzymology*; D013379:Substrate Specificity
D006836:Hydro-Lyases; C048831:nitrile hydratase