摘要: 腈水解酶催化的腈水解具有反应高效、条件温和、环境污染小和成本低等优点，在有机合成、材料合成、医药、食品、农业、畜牧业及环境等污染方面有着重要的应用前景。本研究利用初步活化和分步胁迫富集，通过Berthelot法高通量筛选与高压液相精细筛选获得一株底物对4-羟基苯乙腈具有较好的催化活性的菌株；经过培养基初步优化，产酶量达到28.47 U/ml；铜离子对该酶具有较强的抑制作用，酶学性质研究表明该菌株能够在pH 6.2~pH 7.3之间酶活性能够保持在最高酶活性的80%以上，在35℃~45℃范围内催化活性大于最大酶活性的80%，反应18 h后离心菌体二次催化几乎不丧失活性；该菌株在开发对羟基苯乙腈合成对羟基苯乙酸具有较高的开发价值。
Abstract:
Nitrilases can hydrolyze nitrile efficiently under mild conditions. The enzymatic methods have the advantages of less pollution, low cost compared with the chemical methods. Nitrilases are potentially applied in agriculture, industry, environment, and biomedicine. In this study, Berthelot method and high performance liquid chromatography were used to screen new stains for nitrilases. A nitrilase with high substrate specificity for 4-hydroxyphenylcyanide was found from Pseudomonas sp. 6-1. The stain produced 28.47 U/ml nitrilase after optimization of the culture conditions. The enzyme remained 80% activities at pH 6.6 to pH 7.6, temperature 35˚C to 45˚C, and it was stable after 18 h incubation. The stain can be potentially used in biosynthesis 4-Hydroxyphenylacetic acid using 4-hydroxyphenylcyanide as substrate.