Affiliation:
1. School of Life and Pharmaceutical Sciences, Dalian University of Technology, Panjin 124221, China
2. Laboratory of Advanced Materials and Catalytic Engineering, School of Chemical Engineering, Dalian University of Technology, Dalian 116024, China
Abstract
Nitrile hydratase (NHase, EC 4.2.1.84) is an excellent biocatalyst that catalyzes the hydration of nitrile substances to their corresponding amides. Given its catalytic specificity and eco-friendliness, NHase has extensive applications in the chemical, pharmaceutical, and cosmetic industries. To improve the affinity between Rhodococcus erythropolis CCM2595-derived NHase ( ReNHase) and adiponitrile, this study used a semirational design to improve the efficiency of ReNHase in catalyzing the generation of 5-cyanopentanamide from adiponitrile. Enzyme kinetics analysis showed that Km of the mutant ReNHaseB:G196Y was 3.265 mmol l−1, which was lower than that of the wild-type NHase. The affinity of the mutant ReNHaseB:G196Y to adiponitrile was increased by 36.35%, and the efficiency of the mutant ReNHaseB:G196Y in catalyzing adiponitrile to 5-cyanopentamide was increased by 10.11%. The analysis of the enzyme-substrate interaction showed that the hydrogen bond length of the mutant ReNHaseB:G196Y to adiponitrile was shortened by 0.59 Å, which enhanced the interaction between the mutant and adiponitrile and, thereby, increased the substrate affinity. Similarly, the structural analysis showed that the amino acid flexibility near the mutation site of ReNHaseB:G196Y was increased, which enhanced the binding force between the enzyme and adiponitrile. Our work may provide a new theoretical basis for the modification of substrate affinity of NHase and increase the possibility of industrial applications of the enzyme.
Funder
Fundamental Research Funds for the Central Universities
Liaoning Revitalization Talent Program
Subject
General Physics and Astronomy,General Biochemistry, Genetics and Molecular Biology,General Materials Science,Biomaterials,General Chemistry
Cited by
1 articles.
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