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Applied Microbiology and Biotechnology

, Volume 89, Issue 3, pp 817–823 | Cite as

A high-throughput screening strategy for nitrile-hydrolyzing enzymes based on ferric hydroxamate spectrophotometry

  • Yu-Cai HeEmail author
  • Cui-Luan Ma
  • Jian-He XuEmail author
  • Li Zhou
Methods and Protocols

Abstract

Nitrile-hydrolyzing enzymes (nitrilase or nitrile hydratase/amidase) have been widely used in the pharmaceutical industry for the production of carboxylic acids and their derivatives, and it is important to build a method for screening for nitrile-hydrolyzing enzymes. In this paper, a simple, rapid, and high-throughput screening method based on the ferric hydroxamate spectrophotometry has been proposed. To validate the accuracy of this screening strategy, the nitrilases from Rhodococcus erythropolis CGMCC 1.2362 and Alcaligenes sp. ECU0401 were used for evaluating the method. As a result, the accuracy for assaying aliphatic and aromatic carboxylic acids was as high as the HPLC-based method. Therefore, the method may be potentially used in the selection of microorganisms or engineered proteins with nitrile-hydrolyzing enzymes.

Keywords

Nitrilase Carboxylic acids High-throughput screening Ferric hydroxamate spectrophotometry Evaluation 

Notes

Acknowledgments

This work was financially supported by the National Natural Science Foundation of China (No. 20506037), the Talent Introduction Foundation of Changzhou University (No. ZMF10020077), and the Students’ Science & Technology Creation Foundation of Changzhou University (No. 10-Chem-A-11). This work was also partially supported by the Open Project Program of the State Key Laboratory of Bioreactor Engineering (No. 2008004) and the Key Laboratory of Fermentation Engineering (Ministry of Education).

Supplementary material

253_2010_2977_MOESM1_ESM.doc (138 kb)
ESM 1 (DOC 137 kb)

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Copyright information

© Springer-Verlag 2010

Authors and Affiliations

  1. 1.Laboratory of Biochemical Engineering, College of Pharmaceutical and Life SciencesChangzhou UniversityChangzhouPeople’s Republic of China
  2. 2.State Key Laboratory of Bioreactor EngineeringEast China University of Science and TechnologyShanghaiPeoples Republic of China
  3. 3.Changzhou Kangpu Pharmaceutical Co., Ltd.ChangzhouPeople’s Republic of China
  4. 4.Key Laboratory of Fermentation Engineering (Ministry of Education)Hubei University of TechnologyWuhanPeople’s Republic of China

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