Purification and characterization of nitrilase from Fusarium solani IMI196840 |
| |
| Institution: | 1. Institute of Microbiology, Centre of Biocatalysis and Biotransformation, Academy of Sciences of the Czech Republic, Vídeňská 1083, CZ-142 20 Prague, Czech Republic;2. Department of Biochemistry, Faculty of Science, Charles University Prague, Hlavova 8, CZ-128 40 Prague, Czech Republic;3. Loschmidt Laboratories, Department of Experimental Biology and Centre of Biocatalysis and Biotransformation, Faculty of Science, Masaryk University, Kamenice 5/A4, CZ-625 00 Brno, Czech Republic;1. Key Laboratory of Bioorganic Synthesis of Zhejiang Province, College of Biotechnology and Bioengineering, Zhejiang University of Technology, Hangzhou, Zhejiang 310014, PR China;2. Engineering Research Center of Bioconversion and Biopurification of the Ministry of Education, Zhejiang University of Technology, Hangzhou, Zhejiang 310014, PR China;1. Key Laboratory of Bioorganic Synthesis of Zhejiang Province, College of Biotechnology and Bioengineering, Zhejiang University of Technology, Hangzhou, 310014, China;2. Engineering Research Center of Bioconversion and Biopurification of Ministry of Education, Zhejiang University of Technology, Hangzhou, 310014, China;1. Faculty of Applied Sciences and Biotechnology, Shoolini University, Solan 173229, India;2. Faculty of Basic Sciences, Shoolini University, Solan 173229, India;1. Departamento de Bioquímica y Biología Molecular III e Inmunología, Universidad de Granada (Campus de Melilla), 52071, Melilla, Spain;2. Laboratorio de Estudios Cristalográficos, CSIC-UGR, 18100, Granada, Spain;1. Laboratory of Biocatalysis and Synthetic Biotechnology, State Key Laboratory of Bioreactor Engineering and Shanghai Collaborative Innovation Center for Biomanufacturing, East China University of Science and Technology, 130 Meilong Road, Shanghai 200237, China;2. Department of Biotechnology, College of Life Sciences, Ritsumeikan University, Shiga 525-8577, Japan |
| |
| Abstract: | Nitrilase activity in Fusarium solani IMI196840 (approx. 1500 U l?1 of culture broth) was induced by 2-cyanopyridine. The enzyme was purified by a factor of 20.3 at a yield of 26.9%. According to gel filtration, the holoenzyme was an approx. 550-kDa homooligomer consisting of subunits with a molecular weight of approximately 40 kDa, as determined by SDS-PAGE. Mass spectrometry analysis of the tryptic fragments suggested a high similarity of this enzyme to the hypothetical CN hydrolases from Aspergillus oryzae, Gibberella zeae, Gibberella moniliformis and Nectria haematococca. Circular dichroism and fluorescence spectra indicated that secondary structure content and overall tertiary structure, respectively, were almost identical in nitrilases from F. solani IMI196840 and F. solani O1. The melting temperatures of the enzymes were 49.3 °C and 47.8 °C, respectively. The best substrates for the purified nitrilase from F. solani IMI196840 were benzonitrile and 4-cyanopyridine, which were hydrolyzed at the rates of 144 and 312 U mg?1 protein, respectively, under the optimum conditions of pH 8 and 45 °C. The enzyme was highly chemoselective, producing ≤2% amides as by-products. |
| |
| Keywords: | |
| 本文献已被 ScienceDirect 等数据库收录！ |
|
