Purification and Characterization of Nitrilase from Fusarium solani IMI196840

Authors

Vejvoda, V., Kubac, D., Davidova, A., Kaplan, O., Sulc, M., Sveda, O., Chaloupkova, R., Martinkova, L.

Source

PROCESS BIOCHEMISTRY 45: 1115-1120 (2010)

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.

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Citation

Vejvoda, V., Kubac, D., Davidova, A., Kaplan, O., Sulc, M., Sveda, O., Chaloupkova, R., Martinkova, L., 2010: Purification and Characterization of Nitrilase from Fusarium solani IMI196840. Process Biochemistry 45: 1115-1120.

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