Cyanide hydratase from Aspergillus niger K10: Overproduction in Escherichia coli, purification, characterization and use in continuous cyanide degradation

A cyanide hydratase from Aspergillus niger K10 was expressed in Escherichia coli and purified. Apart from HCN, it transformed some nitriles, preferentially 2-cyanopyridine and fumaronitrile. Vmax and Km for HCN were ca. 6.8 mmol min−1 mg−1 protein and 109 mM, respectively. Vmax for fumaronitrile and 2-cyanopyridine was two to three orders of magnitude lower than for HCN (ca. 18.8 and 10.3 μmol min−1 mg−1, respectively) but Km was also lower (ca. 14.7 and 3.7 mM, respectively). Both cyanide hydratase and nitrilase activities were abolished in truncated enzyme variants missing 18-34 C-terminal aa residues. The enzyme exhibited the highest activity at 45 °C and pH 8-9; it was unstable at over 35 °C and at below pH 5.5. The operational stability of the whole-cell catalyst was examined in continuous stirred membrane reactors with 70-mL working volume. The catalyst exhibited a half-life of 5.6 h at 28 °C. A reactor loaded with an excess of the catalyst was used to degrade 25 mM KCN. A conversion rate of over 80% was maintained for 3 days.