Abstrakt
The transformation of benzonitrile into benzohydroxamic acid was performed by a cascade bienzymatic reaction involving nitrile hydration and acyl transfer of the intermediate benzamide onto hydroxylamine. The first step was catalyzed by either a cell-free extract from Rhodococcus erythropolis A4 (nitrile hydratase) or cell-free extracts from recombinant Escherichia colt strains expressing nitrile hydratases from Raoultella terrigena 77.1 and Klebsiella oxytoca 38.1.2: the biocatalyst in the second step was a cellfree extract from R. erythropolis A4 (amidase).
When using the cell-free extract from R. etythropolis A4 in the first step, the hydrolytic amidase activity was suppressed by ammonium ions, which, however, did not inhibit the acyl transfer reaction catalyzed by the same enzyme. Aromatic and aliphatic nitriles were examined as substrates of the recombinant nitrile hydratases by using a colorimetric assay of hydroxamic acids produced in a coupled acyl transfer reaction.