A fungal isolate from Fusarium proliferatum strain AUF-2 has been found to have a high nitrilase activity (a parts per thousand yen1,000 U/l culture). The present work describes optimization of growth conditions and production medium to achieve maximum nitrilase production. The most effective carbon and nitrogen sources for nitrilase production were glucose and sodium nitrate, respectively. epsilon-Caprolactam was found to be the best inducer for maximum nitrilase production with 80 g/l wet cell biomass and 26 U/g nitrilase activity. An overall nitrilase activity of a parts per thousand yen2,000 U/l culture was obtained in this study, which is one of the best activities reported so far in any Fusarium strain. Chemo-profiling has shown that the strain is versatile in its ability to hydrolyze both aliphatic as well as aromatic nitriles. Efforts are being made to use the strain for biotransformation of pharmaceutical substrates.