Development of nitrilase-mediated process for phenylacetic acid production from phenylacetonitrile

This study aimed at developing an efficient biotransformation process for phenylacetic acid production from phenylacetonitrile by using recombinant Escherichia coli M15 harboring a double mutant MG nitrilase (I113M/Y199G) from Burkholderia cenocepacia J2315. A yield of 2310 U/mL nitrilase was obtained by fermentation after the optimization of cultivation conditions, with a specific activity of 64 U/mg dcw. The MG nitrilase showed high substrate tolerance and completely hydrolyzed 100 mM phenylacetonitrile in 30 min under optimal conditions. To alleviate substrate inhibition, periodic or continuous batch-feeding of substrate was used during the biotransformation. Up to 164 g/L substrate was completely hydrolyzed in 9 h with continuous batch-feeding using resting cells, corresponding to 400 U/mL of nitrilase activity, and leading to production of 163.4 g/L phenylacetic acid. The hydrolysis process has potential application for phenylacetic acid production on a large scale.