Methionine is not only an essential amino acid needed for human health,also an important feed additive.The recombinant E.coli BL21(DE3) /pET-28b-NIT producing nitrilase was investigated.Based on the data obtained from the cultivation using shake flask,the fed-batch fermentation strategy for producing nitrilase by high cell density cultivation of recombinant E.coli in the 5-Liter fermentor was optimized and constructed.It indicated that the concentration of carbon source,nitrogen source and inducer were the important factors.Under the optimized conditions of 14 h fed-batch fermentation,the cell mass concentration and the specific activity reached 13.7 gDCW/L and 36 990 U/L,which was 4.3 and 5.1 times higher than previous optimization,respectively.