The effects of culturing conditions on phenylalanine ammonia lyase production by a recombinant E. coli strain were investigated by using a controlled fed-batch fermentation system. In a 5 L fermentor, the optimal composition of the batch medium was 2% glucose, 1% yeast extract, 0.7% K2HPO4, 0.8% KH2PO4, 0.5% (NH4)2SO4, 0.1% MgSO4·7H2O. The optimal feed glucose solution was 50%. Glucose concentration and pH of the culture broth were maintained at about 2.0 g/L and 7.0 during the fed-batch phase, respectively. Following 24-h cultivation, 0.2 mmol/L isopropyl-β-D-thiogalactopyranoside (IPTG) was added and temperature was shifted from 37 ℃ to 42 ℃ to induce pal gene expression. Under optimal conditions, a high productivity of 300 U/g could be achieved after 48 h culture, and a cell density of OD600 about 82 was obtained at 52 h culture at 500 r/m stirrer speed and 1 vvm, respectively.