Escherichia coli TG1 transformed with an expression plasmid pMS carrying the gene encoding Phytolacca insularis protein (PIP) under the control of the tac promoter was cultivated in Various conditions to improve the productivity of PIP. Production of PIP was dramatically increased by the addition of yeast extract as a nitrogen source in both batch and fed-batch cultures. High levels of PIP were obtained by adopting a fed-batch process and an optimized induction strategy. The optimized fed-batch culture resulted in 59.0 g of cell mass and 466.5 mg of soluble PIP per litre of culture, corresponding to a 9.1 fold increase in cell mass and a 17.3 fold enhancement in PIP production compared with a simple batch culture in Luria broth. (C) 2001 Elsevier Science Ltd. All rights reserved.