Enhanced expression of cytochrome P450s from lac-based plasmids using lactose as the inducer

The cytochrome P450 expression systems used in Escherichia coli are highly regulated and involve the use of the lac repressor to control expression. I nduction in these systems utilizes the nonmetabolizable analog of lactose, isopropyl-beta -D-thiogalactopyranoside (IPTG), which is the most expensive compound required for an E. coli expression system. To determine if the na tural inducer lactose could be used to induce cytochrome P450 expression we examined the expression of three P450 enzymes in E. coli using two differe nt expression systems, pTrc99A and the T7-based PET22b vector. For both sys tems lactose was found to induce expression of active P450 to concentration s that exceeded the levels achieved with IPTG. A 20-liter fermentation of a P450 expression system in the pTrc plasmid in which lactose was used as th e inducer resulted in 2.4 mu mol P450/liter, with a total yield of 2 g of c ytochrome P450. The use of lactose for protein expression in E. coli should be broadly useful for the inexpensive, large-scale production of heterolog ous proteins in E. coli. (C) 2001 Academic Press.