Iron chelator-inducible expression system for Escherichia coli

Abstract

The P-entC promoter of the entCEBA operon encoding enzymes for enterobactin biosynthesis in Escherichia coli is tightly regulated by the availability of iron in the culture medium. In iron-rich conditions, the P-entC promoter activity is strongly repressed by the global transcription regulator Fur (ferric uptake regulator), which complexes with ferrous ions and binds to the Fur box 19-bp inverted repeat. In this study, we have constructed the expression vector pOS2 containing the P-entC promoter and characterized its repression, induction, and modulation by quantifying the expression of the lacZ, reporter gene encoding beta-galactosidase. beta-Galactosidase activities of E. coli transformants harboring pOS2-lacZ were highly induced in the presence of divalent metal ion chelators such as 2,2'-dipyridyl and EDTA, and were strongly repressed in the presence of excess iron. It was also shown that the basal level P-galactosidase expression by the P-entC promoter was drastically decreased by incorporating the fur gene into the expression vector. Since the newly developed iron chelator-inclucible expression system is efficient and cost-effective, it has wide applications in recombinant protein production.