Site-directed mutation effect of the symmetry region at the mRNA 5′-end of Escherichia coli aeg-46.5 gene

The age-46.5 gene of Escherichia coli is induced by nitrate ion and regulated by Fnr, NarL, and NarP during anaerobic growth. aeg-46.5::lacZ fusion gene shows its maximum expression in narL host after two hours of aerobic to anaerobic switch in M9-Glc-nitrate medium. Fnr and NarP act as positive regulators, and NarL acts as a negative regulator. The control region of the aeg-46.5 was identified and the binding sites of regulator proteins have been predicted (Reznikoff and Choe (1993)). It has two symmetry regions. One is located at -52∼-37 bp from the anaerobic mRNA 5′-end, which is the binding site of NarL and NarP. The other is located at +37∼+56 bp from the 5′-end of mRNA. In this study, the downstream symmetry region from the mRNA 5′-end was investigated by site-directed mutagenesis. The destruction of the symmetry region increases the expression level of aeg-46.5. We propose that the symmetry region interferes with the expression of aeg-46.5 possibly by forming a stem-and-loop structure.