The nrfA and nirB Nitrite Reductase Operons in Escherichia coli Are Expressed Differently in Response to Nitrate than to Nitrite

Abstract

ABSTRACT Escherichia coli possesses two distinct nitrite reductase enzymes encoded by the nrfA and nirB operons. The expression of each operon is induced during anaerobic cell growth conditions and is further modulated by the presence of either nitrite or nitrate in the cells' environment. To examine how each operon is expressed at low, intermediate, and high levels of either nitrate or nitrite, anaerobic chemostat culture techniques were employed using nrfA-lacZ and nirB-lacZ reporter fusions. Steady-state gene expression studies revealed a differential pattern of nitrite reductase gene expression where optimal nrfA-lacZ expression occurred only at low to intermediate levels of nitrate and where nirB-lacZ expression was induced only by high nitrate conditions. Under these conditions, the presence of high levels of nitrate suppressed nrfA gene expression. While either NarL or NarP was able to induce nrfA-lacZ expression in response to low levels of nitrate, only NarL could repress at high nitrate levels. The different expression profile for the alternative nitrite reductase operon encoded by nirBDC under high-nitrate conditions was due to transcriptional activation by either NarL or NarP. Neither response regulator could repress nirB expression. Nitrite was also an inducer of nirB and nrfA gene expression, but nitrate was always the more potent inducer by >100-fold. Lastly, since nrfA operon expression is only induced under low-nitrate concentrations, the NrfA enzyme is predicted to have a physiological role only where nitrate (or nitrite) is limiting in the cell environment. In contrast, the nirB nitrite reductase is optimally synthesized only when nitrate or nitrite is in excess of the cell's capacity to consume it. Revised regulatory schemes are presented for NarL and NarP in control of the two operons.