ArnR, a novel transcriptional regulator, represses expression of the narKGHJI operon in Corynebacterium glutamicum.

Abstract

The narKGHJI operon that comprises putative nitrate/nitrite transporter (narK) and nitrate reductase (narGHJI) genes is required for the anaerobic growth of Corynebacterium glutamicum with nitrate as a terminal electron acceptor. In this study, we identified a gene, arnR, which encodes a transcriptional regulator that represses the expression of the narKGHJI operon in C. glutamicum cells under aerobic conditions. Disruption of arnR induced nitrate reductase activities of C. glutamicum cells and increased narKGHJI mRNA levels under aerobic growth conditions. DNA microarray analyses revealed that besides the narKGHJI operon, the hmp gene, which encodes flavohemoglobin, is negatively regulated by ArnR under aerobic conditions. Promoter-reporter assays indicated that arnR gene expression was positively autoregulated by its gene product, ArnR, under both aerobic and anaerobic conditions. Electrophoretic mobility shift assay experiments showed that purified hexahistidyl-tagged ArnR protein specifically binds to promoter regions of the narKGHJI operon and the hmp and arnR genes. A consensus sequence, TA(A/T)TTAA(A/T)TA, found in the promoter regions of these genes was demonstrated to be involved in the binding of ArnR. Effects on LacZ activity by deletion of the ArnR binding sites within the promoter regions fused to the reporter gene were consistent with the view that the expression of the narKGHJI operon is repressed by the ArnR protein under aerobic conditions, whereas the expression of the arnR gene is autoinduced by ArnR.