Solution structure of the C-terminal transcriptional activator domain of FixJ from Sinorhizobium meliloti and its recognition of the fixK promoter.

FixJ is a response regulator of the two-component signal transduction pathway involved in the transcriptional activation of nitrogen fixation genes of Sinorhizobium meliloti. Upon phosphorylation, FixJ transcriptionally activates the fixK and nifA promoters. We identified a FixJ recognition sequence of 16 bp in the high affinity binding site of the fixK promoter by means of a gel shift assay.

The uncoupling of oxygen sensing, phosphorylation signalling and transcriptional activation in oxygen sensor FixL and FixJ mutants.

The rhizobial FixL/FixJ system, a member of the superfamily of bacterial two-component signal transducing systems, regulates the expression of nitrogen fixation-related genes by sensing environmental oxygen tension. Oxygen-free (deoxy) FixL is autophosphorylated at an invariant histidine residue with ATP, and the phosphoryl group is transferred to FixJ, leading to an enhancement in transcriptional activity at low oxygen tensions, but the histidine kinase activity of the oxygen-bound (oxy) form is inhibited. To investigate the mechanism of oxygen sensing, we established a FixL/FixJ-mediated PfixK-lacZ reporter system in Escherichia coli, and isolated FixL and FixJ mutations conferring an upregulation of lacZ gene expression on the reporter cells even under aerobic conditions.