Fine-Tuning Modulation of Oxidation-Mediated Posttranslational Control of FixK Transcription Factor.

FixK is a CRP/FNR-type transcription factor that plays a central role in a sophisticated regulatory network for the anoxic, microoxic and symbiotic lifestyles of the soybean endosymbiont . Aside from the balanced expression of the gene under microoxic conditions (induced by the two-component regulatory system FixLJ and negatively auto-repressed), FixK activity is posttranslationally controlled by proteolysis, and by the oxidation of a singular cysteine residue (C183) near its DNA-binding domain. To simulate the permanent oxidation of FixK, we replaced C183 for aspartic acid. Purified C183D FixK protein showed both low DNA binding and in vitro transcriptional activation from the promoter of the operon, required for respiration under symbiosis. However, in a strain coding for C183D FixK, expression of a fusion was similar to that in the wild type, when both strains were grown microoxically. The C183D FixK encoding strain also showed a wild-type phenotype in symbiosis with soybeans, and increased gene expression levels and FixK protein abundance in cells. These two latter observations, together with the global transcriptional profile of the microoxically cultured C183D FixK encoding strain, suggest the existence of a finely tuned regulatory strategy to counterbalance the oxidation-mediated inactivation of FixK in vivo.