Phosphorylation-induced dimerization of the FixJ receiver domain

The 'two-component' transcriptional activator FixJ controls nitrogen fixati on in Sinorhizobium meliloti, Phosphorylation of FixJ induces its dimerizat ion, as evidenced by gel permeation chromatography and equilibrium sediment ation analysis. Phosphorylation-induced dimerization is an intrinsic proper ty of the isolated receiver domain FixJN, Accordingly, chemical phosphoryla tion of both FixJ and FixJN are second-order reactions with respect to prot ein concentration. However, the second-order phosphorylation constant is 44 -fold higher for FixJN than for FixJ, Therefore, the C-terminal transcripti onal activator domain FixJC inhibits the chemical phosphorylation of the re ceiver domain FixJN. Conversely, FixJN has been shown previously to inhibit FixJC activity approximate to 40-fold, reflecting the interaction between FixJN and FixJC. Therefore, we propose that modulation of FixJ activity inv olves both its dimerization and the disruption of the interface between Fix JN and FixJC, resulting in the opening of the protein structure. Alanine sc anning mutagenesis of FixJN indicated that the FixJ similar to P dimerizati on interface involves Val-91 and Lys-95 in helix alpha 4. Dimerization was required for high-affinity binding to fixK promoter DNA.