FIG. 4.

Comparison of wild-type OmpR and PrS₂-OmpR. (a) Complex formation between EnvZc and PrS₂-OmpR. EnvZc and PrS₂-OmpR (lane 4) or OmpR (lane 2) were mixed and incubated in the reaction buffer at room temperature for 5 min. The samples were subjected to 10% native PAGE. +, present; −, absent. (b) Phosphotransfer from EnvZc-P to PrS₂-OmpR. Purified ³²P-labeled EnvZc was mixed with OmpR, PrS₂-OmpR, or the mixture of OmpR and PrS₂-OmpR in the reaction buffer. The final reaction mixtures were incubated at room temperature. Aliquots were removed at 20, 40, 60, and 120 s, and the reaction was stopped with 5× SDS loading buffer. (c) Dephosphorylation of PrS₂-OmpR-P by EnvZc. First, purified ³²P-labeled EnvZc-P was mixed with OmpR, PrS₂-OmpR, or the mixture of OmpR and PrS₂-OmpR, and the final mixture was incubated in the reaction buffer at room temperature for 2 min to generate OmpR-P or PrS₂-OmpR-P. After the addition of ADP (1 mM), aliquots were removed at 20, 40, 60, and 120 s and the reaction was stopped with 5× SDS loading buffer.