Native gel electrophoresis for verifying the binding of wild-type and mutant PmrD with PmrA. Sample solutions with various ratios of PmrD to PmrA were prepared in 20 μL of reaction buffer (20 mM Tris–HCl, pH 7.8, 5 mM MgCl2, and 100 mM KCl) and incubated at 4°C for 1 h. The samples were loaded onto a 4% native polyacrylamide gel that was pre-run at 12 V/cm for 20 min, and they were analyzed at room temperature at 12 V/cm for 80 min in 1X TBE buffer. The gels were stained with Coomassie Blue. (A) Wild-type PmrD. (B) Mutant PmrD.