Fig. 2.

DNA binding facilities CcrM-Lon recognition. (A) Schematic view of DNA probe designs according to genome locus. Probe 2 is the same as probe 1 except with the mutation of GATTC to AATAC. Probe 3 contains the upstream sequence of pliA. P1-P2 and P3-P4 are primer pairs to amplify probe 1 (or probe 2) and probe 3, respectively. CcrM methylation sites are shown circled “M.” (B) The direct binding of purified LonS674A to CcrM or probe 1 was measured in vitro by microscale thermophoresis. LonS674A was fluorescently labeled with Atto-488 dye. The concentration of LonS674A₆ was held constant at 20 nM while CcrM or probe 1 was titrated in 2-fold serial dilutions against it. The purified proteins were incubated at room temperature for 10 min before the binding assay. The data report the fraction of LonS674A₆ that is bound at each concentration of CcrM or probe 1. See Materials and Methods for description of curve fits. (C) A schematic view showing affinities between CcrM, Lon, and DNA. CcrM and Lon have higher affinities to DNA than that of direct interaction.