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. 2024 Jun 6;15(7):e01220-24. doi: 10.1128/mbio.01220-24

Fig 7.

EMSA results for BT4124 or BT4663 and variants AG and AD interacting with their respective promoter DNA; DNA binding curves for the same HTCS RR proteins under unphosphorylated and phosphorylated conditions.

REC-DBD interface substitutions relieve inhibition and allow DNA binding. (A–B) Binding of HTCS-RR proteins to the DNA detected by EMSAs. EMSAs were done with PCR-generated fluorescent DNA fragments in the presence of 0, 0.6, 1.2, 1.8, 2.4, 4, and 8 µM concentrations of the indicated proteins. Promoter DNA from bt4114 was used for BT4124 proteins (A), and promoter DNA from bt4662 was used for BT4663 proteins (B). A non-specific DNA fragment (“ns”) present in all lanes in panel A did not show any shift. (C–D) Comparison of DNA-binding curves of HTCS RR proteins. Percent decreases of band intensities for the free unbound DNA were quantified. RR, RRAG, and RRAD are colored black, violet, and pink, respectively. Data are shown as mean ± SD from at least three independent experiments. Solid (unphosphorylated) and dashed (phosphorylated) lines represent data fitted with the Hill equation.