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. Author manuscript; available in PMC: 2020 Apr 30.
Published in final edited form as: Biochemistry. 2018 Nov 27;57(49):6797–6805. doi: 10.1021/acs.biochem.8b01072

Figure 6.

Figure 6.

Phosphorylation of LiaS increases affinity for LiaR. Microscale Thermophoresis was used to measure E. faecium LiaS binding to a LiaR mimetic peptide and to full length LiaR. The resulting Kd values were based on the average from at least three independent measurements. Bars indicate the SEM (standard error of the mean). The fraction bound was plotted on the y-axis in [%] unit against the total concentration of the titrated partner on a log10 scale on the x-axis27. (a) To determine Kd, increasing concentrations of LiaS (blue circles), LiaS in the presence of 0.2 mM ATP (light blue triangles) or adaptive mutant LiaST120A (magenta diamonds) were added to 84 nM of fluorescently labeled Peptide 3. (b) To determine Kd, we added increasing concentrations of Efm LiaR to 20 nM of fluorescently labeled LiaS (blue circles), LiaS in the presence of 0.2 mM ATP (light blue triangles) or in the presence of 2 mM AMP-PNP (light green squares).