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. Author manuscript; available in PMC: 2016 Aug 20.
Published in final edited form as: Mol Cell. 2015 Jul 23;59(4):541–552. doi: 10.1016/j.molcel.2015.06.030

Figure 4. Analysis of ATP binding to the loading protomers by single molecule FRET.

Figure 4

(A) Schematics for the single molecule FRET measurements.

(B) Left column: smFRET Histograms for Ded1p binding to the RNA with increasing concentrations of ATP/Mg2+, as indicated. Grey curves represent best fits to individual Gaussian distributions, black curves the sum of the individual distributions. Right column: representative time traces for reactions in the presence of increasing concentrations of ATP/Mg2+, as indicated in the histograms. Dashed lines mark the three FRET states.

(C) Association and dissociation rate constants for ATP/Mg2+, and associated equilibrium dissociation constants.

(D) Fraction of each FRET state calculated from smFRET histograms as function of the ATP/Mg2+ concentration. Lines are the best fit of the experimental data to the model shown in panel (C). See also Figure S4.