Table 2 MD-predicted distances for the three FRET dye pairs for each replica (R1, R2, R3) in comparison to the experimental values

From: Single-molecule imaging and molecular dynamics simulations reveal early activation of the MET receptor in cells

Dye pair(a,b)

R1

R2

R3

Experiment

Fixed cell

Live cell

T-T

3.6 ± 0.1

4.1 ± 0.1

4.2 ± 0.2

4.8 ± 0.3

4.4 ± 0.4

T-H/H-T

5.9 ± 0.1

5.8 ± 0.1

6.3 ± 0.1

6.2 ± 0.4

6.1 ± 0.4 (H-T)

H-H

10.2 ± 0.1

9.8 ± 0.1

9.7 ± 0.1

11.4 ± 4.2

-(c)

  1. (a)The errors on the predicted distances were computed by summing the variances describing the statistical contribution and the experimental uncertainty on the R0 value and taking the square root of the resulting variance. The statistical uncertainty corresponds to a 66% confidence interval estimated by bootstrapping the distance distributions. We resampled the FRET efficiency 1000 times, computed using FRETpredict45 with repetition, and calculated the mean of each sample. We converted the 1000 bootstrapped means to distances (using the experimental R0 value for the FRET pair), and computed the standard deviation. For the T-H/H-T distance, we first converted the MD-calculated efficiency into distance prior to bootstrapping. We then applied bootstrapping to the merged distance series and computed the standard deviation. Our estimate of the uncertainty arising from the MD simulations does not include potential forcefield inaccuracies and should be considered a lower-bound estimate.
  2. (b)The experimental FRET distance uncertainty is calculated according to Hellenkamp et al. 56.
  3. (c)Live-cell FRET was measured using the smFRET-RAP approach. No FRET signal was detected for H-H labeled MET dimers in smFRET measurements in living cells.
  4. All values are given in nm.
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