FIG. 3.

Comparison of measured apparent D-A distances, ${\overline{r}}_{D-A,E}$, with expected D-A distances, ${\overline{r}}_{D-A}$. The experimentally derived ${\overline{r}}_{D-A,E}$ values of different conformational states for each D-A labeled lacCONS constructs are compared with the expected D-A distances from RPo crystal structure (pdb code: 4XLN) in free DNA (black dot) and in RPo (red dot). Two conformational states were found (with sub-millisecond underlying interconversion dynamics) in the RNAP-bound fraction. Their derived distance values are shown (magenta open squares) for the subset that defines a conformation closest to the known RPo structure. Distances for the other conformation are also shown (blue open diamonds). The values of the free promoter fraction are also presented (grey triangles; for the D-A labeled promoter constructs that do not exhibit quenched FRET). Quenched FRET constructs had an expected D-A distance below 0.5R₀ (shown as shaded area). The error bars in the measured distances represent experimental standard error (Eq. S21 of the supplementary material). The error bars in the expected distances represent the D-A standard deviation, as calculated from all possible donor and acceptor positions in space (dye AVs).