Fig. 3: Single-turnover observations of remodeling with LAGOON.

a, Representation of the labeling scheme for observing the DNA movement at the entry side during nucleosome sliding. The Cy5 fluorophore (red) attached to the exit-side H2B-120C is excited with the 638-nm laser and acts as a FRET donor, whereas the Alexa750 fluorophore (purple) attached to the entry-side DNA acts as a FRET acceptor. b, FRET histograms before (n = 855) and after (n = 726) the addition of Chd1 and 1 mM ATP. c, Representative donor (green), acceptor (red) and FRET (blue) time traces showing the remodeling of individual nucleosomes by Chd1 in the presence of 1 mM (left, n = 145) or 10 µM (right, n = 89) ATP. d, Representative donor (green), acceptor (red) and FRET (blue) time traces showing the remodeling of individual nucleosomes by Chd1 in a LAGOON experiment in the presence of 10 mM DMNPE-ATP with 0.30 W cm⁻² (left, n = 93), 1 W cm⁻² (middle, n = 91) or 3 W cm⁻² (right, n = 72) continuous uncaging. e, A simplified schematic of the single-turnover experiment. Average times are marked for stochastic processes. f, Representative donor (green), acceptor (red) and FRET (blue) time traces showing entry-side DNA movement during nucleosome sliding in the single-turnover regime in the presence of 10 mM DMNPE-ATP (n = 209). The purple dashed lines mark uncaging laser pulses (90-ms duration every 8 seconds, 100 W cm⁻² uncaging power density). Red lines in the FRET time traces represent the steps identified in the data using the AutoStepFinder method⁴⁸. g, Histograms of FRET change in the first observed step upon pulsed ATP uncaging with a pulse duration of 30 ms (top, n = 453), 90 ms (middle, n = 206) or 180 ms (bottom, n = 254). Black lines represent fits of the histograms with two Gaussian peaks. Pulse period, 8 seconds. The asterisk indicates that, for the second peak, its center and width were fixed for the 30-ms and 90-ms histograms to match the parameters derived from the second peak of the 180-ms histogram. a.u., arbitrary unit.