Figure 1: A single-molecule FRET assay for monitoring telomerase–DNA interactions.
From: Coordinated DNA dynamics during the human telomerase catalytic cycle
(a) A model for telomere DNA repeat synthesis and RAP during the telomerase catalytic cycle. The precise location of the primer outside of the template hybrid is not well characterized and is therefore represented by a dashed line to denote an arbitrary path for the primer as it exits complex. (b) Cartoon schematics of the two hTR fragments used in telomerase reconstitutions. The pseudoknot fragment is shown with a U42-Cy3 modification. (c) (left) Schematic illustration of experimental geometry during smFRET telomerase binding experiments using total internal reflection fluorescence microscopy. (right) A representative single-molecule trajectory of U42-Cy3-labelled telomerase bound to the 18GGG/T13-Cy5 primer. (top) Raw donor Cy3 (green) and acceptor Cy5 (red) dye intensities and (bottom) calculated FRET values are plotted as a function of time. The sudden drop in the dye intensity traces represents irreversible photobleaching of the FRET dyes.
