Fig. 3: Investigation of Mn2+ riboswitch docking in the presence and absence of RNAP by smFRET.
From: A nascent riboswitch helix orchestrates robust transcriptional regulation through signal integration
a smFRET experimental setup. Paused Elongation Complex (PEC) immobilization utilizes biotinylated E. coli RNAP. Locked Nucleic Acid (LNA) immobilization utilizes a complementary biotinylated LNA capture probe. Location of donor (Cy3 – green) and acceptor (Cy5 – red) fluorophores are indicated. Representative donor-acceptor (top) and smFRET (bottom) traces for PEC-104 in the absence (b) and presence (c) of 0.5 mM Mn2+. Probability FRET histogram for each trace is indicated on the right. smFRET histograms of PEC-104 (d), LNA-104 (e) in the presence of only Mg2+ (middle panel) and in the presence of both Mg2+ and Mn2+ (right panel). Cartoon of each construct is indicated on the left. The SD of each FRET value is reported. The total number of traces (N) included in each histogram is indicated on top right corner. Source data are provided with this paper at https://doi.org/10.7302/22513.
