Journal: Nature Communications

Article Title: Sequence-specific dynamic DNA bending explains mitochondrial TFAM’s dual role in DNA packaging and transcription initiation

doi: 10.1038/s41467-024-49728-6

Figure Lengend Snippet: A LSP DNA labeled with a donor/acceptor dye pair for single-molecule FRET (smFRET) assay. B Schematic of smFRET assay to study real-time, single-molecule DNA bending dynamics of TFAM. C , D , G A sample smFRET time trace (left) and FRET histogram (right) for LSP DNA alone (N Trace = 288), LSP + TFAM (N Trace = 420), and LSP + TFAM C-tail mutant (ΔCT) (N Trace = 219), respectively. E FRET transition density plot (TDP) for LSP + TFAM, showing two-state kinetics. F Dwell time histograms of the low FRET state, i.e., P-state (left), and the high FRET state, i.e., F-state (right). Red curves indicate the fitted single- and double-exponential models for P- and F-state, respectively. H Ensemble FRET spectra, showing TFAM-induced DNA bending at the ensemble level in solution. I Diagram summarizes the two-state DNA bending kinetics of the LSP-TFAM complex. The forward and reverse rate constants (~24 s −1 and ~1 s −1 ) were calculated from the inverse of P-state and F-state mean dwell times (see Supplementary Table ). smFRET time traces were fitted with Hidden Markov Model using vbFRET or ebFRET MATLAB software packages to plot FRET histograms, TDP, and dwell time histograms in this figure.

Article Snippet: The forward and reverse rate constants (~24 s −1 and ~1 s −1 ) were calculated from the inverse of P-state and F-state mean dwell times (see Supplementary Table ). smFRET time traces were fitted with Hidden Markov Model using vbFRET or ebFRET MATLAB software packages to plot FRET histograms, TDP, and dwell time histograms in this figure.

Techniques: Labeling, Smfret Assay, Mutagenesis, Software