High-throughput smFRET analysis of freely diffusing nucleic acid molecules and associated proteins

Abstract

High-throughput smFRET analysis of freely diffusing nucleic acid molecules and associated proteins, Methods (2019), doi:

Integrating single-molecule spectroscopy and simulations for the study of intrinsically disordered proteins

“…Similarly, microfluidic devices with fast mixing allows following the kinetics of the system of interest, at the single-molecule level, from hundreds of microseconds up to tens of seconds. [ 171 – 177 ]. Recurrence analysis of single particles (RASP) also captures the kinetics of freely diffusing molecules by identifying those molecules that after passing through the confocal volume re-enter in the confocal volume.…”

Dynamical control of denaturation bubble nucleation in supercoiled DNA minicircles

“…For example, the bacteriophage λ O protein and gal Repressor, which are well-characterized DNA replication initiator and E. coli transcription factors, were shown to induce excess linking numbers Lk = −0.222 ± 0.016 and Lk = −0.169 ± 0.004 on plasmid of about 4.8 kbps containing 16 and 18 sites, respectively [40]. We predict that the binding of such proteins on DNA molecules containing between 200 and 400 base-pairs, as studied here, would induce long-lived thermally assisted DNA denaturation bubbles with characteristic times τ closure in the order of hundreds of nanoseconds for the λ O protein (respectively the millisecond for the gal Repressor) and τ opening in the order of tens of milliseconds, which could be measured by smFRET on freely diffusing DNA [41].…”