TY - JOUR
T1 - The impact of DNA intercalators on DNA and DNA-processing enzymes elucidated through force-dependent binding kinetics
AU - Biebricher, A.S.
AU - Heller, I.
AU - Roijmans, R.F.H.
AU - Hoekstra, T.P.
AU - Peterman, E.J.G.
AU - Wuite, G.J.L.
PY - 2015
Y1 - 2015
N2 - DNA intercalators are widely used as fluorescent probes to visualize DNA and DNA transactions in vivo and in vitro. It is well known that they perturb DNA structure and stability, which can in turn influence DNA-processing by proteins. Here we elucidate this perturbation by combining single-dye fluorescence microscopy with force spectroscopy and measuring the kinetics of DNA intercalation by the mono- and bis-intercalating cyanine dyes SYTOX Orange, SYTOX Green, SYBR Gold, YO-PRO-1, YOYO-1 and POPO-3. We show that their DNA-binding affinity is mainly governed by a strongly tension-dependent dissociation rate. These rates can be tuned over a range of seven orders of magnitude by changing DNA tension, intercalating species and ionic strength. We show that optimizing these rates minimizes the impact of intercalators on strand separation and enzymatic activity. These new insights provide handles for the improved use of intercalators as DNA probes with minimal perturbation and maximal efficacy.
AB - DNA intercalators are widely used as fluorescent probes to visualize DNA and DNA transactions in vivo and in vitro. It is well known that they perturb DNA structure and stability, which can in turn influence DNA-processing by proteins. Here we elucidate this perturbation by combining single-dye fluorescence microscopy with force spectroscopy and measuring the kinetics of DNA intercalation by the mono- and bis-intercalating cyanine dyes SYTOX Orange, SYTOX Green, SYBR Gold, YO-PRO-1, YOYO-1 and POPO-3. We show that their DNA-binding affinity is mainly governed by a strongly tension-dependent dissociation rate. These rates can be tuned over a range of seven orders of magnitude by changing DNA tension, intercalating species and ionic strength. We show that optimizing these rates minimizes the impact of intercalators on strand separation and enzymatic activity. These new insights provide handles for the improved use of intercalators as DNA probes with minimal perturbation and maximal efficacy.
UR - https://www.scopus.com/pages/publications/84935036158
UR - https://www.scopus.com/inward/citedby.url?scp=84935036158&partnerID=8YFLogxK
U2 - 10.1038/ncomms8304
DO - 10.1038/ncomms8304
M3 - Article
SN - 2041-1723
VL - 6
SP - 7304
JO - Nature Communications
JF - Nature Communications
M1 - 7304
ER -