The impact of DNA intercalators on DNA and DNA-processing enzymes elucidated through force-dependent binding kinetics

Andreas S. Biebricher, Iddo Heller, Roel F. H. Roijmans, Tjalle P. Hoekstra, Erwin J. G. Peterman and Gijs J. L. Wuite ()
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Andreas S. Biebricher: LaserLaB Amsterdam, VU University Amsterdam
Iddo Heller: LaserLaB Amsterdam, VU University Amsterdam
Roel F. H. Roijmans: LaserLaB Amsterdam, VU University Amsterdam
Tjalle P. Hoekstra: LaserLaB Amsterdam, VU University Amsterdam
Erwin J. G. Peterman: LaserLaB Amsterdam, VU University Amsterdam
Gijs J. L. Wuite: LaserLaB Amsterdam, VU University Amsterdam

Nature Communications, 2015, vol. 6, issue 1, 1-12

Abstract: Abstract 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.

Date: 2015
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DOI: 10.1038/ncomms8304

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