Journal
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA
Volume 110, Issue 10, Pages 3859-3864Publisher
NATL ACAD SCIENCES
DOI: 10.1073/pnas.1213676110
Keywords
DNA structure; base-pair stability; DNA melting; single molecules; optical trapping
Categories
Funding
- Nederlandse Organisatie voor Wetenschappelijk Onderzoek (NWO)
- European Research Council
- NWO
- European Molecular Biology Organisation
- ATLAS, a European Commission
- Royal Netherlands Academy of Arts and Sciences
- Fondation ARC pour la Recherche sur le Cancer
- Agence Nationale de la Recherche
- Marie Curie early-stage training network
Ask authors/readers for more resources
Mechanical stress plays a key role in many genomic processes, such as DNA replication and transcription. The ability to predict the response of double-stranded (ds) DNA to tension is a cornerstone of understanding DNA mechanics. It is widely appreciated that torsionally relaxed dsDNA exhibits a structural transition at forces of similar to 65 pN, known as overstretching, whereby the contour length of the molecule increases by similar to 70%. Despite extensive investigation, the structural changes occurring in DNA during overstretching are still generating considerable debate. Three mechanisms have been proposed to account for the increase in DNA contour length during overstretching: strand unpeeling, localized base-pair breaking (yielding melting bubbles), and formation of S-DNA (strand unwinding, while base pairing is maintained). Here we show, using a combination of fluorescence microscopy and optical tweezers, that all three structures can exist, uniting the often contradictory dogmas of DNA overstretching. We visualize and distinguish strand unpeeling and melting-bubble formation using an appropriate combination of fluorescently labeled proteins, whereas remaining B-form DNA is accounted for by using specific fluorescent molecular markers. Regions of S-DNA are associated with domains where fluorescent probes do not bind. We demonstrate that the balance between the three structures of overstretched DNA is governed by both DNA topology and local DNA stability. These findings enhance our knowledge of DNA mechanics and stability, which are of fundamental importance to understanding how proteins modify the physical state of DNA.
Authors
I am an author on this paper
Click your name to claim this paper and add it to your profile.
Reviews
Recommended
No Data Available