A polarized view on DNA under tension.

Area of Science:

• Molecular Biology
• Biophysics
• Polymer Physics

Background:

• Fluorescence microscopy and single-molecule manipulation have advanced DNA dynamics and mechanics studies.
• Intercalating dyes enable quantitative measurement of DNA thermal fluctuations.
• Understanding DNA conformational dynamics under tension is crucial.

Purpose of the Study:

• To investigate DNA conformational dynamics under varying tensions using a combined approach.
• To disentangle rapid dye dynamics from DNA dynamics by controlling dye alignment.
• To resolve inconsistencies in intercalated dye orientation reports.

Main Methods:

• Utilized polarized fluorescence microscopy and optical-tweezers manipulation of YOYO-intercalated DNA.
• Controlled DNA tension to align YOYO dyes.
• Measured dye orientation and orientational dynamics relative to DNA tension.

Main Results:

• Intercalated dyes are oriented perpendicular to DNA but exhibit fast dynamics.
• No significant changes in dye orientation or dynamics were observed during DNA overstretching.
• Depolarization beyond overstretching suggests average DNA base pair tilting.

Conclusions:

• The combined approach provides unprecedented control over DNA alignment and dynamics measurement.
• Resolved dye orientation inconsistencies and characterized dye dynamics.
• Identified base pair tilting beyond the DNA overstretching transition, elucidating DNA molecular dynamics.