Reduced DNA flexibility in complexes with a type II DNA binding protein.

Area of Science:

• Molecular biology
• Biophysics
• Structural biology

Background:

• Bacillus subtilis phage SPO1 DNA contains (hydroxymethyl)uracil instead of thymine.
• Internal molecular motions and torsional flexibility are key DNA properties.
• Transcription factor 1 (TF1) is a procaryotic DNA binding protein encoded by phage SPO1.

Purpose of the Study:

• To investigate the internal molecular motions and torsional flexibility of Bacillus subtilis phage SPO1 DNA.
• To determine the effect of transcription factor 1 (TF1) binding on DNA torsional flexibility.
• To explore the mechanism underlying TF1-mediated changes in DNA flexibility.

Main Methods:

• Time-resolved fluorescence polarization anisotropy (FPA) was used to study DNA.
• Ethidium was used as an intercalating fluorescent probe.
• An elastic DNA model was employed to fit FPA decay data.

Main Results:

• The torsional flexibility of SPO1 DNA is comparable to natural thymine-containing DNAs.
• TF1 binding significantly decreases DNA torsional flexibility, indicated by enhanced FPA.
• This effect is observed at low TF1 binding ratios and increases torsional rigidity up to 7-fold.
• Reduced ethidium binding affinity correlates with increased FPA.

Conclusions:

• TF1 binding substantially reduces the torsional flexibility of SPO1 DNA.
• Protein-induced DNA bending is proposed as the primary mechanism at low TF1 densities.
• These findings may offer insights into the biological role of TF1 in phage gene regulation.