NMR-derived solution structure of a 17mer hydroxymethyluracil-containing DNA.

Area of Science:

• Molecular Biology
• Structural Biology
• Biophysics

Background:

• The bacteriophage SPO1 of Bacillus subtilis utilizes 5-(hydroxymethyl)-2 ahydrodeoxyuridine (hmU) in its DNA instead of thymine.
• This modification allows viral DNA to selectively bind to transcription factor 1 (TF1).

Purpose of the Study:

• To synthesize and structurally characterize a DNA molecule containing hmU at a TF1 binding site.
• To elucidate the structural basis for hmU-mediated selective binding to TF1.

Main Methods:

• Synthesis of a DNA duplex containing hmU.
• Nuclear Magnetic Resonance (NMR) spectroscopy for structural analysis.
• Restrained molecular dynamics calculations.

Main Results:

• Sequential assignment of proton chemical shifts for the hmU-containing DNA duplex.
• NMR data indicated the absence of Watson-Crick hydrogen bonding at the terminal base pairs.
• Molecular dynamics simulations generated a family of B-DNA structures with specific helical parameters.
• The hydroxyl groups of hmU influenced base orientation, with a more positive chi value compared to other nucleosides.
• Positive roll angles and small twists at hmU sites suggested opening of hmU-A base pairs towards the minor groove.

Conclusions:

• The hmU modification in DNA induces specific structural alterations, including changes in base pairing and helical geometry.
• These structural changes are likely responsible for the selective binding of viral DNA to TF1.
• The study provides insights into the structure-function relationship of modified nucleosides in DNA-protein interactions.