Probing telomeric-like G4 structures with full or partial 2'-deoxy-5-hydroxyuridine substitutions

Area of Science:

• Biochemistry
• Molecular Biology
• Chemical Biology

Background:

• Guanine quadruplexes (G4s) are crucial secondary DNA structures.
• Understanding G4 stability and interactions is vital for therapeutic development.

Purpose of the Study:

• To investigate the impact of 2 -deoxy-5-hydroxyuridine (H) incorporation on G4 structure and stability.
• To assess the influence of H substitutions on G4 disruption by BLM helicase and ligand binding.

Main Methods:

• Synthesis and phosphoramidite incorporation of 2 -deoxy-5-hydroxyuridine into telomeric repeat G4 models.
• Characterization using CD, UV-Vis spectroscopy, native gel electrophoresis, and kinetic assays.
• Evaluation of BLM helicase activity and G4 ligand (PhenDC3) interactions.

Main Results:

• Partially H-substituted G4s adopted G4-like structures but showed reduced thermal stability.
• Central tetrad substitution led to denaturation; full substitution's structure remained uncertain.
• BLM helicase activity was unaffected by H substitution but influenced by topology and PhenDC3.

Conclusions:

• The H modification can be incorporated into G4s at specific positions without compromising stability.
• 2 -deoxy-5-hydroxyuridine may destabilize DNA and RNA quadruplexes, similar to 5-hydroxyuracil.
• The developed kinetic assay is a versatile tool for comparing G4 stabilities and ligand interactions.