Aminoglycoside binding to Oxytricha nova telomeric DNA

Area of Science:

• Biochemistry
• Medicinal Chemistry
• Molecular Biology

Background:

• Telomeric DNA sequences form G-quadruplex structures crucial for cancer cell survival.
• G-quadruplexes are targets for anti-cancer drug development due to their role in telomerase activity.
• Understanding ligand-DNA interactions with G-quadruplexes is key for designing potent inhibitors.

Purpose of the Study:

• Investigate the binding of aminoglycosides to the Oxytricha nova G-quadruplex DNA sequence.
• Determine the binding affinity and stoichiometry of neomycin and paromomycin with the quadruplex.
• Explore the binding site and mechanism of aminoglycoside interaction with G-quadruplex DNA.

Main Methods:

• Isothermal titration calorimetry (ITC) to quantify binding affinity and stoichiometry.
• Circular dichroism (CD) titration to assess structural changes upon ligand binding.
• Fluorescence intensity displacement (FID) assays to evaluate ligand displacement of a probe.
• Nuclear magnetic resonance (NMR) footprinting to identify ligand-DNA interaction sites.
• Computational docking studies to predict binding modes.

Main Results:

• Neomycin binds to the G-quadruplex in a 1:1 ratio with an association constant (Ka) of approximately 10^5 M^-1.
• Paromomycin showed a 2-fold weaker binding affinity compared to neomycin.
• FID assays identified neomycin as the most effective aminoglycoside studied for quadruplex binding.
• CD titrations with neomycin showed minimal changes in the CD signal.
• NMR and docking studies suggest aminoglycosides bind within the wide groove of the G-quadruplex.

Conclusions:

• Aminoglycosides, particularly neomycin, effectively bind to G-quadruplex DNA structures.
• The binding occurs in a 1:1 stoichiometry and primarily within the wide groove of the quadruplex.
• These findings support the development of aminoglycoside-based drugs for targeting G-quadruplexes in cancer therapy.