HgII binds to C-T mismatches with high affinity

Area of Science:

• Biochemistry
• Molecular Biology
• Chemical Biology

Background:

• DNA mismatches are common genetic alterations.
• Metal ions can interact with DNA, influencing its structure and function.
• Understanding metal-DNA interactions is crucial for various biological and therapeutic applications.

Purpose of the Study:

• To characterize the binding of Mercury(II) (HgII) and Silver(I) (AgI) ions to pyrimidine-pyrimidine mismatches in DNA.
• To compare the binding kinetics and thermodynamics of HgII and AgI to different mismatch types.
• To evaluate the utility of fluorescent nucleobase analogs for detecting metal-mediated base pairs.

Main Methods:

• Fluorescent nucleobase analogs
• Thermal denaturation assays
• Proton nuclear magnetic resonance (1H NMR) spectroscopy

Main Results:

• HgII exhibited stoichiometric, site-specific binding to C-T mismatches, unlike AgI.
• HgII binding kinetics were faster for C-T mismatches (kon ≈ 10^5 M^-1s^-1, koff ≈ 10^-3s^-1) compared to T-T mismatches (kon ≈ 10^4 M^-1s^-1, koff ≈ 10^-4s^-1).
• Equilibrium binding affinities (Kd ≈ 10-150 nM) were similar for both mismatch types, despite significant differences in thermal stability changes observed via denaturation assays.

Conclusions:

• C-HgII-T base pairs possess high thermodynamic and kinetic stability.
• Fluorescent nucleobase analogs provide a sensitive method for detecting and characterizing metal-mediated base pairs.
• This technique is effective even when metal binding does not significantly alter DNA duplex thermal stability.