Single-Molecule Analysis of Human Telomere Sequence Interactions with G-quadruplex Ligand

Area of Science:

• Biophysics
• Nanotechnology
• Medicinal Chemistry

Background:

• G-quadruplexes in human telomeres are key targets for anticancer drugs that inhibit telomerase.
• Understanding ligand interactions with telomeric G-quadruplexes at the single-molecule level is crucial for drug development.

Purpose of the Study:

• To analyze the interaction between the small molecule ligand pyridostatin (PDS) and human telomeric DNA G-quadruplexes using a single-molecule nanopore technique.
• To propose a nanopore thermodynamic analytical method for quantifying G-quadruplex/ligand interactions.

Main Methods:

• Utilized an α-hemolysin protein nanopore to study human telomeric DNA interactions with pyridostatin (PDS).
• Employed a nanopore thermodynamic analytical method to analyze binding and stabilization effects.
• Conducted translocation studies and free-energy change analysis to determine the ligand-binding mode.

Main Results:

• Pyridostatin (PDS) binding significantly prolonged the unraveling time of telomeric DNA G-quadruplexes, indicating potent stabilization.
• Observed two-level electronic blocks characteristic of a two-state unraveling process for K(+)-PDS-G-quadruplex complexes.
• Demonstrated a binding mode where PDS and K(+) simultaneously bind to the G-quadruplex, cooperatively stabilizing the structure.

Conclusions:

• The single-molecular nanopore platform provides an efficient, label-free method for determining ligand affinity constants for G-quadruplexes.
• This approach offers a general analytical tool for monitoring and quantifying G-quadruplex/ligand interactions.
• Findings have significant implications for the design and screening of novel anticancer drugs targeting telomeric G-quadruplexes.