Raman Scattering Reveals Ion-Dependent G-Quadruplex Formation in the 15-mer Thrombin-Binding Aptamer upon Association...

Area of Science:

• Biochemistry
• Biophysics
• Analytical Chemistry

Background:

• DNA aptamers enable biosensing through selective binding and conformational changes.
• Characterizing immobilized aptamer conformations and their in situ responses is challenging.
• Thrombin-binding aptamer (TBA) forms a G-quadruplex for α-thrombin detection, influenced by metal cations.

Purpose of the Study:

• To develop a Raman spectroscopy method for characterizing immobilized aptamer conformations.
• To quantify the effects of K+ and Li+ on TBA G-quadruplex versus unfolded populations.
• To investigate aptamer conformational changes upon α-thrombin binding in different ionic conditions.

Main Methods:

• Developed a structurally informative Raman spectroscopy method.
• Immobilized the 15-mer thrombin-binding aptamer (TBA) on porous silica surfaces.
• Quantified aptamer conformations using Raman spectroscopy in the presence of K+ and Li+.

Main Results:

• In K+ solutions, TBA remained in a G-quadruplex conformation before and after α-thrombin binding.
• In Li+ solutions, TBA was unfolded but formed a G-quadruplex upon α-thrombin association.
• Raman scattering detected α-thrombin binding to immobilized aptamers.

Conclusions:

• Raman spectroscopy can determine immobilized aptamer conformations and their response to target binding.
• Aptamer conformational changes are dependent on ionic environment and target presence.
• This method advances understanding of aptamer-based biosensing mechanisms.