Proximity-Dependent Switchable ATP Aptasensors Utilizing a High-Performance FRET Reporter

Area of Science:

• Biochemistry
• Molecular Biology
• Nanotechnology

Background:

• Förster Resonance Energy Transfer (FRET) systems are crucial for molecular sensing.
• Developing FRET pairs with desirable properties like large Stokes shift and photostability is an ongoing challenge.
• DNA aptasensors offer high specificity for target molecule detection.

Purpose of the Study:

• To develop a high-performance FRET system using fluorescent molecular rotors for proximity-dependent DNA aptasensors.
• To create a pH-insensitive and photostable FRET pair for enhanced biosensing applications.
• To construct a pH-switched, proximity-induced fluorescent ATP aptasensor.

Main Methods:

• Synthesis of carboxylated benzothiazole-based molecular rotors for DNA labeling.
• Attachment of the developed FRET pair to a split ATP aptamer on a DNA nanoscaffold.
• Utilizing a bimolecular i-motif for pH-controlled switching of DNA proximity.
• Systematic investigation of proximity effects by altering linker length.

Main Results:

• Successful synthesis of green and red fluorescent DNA-labeled molecular rotors.
• Demonstration of efficient FRET between the synthesized dyes, comparable to Cy3/Cy5.
• Construction of a pH-switched ATP aptasensor with high sensitivity, selectivity, and reconfiguration.
• Insights into DNA proximity reactions and their physiological relevance.

Conclusions:

• Fluorescent molecular rotors enable a new class of high-performance FRET systems for DNA aptasensors.
• The developed FRET pair and ATP aptasensor exhibit promising characteristics for biosensing.
• Systematic linker length studies provide valuable understanding of DNA proximity dynamics.