A Broccoli aptamer chimera yields a fluorescent K⁺ sensor spanning physiological concentrations.

Area of Science:

• Biochemistry
• Molecular Biology
• RNA Therapeutics

Background:

• RNA aptamers are crucial in molecular biology and therapeutics.
• The Broccoli aptamer is known for its fluorescence properties.
• Incorporating modified nucleotides can alter RNA function.

Purpose of the Study:

• To investigate the effect of 2'fluorinated pyrimidine incorporation on the Broccoli RNA aptamer.
• To assess the impact of these modifications on potassium ion (K+) binding and fluorescence.
• To develop an improved RNA-based sensor for physiological potassium concentrations.

Main Methods:

• Synthesis of Broccoli RNA variants with 2'fluorinated pyrimidines.
• Characterization of RNA structure and fluorescence in the presence of potassium and DFHBI.
• Affinity measurements for potassium ions.
• Construction and testing of a chimeric RNA sensor.

Main Results:

• 2'fluorinated pyrimidine incorporation did not perturb Broccoli's structure or fluorescence with potassium and DFHBI.
• Apparent affinity for potassium ions decreased more than 30-fold.
• A chimeric RNA with mixed chemistries showed linear fluorescent gain across physiological potassium concentrations.

Conclusions:

• Modified Broccoli aptamers retain key functional properties while enabling new sensor applications.
• Chimeric RNA aptamers offer a promising platform for developing sensitive, RNA-based ion sensors.
• This work advances the design of functional RNA molecules for biological sensing.