A modular strategy for tailoring fluorescent biosensors from ribonucleopeptide complexes.

Area of Science:

• Biochemistry
• Molecular Biology
• Chemical Biology

Background:

• Fluorescent biosensors are vital for sensitive, reagentless detection of small molecules in therapeutics and diagnostics.
• Developing biosensors with specific detection wavelengths and concentration ranges from macromolecular receptors is challenging.

Purpose of the Study:

• To engineer a fluorescent adenosine triphosphate (ATP) sensor from an ATP-binding ribonucleopeptide (RNP) receptor.
• To establish a combinatorial strategy for creating tailored fluorescent biosensors for specific ligands.

Main Methods:

• Converted an ATP-binding RNP receptor into a fluorescent ATP sensor by complexing its RNA subunit with a pyrenyl-modified peptide.
• Generated libraries of fluorescent ATP-binding RNPs using selected RNA subunits and fluorophore-modified peptides.
• Screened libraries based on fluorescence intensity changes upon ligand binding to identify ATP or guanosine triphosphate (GTP) sensors.

Main Results:

• A stable fluorescent RNP complex was formed, exhibiting increased fluorescence intensity upon ATP binding.
• Screening yielded fluorescent sensors with emission wavelengths ranging from 390 to 670 nm.
• Titration analysis enabled the development of ATP sensors responsive to a wide range of ATP concentrations.

Conclusions:

• A modular RNP receptor strategy allows for the creation of customized fluorescent sensors for specific ligands.
• This method bypasses the need for detailed structural information of the macromolecular receptor.
• The developed fluorescent biosensors offer sensitive and reagentless detection capabilities.