Sequence Requirements of Intrinsically Fluorescent G-Quadruplexes

Area of Science:

• Biochemistry
• Molecular Biology
• Biophysics

Background:

• G-quadruplexes are four-stranded nucleic acid structures.
• These structures exhibit intrinsic fluorescence, suggesting potential applications in biosensing.
• Understanding factors influencing G-quadruplex fluorescence is crucial for sensor development.

Purpose of the Study:

• To systematically investigate how mutations affect the fluorescence intensity and emission wavelength of DNA G-quadruplexes.
• To explore the relationship between G-quadruplex sequence, structure, and fluorescence properties.
• To identify mutations that can enhance fluorescence for potential use in label-free sensors.

Main Methods:

• Generated and analyzed over 500 DNA G-quadruplex sequence variants.
• Systematically mutated tetrads, loops, and overhanging nucleotides.
• Measured fluorescence intensity and maximum emission wavelength for each variant.

Main Results:

• Certain mutations increased fluorescence intensity, while others shifted the maximum emission wavelength.
• A trade-off was observed between increased fluorescence intensity and shifted emission wavelength.
• Fluorescence properties correlated with the G-quadruplex's multimeric state (monomer vs. dimer).
• Oligonucleotides with multiple G-quadruplexes showed varied fluorescence depending on spacer sequences.

Conclusions:

• Mutations can modulate G-quadruplex fluorescence properties, offering distinct effects on intensity and wavelength.
• The multimeric state significantly influences fluorescence characteristics.
• These findings provide insights into fluorescent G-quadruplexes and support the development of novel label-free nucleic acid sensors.