dTAT1: An Unnatural Nucleoside Exhibiting Low Photocytotoxicity for Genetic Code Expansion

Area of Science:

• Biochemistry
• Molecular Biology
• Synthetic Biology

Background:

• Synthetic biology seeks to expand the genetic code for enhanced cellular information processing.
• Unnatural base pairs (UBPs) are key tools for genetic code expansion.
• dTAT1-dNaM is a novel UBP with improved photo- and thermostability over previous systems.

Purpose of the Study:

• To investigate the photophysical properties and cellular cytotoxicity of the dTAT1-dNaM unnatural base pair under UV light.
• To assess the safety and potential applications of dTAT1 in biological systems.

Main Methods:

• UV-Vis spectroscopy to analyze photophysical properties.
• Cellular assays to evaluate cytotoxicity and reactive oxygen species (ROS) production.
• Measurement of triplet state lifetime and singlet oxygen quantum yield.

Main Results:

• dTAT1 populates the triplet state upon 390 nm excitation.
• dTAT1 exhibits minimal cytotoxicity in cellular models.
• Low singlet oxygen quantum yield (17%) and superoxide generation were observed.
• dTAT1 possesses a 2.7-fold shorter triplet lifetime compared to dTPT3, correlating with reduced phototoxicity.

Conclusions:

• dTAT1-dNaM demonstrates favorable photophysical characteristics and low cellular toxicity.
• Its reduced photocytotoxicity suggests significant potential for safe genetic code expansion.
• Findings provide crucial data for designing next-generation UBPs for therapeutic and biotechnological applications.