A Size-Expanded Fluorescent Ribonucleoside Genetic Set Bearing Canonical Watson-Crick Base Pairing Groups

Fluorescent ribonucleosides have found widespread use as probes and sensors to study the intricate properties of native RNAs in biological systems.  However, many of these molecules sacrifice the canonical Watson-Crick base pairing groups in their design for functionality that enhance their photophysical properties.  Decades ago, a novel set of adenosine and guanosine analogs, in which a benzene ring is incorporated into the purine scaffold thus expanding their size by 2.4 angstroms, were described and found to be excellent fluorophores.  However, their photophysical properties were not fully explored and the analogous benzo-expanded pyrimidine set have never been reported. 

In order to investigate the use of these size-expanded RNAs (xRNAs) as steric and fluorescent probes in RNA biology, we sought to explore an efficient synthesis of the entire xRNA genetic set.  Furthermore, we were interested in measuring their photophysical properties in various solvents in order to evaluate their utility in biological systems.  This project involves the use of chemical synthesis and biophysical techniques in order to acquire the data of interest.

Overall, we found that the xRNA free nucleosides are efficient fluorophores with red-shifted emission maxima of 369-411 nm and quantum yields ranging from 0.27 to 0.48.  Their extinction coefficients and fluorescence lifetimes were also determined.  In summary, the xRNA genetic set has been synthesized in excellent yield and purity.  Furthermore, we found that these fluorophores have a broad range of photophysical properties.  Future work will be directed toward the use of these compounds as nucleotide probes and in unnatural oligoribonucleotides.