Nucleic acid function relies on chemical change and conformational dynamics, including While performing their essential functions, nucleic acids encounter a variety of disturbances, such as strand cleavage and ligation, local conformational changes, base damage, flipping and chemical modification. Fluorescent nucleosides continue to play a major role in the biophysical study of these processes. Some fluorescent nucleoside analogues maintain their brightness when base stacked, while others are quenched. Our lab has Purse Lab designed and synthesized a tricyclic cytidine analogue, 8-diethylamino-tC, which increases its fluorescence 20-fold once upon incorporation into a DNA duplex with the correctmatched Watson-Crick base pairing. This turn-on response expends extends the applications of this analogue as a fluorescent turn-on probe for base pairing or enzymatic nucleic acid synthesispolymerization. While this analogue has been somewhat studied in DNA, it has not been characterized thoroughly in RNA, like most reported base analogueswhere structural dynamics are even more important for function. By preparing and studying DNA–RNA heteroduplexes containing DEAtC, we show that DEAtC’s fluorescence turn-on properties are enhanced when base paired with RNA and in an A-form conformation. Here we report on the synthesis of a DEAtC ribonucleoside phosphoramidite for incorporation into RNA, the properties of the resulting RNA oligonucleotides, and the fluorescence of DEAtC in this context. These advanced studies will giveprovide a foundation for future applications of fluorescent nucleosides to the investigation of nucleic acid transport, metabolism, regulatory processes, and interactions with proteins and small molecules.

Fluorescence turn-on sensing of nucleic acid hybridization by a tricyclic cytidine analogue

ACS Spring 2022

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