Abstract
Characterized by their high structural similarity to their native counterparts, yet endowed with useful photophysical features, isomorphic fluorescent nucleosides present enormous opportunities. Here we discuss their fundamental design features, compare and contrast them to other families of fluorescent nucleoside analogs, as well as illustrate their utility and highlight potential limitations. The isomorphic nature of the analogs and their faithful representation of the canonical nucleosides enable their enzymatic, sequence-specific incorporation into RNA and DNA oligonucleotides, typically resulting in minimal structural perturbations. Their photophysical responsiveness, signaling changes in their microenvironment, can therefore be exploited for biophysical and biochemical applications, and expanded to high-throughput screening settings. The chapter specifically illustrates their application for fluorescently monitoring metabolically relevant enzymatic reactions involving nucleobases, nucleosides, cofactors, and oligonucleotides, as well as their utilization for biophysically studying folding and recognition of RNA and DNA oligonucleotides. Focusing on two families of fluorescent nucleoside analogs built upon thieno [3,4-d] pyrimidines and isothiazolo [4,3-d]-pyrimidines, which differ from one another by a single N atom, we further illustrate their utility as mechanistic probes, providing insight into the tolerance of diverse enzymes to minimal structural perturbations.
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Steinbuch, K.B., Tor, Y. (2023). Isomorphic Fluorescent Nucleoside Analogs. In: Sugimoto, N. (eds) Handbook of Chemical Biology of Nucleic Acids. Springer, Singapore. https://doi.org/10.1007/978-981-19-9776-1_17
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