Please use this identifier to cite or link to this item: http://hdl.handle.net/1946/13212
To understand the function of nucleic acids in biological systems, their structural information is important. Electron paramagnetic resonance (EPR) spectroscopy has been increasingly used to study spin-labeled nucleic acids. Spin labels are usually incorporated site-specifically through covalent bonding, which can be difficult and time consuming. A new method was previously developed in our lab using noncovalent interactions where a spin label binds to an abasic site in a duplex DNA at low temperatures. However, the spin label (ҫ) that was used has a lengthy synthetic route and limited solubility in aqueous solutions. To search for spin labels with higher affinity for abasic sites and easier synthesis we have incorporated several nitroxides into the 5-position of pyrimidines. Two were made by an azide-alkyne Huisgen-Meldal-Sharpless (3+2) cycloaddition reaction (click reaction) and another two using palladium-catalyzed Sonogashira coupling. Among the click spin-labels, a U-analogue was fully bound at -30 °C while ca. 60% of a C-analogue bound to a duplex DNA containing an abasic site when placed opposite to A and G, respectively. However, 5-alkyne-linked pyrimidine spin-labeled derivatives of U and C bind only ca. 30% and 90% when paired with A and G, respectively. A C-analogue prepared by coupling 4-amino-TEMPO to 4-TPS U-derivative did not bind at all. Combined, our results indicate that stacking interactions contribute significantly to noncovalent binding of spin-labels at abasic sites in duplex DNA. Further work involving conjugation of a polyamine linker and an intercalator to the 1-position of the pyrimidines was initiated.
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