Purine-Derived Nitroxides for Noncovalent Spin-Labeling of Abasic Sites in Duplex Nucleic Acids

Abstract

A series of purine-based spin labels was prepared for noncovalent spin-labeling of abasic sites of duplex nucleic acids through hydrogen bonding to an orphan base on the opposing strand and π-stacking interactions with the flanking bases. Both 1,1,3,3-tetramethylisoindolin-2-yloxyl and 2,2,6,6-tetramethylpiperidine-1-oxyl (TEMPO) were conjugated to either the C2- or C6-position of the purines, yielding nitroxide derivatives of guanine, adenine, or 2,6-diaminopurine. The isoindoline-derived spin labels showed extensive or full binding to abasic sites in RNA duplexes, whereas the TEMPO-derived spin labels showed limited binding. An adenine-derived spin label (5) bound fully at low temperature to abasic sites in both DNA and RNA duplexes when paired with thymine and uracil, respectively, complementing the previously described guanine-derived spin label Ǵ, which binds efficiently opposite cytosine. Compound Ǵ was also shown to bind to abasic sites in DNA-RNA hybrids, either in the DNA- or the RNA-strand. Ǵ showed only a minor flanking-sequence effect upon binding to abasic sites in RNA. When the abasic site was placed close to the end of the RNA duplex, the affinity of the spin label Ǵ was reduced; full binding was observed at the fourth position from the duplex end. In summary, spin labels 5 and Ǵ showed full binding to abasic sites in both DNA and RNA duplexes and are promising spin labels for structural studies of nucleic acids by pulsed EPR methods.

Keywords: DNA/RNA; EPR spectroscopy; noncovalent interactions; spin labels; structure elucidation.