Probing specific RNA bulge conformations by modified fluorescent nucleosides

Recently, RNA bulges, three-dimensional structural motifs that function as molecular handles in RNA, were identified in various sequences and have been proposed as the interaction site for RNA-binding drugs. However direct and sensitive monitoring methods specific to RNA bulges have not been well developed. We describe here pyrene-labeled uridine derivatives which were incorporated into HIV TAR RNA bulge sites resulting in greatly enhanced fluorescence, allowing the effective monitoring of base conformations in HIV TAR RNA. To clarify this fluorescence enhancement, Hartree-Fock calculations were performed for natural and pyrene-labeled HIV TAR RNA. The calculated results showed a considerable rotation of the pyrene moiety at U25 due to argininamide-induced conformational changes in the RNA bulge. This simple and efficient method can be used to elucidate conformational changes in specific bulge regions.