The bulge region of HIV-1 TAR RNA binds metal ions in solution

Binding of Mg(2+), Ca(2+) and Co(NH(3))(6)(3+) ions to the HIV-1 TAR RNA in solution was analysed by (19)F NMR spectroscopy, metal ion-induced RNA cleavages and Brownian dynamics (BD) simulations. Chemically synthesised 29mer oligoribonucleotides of the TAR sequence labelled with 5-fluorouridine (FU) were used for (19)F NMR-monitored metal ion titration. The chemical shift changes of fluorine resonances FU-23, FU-25 and FU-40 upon titration with Mg(2+) and Ca(2+) ions indicated specific, although weak, binding at the bulge region with the dissociation constants (K(d)) of 0.9 ± 0.6 and 2.7 ± 1.7 mM, respectively. Argininamide, inducing largest (19)F chemical shifts changes at FU-23, was used as a reference ligand (K(d) = 0.3 ± 0.1 mM). In the Pb(2+)-induced TAR RNA cleavage experiment, strong and selective cleavage of the C24-U25 phosphodiester bond was observed, while Mg(2+) and Ca(2+) induced cuts at all 3-nt residues of the bulge. The inhibition of Pb(2+)-specific TAR cleavage by di- and trivalent metal ions revealed a binding specificity [in the order Co(NH(3))(6)(3+) > Mg(2+) > Ca(2+)] at the bulge site. A BD simulation search of potential magnesium ion sites within the NMR structure of HIV-1 TAR RNA was conducted on a set of 20 conformers (PDB code 1ANR). For most cases, the bulge region was targeted by magnesium cations.