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

Binding of Mg2+, Ca2+ and Co(NH3)63+ ions to the HIV‐1 TAR RNA in solution was analysed by 19F 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 19F NMR‐monitored metal ion titration. The chemical shift changes of fluorine resonances FU‐23, FU‐25 and FU‐40 upon titration with Mg2+ and Ca2+ ions indicated specific, although weak, binding at the bulge region with the dissociation constants (Kd) of 0.9 ± 0.6 and 2.7 ± 1.7 mM, respectively. Argininamide, inducing largest 19F chemical shifts changes at FU‐23, was used as a reference ligand (Kd = 0.3 ± 0.1 mM). In the Pb2+‐induced TAR RNA cleavage experiment, strong and selective cleavage of the C24‐U25 phosphodiester bond was observed, while Mg2+ and Ca2+ induced cuts at all 3‐nt residues of the bulge. The inhibition of Pb2+‐specific TAR cleavage by di‐ and trivalent metal ions revealed a binding specificity [in the order Co(NH3)63+ > Mg2+ > Ca2+] 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.