title: The bulge region of HIV-1 TAR RNA binds metal ions in solution.
abstract: Binding of Mg2+, Ca2+ and Co(NH3)6(3+) 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 (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 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)6(3+) > 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.
mesh_terms: D001120:Arginine / Q000031:analogs & derivatives* / Q000378:metabolism; D001665:Binding Sites; D001667:Binding, Competitive; D002118:Calcium / Q000378:metabolism; D003035:Cobalt / Q000378:metabolism; D005461:Fluorine; D005472:Fluorouracil / Q000737:chemistry; D015696:Gene Products, tat / Q000378:metabolism; D015497:HIV-1 / Q000235:genetics*; D006801:Humans; D008274:Magnesium / Q000378:metabolism; D009682:Magnetic Resonance Spectroscopy / Q000379:methods; D008670:Metals / Q000378:metabolism*; D009690:Nucleic Acid Conformation; D012367:RNA, Viral / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D054322:tat Gene Products, Human Immunodeficiency Virus
chemical_list: D015696:Gene Products, tat; D008670:Metals; D012367:RNA, Viral; D054322:tat Gene Products, Human Immunodeficiency Virus; D005461:Fluorine; C069339:argininamide; D003035:Cobalt; D001120:Arginine; D008274:Magnesium; D002118:Calcium; D005472:Fluorouracil